Triparna De Vreede

Imed Boughzala
email: imed.boughzala@it-sudparis.eu

Gert-Jan De Vreede

Roni Reiter-Palmon
email: rreiter-palmon@unomaha.edu

A Model and Exploratory Field Study on Team Creativity

Organizations increasingly rely on teams to solve problems creatively or design new products and services. Research to date has mostly focused on individual creativity, rather than team creativity. This paper introduces the Team Creativity Model (TCM) to understand the antecedents of team creativity. TCM posits that both individual creativity and shared mental models (SMMs) contribute to team creativity. SMMs act as a mediator between knowledge sharing and team creativity. Antecedents to individual creativity include an individual's propensity to be creative and individual knowledge. Individual knowledge also is an antecedent to knowledge sharing, as are an individual's propensity to share knowledge and trust within the team. In an exploratory study at a telecom company, a team of design experts participating in four creative sessions provided initial support for the TCM constructs and their relationships. The findings suggest that further exploratory and empirical research on TCM is justified. Some tentative implications for research and practice are presented.

Introduction

Organizations face problems today that are too complex for one individual or a mono-disciplinary team to solve [START_REF] Kozlowski | Team learning, development, and adaptation[END_REF]. Therefore, the past decade has seen an increase in the use of teams by organizations [START_REF] Galbraith | Designing matrix organizations that actually work[END_REF]. For instance, in 1999 it was reported that slightly less than 50% of organizations used teams [START_REF] Devine | Teams in organizations: Prevalence, characteristics, and effectiveness[END_REF]. Current reports suggest that this figure has jumped to over 70% of companies in 2003, and 80% in 2004 [START_REF] Edmondson | Overcoming barriers to collaboration: Psychological safety and learning in diverse teams[END_REF]. In addition, recent advances in sciences and technologies have enabled organizations to become global. Geographically dispersed locations and multidisciplinary distributed teams have become realistic work arrangements for many modern organizations.

In response to the increased focus on team work, many collaboration techniques and technology solutions have been developed to enable teams to work together productively. Team collaboration refers to the joint effort of team members in achieving a goal. The quality of collaboration in an organization directly affects the organization's outcomes and performance [START_REF] Jordan | Workgroup emotional intelligence: Scale development and relationship to team process effectiveness and goal focus[END_REF]. Team collaboration is important for team functioning when tasks are complex, ambiguous and dynamic [START_REF] Burke | Understanding team adaptation: A conceptual analysis and model[END_REF]. Collaboration is particularly beneficial when the team members are required to be open, share information with others, or be creative.

Team collaboration is especially important at the early stages of the creative process [START_REF] Hoegl | Interteam Coordination, Project Commitment, and Teamwork in Multiteam R & D Projects: A Longitudinal Study[END_REF]. Creativity is quickly becoming an essential part of non-routine project teams who are tasked with the development of innovative products or services [START_REF] Leenders | Virtuality, Communication, and New Product Team Creativity: A Social Network Perspective[END_REF]. High levels of market competition, rapid technology developments, and short product life spans, set increasingly challenging requirements to create new product specifications. Thus, team members have to display high levels of creativity in order to remain ahead of the curve [START_REF] Leenders | Virtuality, Communication, and New Product Team Creativity: A Social Network Perspective[END_REF]. Therefore, organizations are creating project based teams consisting of individuals tasked with generating innovative ideas and then transferring these ideas to create marketable technology, products, and services [START_REF] Iansiti | Technology Integration: Turning Great Research into Great Products[END_REF][START_REF] Thamhain | Managing Innovative R & D Teams[END_REF]. Research on the relationship between team collaboration, team creativity, and innovation has shown that having team goals emphasizing collaboration produces more new ideas and results in an improvement of team innovation [START_REF] Mitchell | The impact of goal structure in team knowledge creation[END_REF][START_REF] Pearce | A reciprocal and longitudinal investigation of the innovation process: The central role of shared vision in product and process innovation teams (PPITs)[END_REF].

Despite of the increasing relevance of team creativity, this area has not been extensively researched. Most of the early work on organizational creativity has focused on the individual level. Teams were viewed as a social context within which the individuals function and which could facilitate or inhibit individual creativity [START_REF] Amabile | Creativity in context: Update to the social psychology of creativity[END_REF][START_REF] Woodman | Toward a theory of organizational creativity[END_REF]. There are several reasons why the organizations focus on the role of teams in the development of creative products. First, models have been developed that suggest that some team properties can be emergent [START_REF] Kozlowski | A multilevel approach to theory and research in organizations: Contextual, temporal, and emergent processes[END_REF]. Emergence can be defined as a phenomenon that "originates in the cognition, affect, behaviors, or other characteristics of individuals, is amplified by their interactions, and manifests as a higher-level, collective phenomenon" [42, p.55]. This definition suggests that team creativity might be an entirely different process than individual creativity and should not merely be viewed as a background or a social process. Second, a focus on team creativity enables teams to capitalize on the social interactions of the team members [START_REF] Tesluk | Influences of organizational culture and climate on individual creativity[END_REF]. Project teams represent a complex social system that may involve multiple stakeholders from different parts of the organization. One of the critical aspects of innovation is the integration of the existing knowledge and ideas into an innovative business model. Studying creativity at the team level allows organizations to better understand the social interactions and allows them to exploit and integrate the expertise of the team members in order to generate more creative ideas. Finally, encouraging team creativity enables team members to share their thoughts and ideas and thereby leading to a shared understanding of the product, market, and customer requirements [START_REF] Gardner | Creativity: An interdisciplinary perspective[END_REF]. Also, team work creates opportunities for the team members to participate in group decision making and problem solving which in turn allows the team members to utilize their varied skill sets and experiences [START_REF] King | Innovation in Working Groups[END_REF].

One of the aspects of team creativity that requires attention concerns the antecedents of team creativity. While it is being accepted that team creativity is beneficial for organizations, there have not been many studies that isolate the factors that encourage or inhibit team creative outputs. It is important to understand the antecedents of team creativity as it will help organizations to create an environment and select team members that maximize the chances of innovation. Without a clear understanding of the factors affecting creativity, organizations cannot effectively utilize the expertise of their teams.

Therefore, the purpose of this paper is to explore and understand the antecedents of team creativity. We present a theoretical model, the Team Creativity Model (TCM), to explain the constructs that affect team creativity. We then evaluate this model through an exploratory field study. Through this field study we compare the constructs and relationships proposed by the TCM with the attitudes, opinions, and reported actions of 8 experts from different areas involving in four creativity sessions within an International Telecom company to come up with a new and innovative cellular phone model.

The remainder of the paper is structured as follows. In the next section we introduce the TCM, a theoretical model of the antecedents of team creativity. This is followed by the details of the field study including the participants and the process of the study. We then provide an analysis of the session transcripts where we discuss different situations and provide examples that provide support for the TCM constructs and their relationships. The paper concludes with a discussion of some tentative implications based on the model's logic, the study's limitations, and directions for future research.

A Model of Team Creativity

Creativity is often defined in terms of novelty and usefulness. A well accepted definition of creativity is "a product or response that is (a) both a novel and appropriate, useful, correct or valuable response to the task at hand, and (b) the task is heuristic rather than algorithmic in nature" [START_REF] Amabile | The social psychology of creativity[END_REF]. Creativity can exist both at the individual and team level. There appears to be consensus among researchers that when individuals work together to solve problems creatively, the creativity of the team as a whole will be influenced by the creativity of the individuals that make up the team [START_REF] Pirola-Merlo | The relationship between individual creativity and team creativity: aggregating across people and time[END_REF][START_REF] Tiwana | Expertise integration and creativity in information systems development[END_REF]. However, the extent of this influence is debated. Some researchers argue that team creativity can simply be considered as the combined creativity of the individual team members [START_REF] Pirola-Merlo | The relationship between individual creativity and team creativity: aggregating across people and time[END_REF]. Other researchers posit that the collaboration between the team members may lead to higher levels of team creativity than can be explained by the combined individual creativity levels: Combining contributions from the individual team members may lead to richer, more creative ideas at the team level [START_REF] Tagger | Individual creativity and group ability to utilize individual creative resources: A multilevel model[END_REF]. Yet, team creativity results not just from combining individual ideas. Team creativity occurs during a social process of sense-making and collaboration where one individual's actions may inspire the team to devise and follow a more creative process to address the problem at hand resulting in higher levels of creativity [START_REF] Tagger | Individual creativity and group ability to utilize individual creative resources: A multilevel model[END_REF][START_REF] Tiwana | Expertise integration and creativity in information systems development[END_REF]. The Team Creativity Model (TCM) that we introduce below adopts the second perspective, i.e. that team creativity is more than just the combined creativity of the individual team members. Following Amabile's [START_REF] Amabile | The social psychology of creativity[END_REF] definition of individual creativity, we define team creativity as the extent to which a team's ideas in response to a problem solving task are both novel and useful. TCM posits that team creativity is directly influenced by individual creativity. The extent to which individual members of a team are capable to generate creative ideas will determine the creativity of the team as a whole since the individual contributions provide the team with its 'raw' materials (see e.g. [START_REF] Santanen | Causal relationship in creative problem solving: Comparing facilitation interventions for ideation[END_REF]). Furthermore, creative individuals, experts in particular, are less likely to have difficulties or be uncomfortable expressing themselves, even under less than optimal conditions [START_REF] Collaros | Effect of perceived expertness upon creativity of members of brainstorming groups[END_REF]. Thus, if teams have higher proportion of creative individuals who are experts, such teams will be more likely to have a high degree of team creativity. However, it is important that the members of the creative team are channeled to express themselves properly and process loss is limited during their discussions as this may result in less than optimal conditions for individual team members to express themselves [START_REF] Kerr | Group performance and decision making[END_REF][START_REF] Diehl | Productivity loss in idea-generating groups: Tracking down the blocking effect[END_REF][START_REF] Nemeth | Differential contributions of majority and minority influence[END_REF].

An individual's personality or propensity to be creative is an important determinant for individual creativity. Research shows that aspects of individual ability, personality, motivational variables, and certain cognitive processes are strongly associated with individual creativity [START_REF] George | Creativity in organizations[END_REF][START_REF] Hennessey | Creativity[END_REF][START_REF] Mumford | Process analytic models of creative thought[END_REF]. Another study revealed that students who were more creative were also rated by their peers to be more sociable and popular than their non-creative counterparts [START_REF] Rivlin | Creativity and the self-attitudes and sociability of high school students[END_REF].

Along with personality and social variables that enable groups to be creative, skill sets which encourage creativity also add to the creative propensity of the individual. Amabile [START_REF] Amabile | The social psychology of creativity[END_REF] identified creativity relevant skills to include skills to break the most commonly used perceptual concepts and be able to abandon the most commonly used strategies and move in new directions. People who could look at problems with a new strategy and approached the problem with a blank canvas were better able to get creative [START_REF] Getzels | The creative vision: A longitudinal study of problem finding in art[END_REF]. Another characteristic that enabled individuals to be creative was the capability to suspend judgment and use wide categories while coming up with creative ideas [START_REF] Cropley | Creativity[END_REF][START_REF] Stein | Several findings of a transactional approach to creativity[END_REF]. Finally, individuals who remembered large amounts of information more accurately also had the capability to be more creative than their counterparts [START_REF] Campbell | Blind variation and selective retention in creative thought as in other knowledge processes[END_REF].

Creativity is also influenced by an individual's knowledge in his/her own discipline. Expertise and domain knowledge have been found to be important contributors to creativity [START_REF] Ericsson | Creative expertise as superior reproducible performance: Innovative and flexible aspects of expert performance[END_REF][START_REF] Vincent | Divergent thinking, intelligence, and[END_REF]. Complete novices tend to be less creative at first but as they gain knowledge, their creativity increases rapidly [START_REF] Stacey | Expertise and creativity in knitwear design[END_REF]. However, this relationship is not linear; it is curvilinear. To a certain point, as expertise increases, so does the individual creativity. Yet increased expertise at some level tends to close people off to options that are beyond their area of expertise and they start to depend on a broad array of conservative options that have been successful in the past [START_REF] Stacey | Expertise and creativity in knitwear design[END_REF].

Individual knowledge not only influences individual creativity, but also affects the amount of knowledge sharing that takes place in a team. Knowledge sharing occurs when group members voluntarily exchange information with the purpose of reaching a broader understanding of their group goal and its accomplishment [START_REF] Gigone | The common knowledge effect: Information sharing and group judgment[END_REF]. One of the core drivers of creativity in organizations is the knowledge-based view where project teams are used as instruments for integrating tacit and explicit knowledge among the team members [START_REF] Grant | Prospering in dynamically competitive environments: Organizational capability as knowledge integration[END_REF]. While possession of knowledge is at an individual level, it is important to integrate the knowledge of the individuals into a collective knowledge for the project to benefit from it [START_REF] Tiwana | Expertise integration and creativity in information systems development[END_REF].

Knowledge sharing is critical for creative teams. The type of knowledge that is most likely to be important for creative teams is the one that is unique to each team member based on his/her area of expertise. Groups that are able to consider more information from diverse sources are more likely to arrive at better solutions than individuals working on their own [START_REF] Gigone | The common knowledge effect: Information sharing and group judgment[END_REF]. Inadequate knowledge sharing leads to less than optimum problem solving by the group because individual group members are not able to evaluate a problem comprehensively with the limited information available to them [START_REF] Lam | Relationship between organizational justice and employee work outcomes: a cross-national study[END_REF]. In order to solve a problem most effectively, all information relevant to the problem has to be taken into account [START_REF] Lam | Relationship between organizational justice and employee work outcomes: a cross-national study[END_REF], and group members need to actively exchange information with each other in order to access that information [START_REF] Stasser | Pooling of unshared information in group decision making: Biased information sampling during discussion[END_REF][START_REF] Toma | Hidden profiles and concealed information: Strategic information sharing and use in group decision making[END_REF].

However, research indicates that the process of sharing knowledge that is not common is difficult. Group members often tend to repeat the information known to all group members as the discussion progresses and do not share information that is not widely known [START_REF] Larson | Diagnosing groups: charting the flow of information in medical decision making teams[END_REF][START_REF] Stasser | Information sampling in structured and unstructured discussions of 3-person and 6-person groups[END_REF][START_REF] Winquist | Information pooling: when it impacts group decision making[END_REF]. Moreover, it has been found that the amount of perceived expertise of the group members influences the knowledge sharing taking place in the group [START_REF] Davenport | Successful knowledge management projects[END_REF]. In other words, individuals with higher levels of expertise are more willing to share their knowledge with their team mates than their less experienced counterparts. Conversely, when individual team members do not perceive themselves to be experts in their discipline, they are less likely to offer their unique observations to the group and contribute to knowledge sharing.

Trust is an important antecedent to knowledge sharing [START_REF] Edmondson | Psychological safety and learning behavior in work teams[END_REF][START_REF] Rank | Three avenues for future research on creativity, innovation, and initiative[END_REF]. Research shows that increased trust allows team members to share knowledge more effectively [START_REF] Edmondson | Psychological safety and learning behavior in work teams[END_REF]. Trust not only influences the degree of knowledge sharing, that is, how willing team members are to share the knowledge they have, but also how the knowledge is viewed and integrated by other team members [START_REF] Abrams | Nurturing interpersonal trust in knowledge-sharing networks[END_REF]. Lack of trust is sometimes an issue with because of such team's diverse makeup. As a result these teams may suffer from less than optimal communication and interactions.

In addition to trust, individuals must have a propensity to share knowledge in order to make the knowledge sharing a part of the team routine. But, there are various factors that may affect this desire to share information. Information is shared differently among group members who are familiar with each other than those who are not, which in turn affects the problem solving capabilities of the group [START_REF] Gruenfeld | Group composition and decision making: How member familiarity and information distribution affect process and performance[END_REF]. There are also certain social dilemmas associated with knowledge sharing which might impact the propensity to share knowledge. For example, if some of the group members have spent considerable time and resources to gather the knowledge, they might be unwilling to share that knowledge in a group setting [START_REF] Cabrera | Knowledge-Sharing Dilemmas[END_REF]. In particular when it is unclear what they can gain in exchange [START_REF] Boughzala | Knowledge sharability in cross-organizational collaboration: an exploratory field study, 44 th Hawaii International Conference on System Sciences[END_REF].

TCM posits that knowledge sharing does not contribute to team creativity directly but it contributes to the development of Shared Mental Models (SMMs), which in turn influences team creativity. SMMs are representations of knowledge structures that are shared among team members [START_REF] Cannon-Bowers | Shared mental models in expert team decision making[END_REF][START_REF] Klimoski | Team mental model: Construct or metaphor[END_REF]. These shared structures enable the team members to find a common ground to describe, explain, and predict the events that occur in their environment [START_REF] Mathieu | The influence of shared mental models on team processes and performance[END_REF]. SMMs can be divided primarily into four areas: (a) knowledge about equipment and tools; (b) knowledge about the team task, goal, and performance requirements; (c) knowledge about other team members' abilities, knowledge, and skills; and (d) knowledge about appropriate team interactions [START_REF] Cannon-Bowers | Shared mental models in expert team decision making[END_REF]. The majority of research on SMMs has focused on one of the above dimensions or has consolidated them into two dimensions, namely, task and team shared mental models.

Even though research on SMMs and team creativity is limited, existing empirical studies suggest that SMMs have a positive relationship with creativity both at individual and team level. For example, Mumford et al. [START_REF] Kozlowski | Team learning, development, and adaptation[END_REF] found that when shared mental models are developed through sharing the same training program, they facilitate higher levels of creative idea generation within the team. Similarly, Pearce and Ensley [START_REF] Pearce | A reciprocal and longitudinal investigation of the innovation process: The central role of shared vision in product and process innovation teams (PPITs)[END_REF] found shared vision to be positively related to product and process innovation ideas in teams. In a related study, Gilson and Shalley [START_REF] Gilson | A little creativity goes a long way: An examination of teams' engagement in creative processes[END_REF] found that increasing the number of shared goals in a team not only resulted in an increase in the creativity but also increased the engagement of the team members in the creative process. However, SMMs are not static. They may not initially be present in a group (see e.g. [START_REF] Majchrzak | Coordinating Expertise Among Emergent Groups Responding to Disasters[END_REF]) and they are developed over time through discussion of issues, sharing knowledge, and learning from past mistakes and successes [START_REF] Burke | Understanding team adaptation: A conceptual analysis and model[END_REF][START_REF] Marks | The impact of cross-training on team effectiveness[END_REF][START_REF] West | Innovation in top management teams[END_REF]. Based on the above discussion, figure 1 depicts the various antecedents and their relationships proposed by the TCM. The next section introduces an exploratory field study with experts from an international telecom company in France that was executed to find support for the TCM constructs and their relationships.

Method

Participants

Eight experts from a telecom company participated in each of four creativity sessions (Table 1). The involvement of industry experts in these sessions [START_REF] Galbraith | Designing matrix organizations that actually work[END_REF] allowed us, in the words of Hevner et al. [START_REF] Edmondson | Psychological safety and learning behavior in work teams[END_REF] p. 80) "to combine relevance and rigor by meeting a business need with applicable knowledge." Each participant held a master degree and three had earned PhDs. Their degrees were in a variety of disciplines: Industrial Design, Engineering, Management, Marketing, Sociology, Computer Science, and Telecom. Participants worked for different departments, including Strategic Marketing, Telecom Solutions, Sales, R & D, Quality, and IT. Their work experience ranged from 10 to 24 years. Their average age was 42 and 62% were male. The participant names are withheld for privacy reasons. Their statements are identified only by their ID and their department. The name of the company and the sessions' factual deliverables are confidential.

Procedures

Participants worked together during four sessions over a four-month period between October 2009 and January 2010 to co-design an innovative cellular phone model. Each session lasted three hours and was facilitated by one of the authors. All sessions were conducted in French. Graduate assistants recorded the sessions and made field notes about critical incidents and verbal statements. Participants recorded their key contributions on Post-It notes, and gave them to the research team at the end of each session. The facilitator made field notes immediately following every session. At the end of each session, participants were asked to provide their personal perceptions about the team creativity process and the intermediate and final deliverables (the cellular phone model).

To minimize the possibility of observation bias, we did not seed the participants with direct questions pertaining to the TCM constructs. Rather, we observed interactions among participants and documented their feedback. The session observers were not briefed on the TCM constructs until after the sessions were over.

The Creativity Method

The sessions used a specific creativity method, the Concept-Knowledge (C-K) method [START_REF] Hatchuel | C-K theory: Notions and applications of a unified design theory[END_REF] and a structured facilitation process. The C-K method helps individuals and teams to innovate by constraining them to get out of the box when they aim to create a new solution, product, or service. The method is based on the concept of "expandable rationality" [START_REF] Hatchuel | Studying creative design: The contribution of C-K theory. Studying Design Creativity[END_REF][START_REF] Hatchuel | C-K theory: Notions and applications of a unified design theory[END_REF]. It allows training people to avoid design illusions, memorizing the history of a design process, and structuring team work in innovative design projects. The participants were not familiar with this type of creativity methods.

The C-K method views expansion as a key focus in a design project. There are two expandable spaces: The Knowledge space (K), consisting of logical propositions relevant to the design project. These propositions are used to formulate new concepts or expand existing concepts. The Concept space (C), consisting of objects that meet some desired design requirements that do not exist yet. These objects typically represent the goal of design activities. The C-K design process lets the C and K spaces co-evolve by (1) allowing concepts in the C-space to expand with new properties based on propositions from the K space, and (2) expanding knowledge in the K-space through new insights from the experiences with the concepts that result from the design process. For further details on the C-K method, please refer to [START_REF] Gillier | Managing innovation fields in a cross-industry exploratory partnership with C-K design theory[END_REF][START_REF] Hatchuel | C-K theory: Notions and applications of a unified design theory[END_REF].

Creativity Sessions

The main goal of the four creativity sessions was to "imagine" and design a new cell phone model without a SIM card (based on the Cloud Computing developments). Each creativity session focused on one step of the C-K method. First, a cell phone was initially defined to consist of the following components: Screen, Keyboard, Microphone, Earphone, Transceiver/antenna, SIM card, Operating System, Services/Apps, and Camera. Second, the SIM card concept was removed. The participants had to try to expand the other components such that they would fulfill the requirements of the new phone model. Finally, the participants had to integrate the expanded components such that they would envision a new cell phone model that still offered the same functionality of a traditional cell phone.

Each session was organized at a different department. Tables and chairs were arranged in a U shape to facilitate eye contact between the participants. Each session opened with a brief presentation by the facilitator to introduce the session agenda and to summarize progress so far in process. Participant also received guidelines on how to share their ideas by writing them on Post-It notes and on justifying their ideas with precise arguments and/or concrete examples. Participants then generated, clarified, reduced, organized, and evaluated ideas concerning the new cell phone design. At the end of each session, participants were asked to provide feedback about the team work and the execution of the C-K method.

Specific details on each of the four sessions are provided in table 2. The final deliverable from the four sessions was a specification report for the new cellular phone model.

Analysis

The facilitator and graduate assistants transcribed the sessions' Post-It notes and field notes into a collection of utterances identified by contributor. They then anonymized this data by replacing the contributors' names with their respective ID. The facilitator then coded the utterances according to the TCM constructs. The utterances were also translated into English and validated by someone fluent in both English and French. An English-speaking researcher then also coded the utterances according to the TCM constructs. There was substantial concurrence between the French and English coding. The few discrepancies were resolved through discussion.

Results

This section presents an analysis of the session transcripts in light of the constructs and relationships proposed by TCM. Many of the utterances from the participants about propensity to share, propensity to create, knowledge sharing, shared mental model, individual and team creativity were consistent with the constructs and relationships proposed by TCM. This section presents qualitative support for that assertion.

Antecedents to knowledge sharing

Propensity to share knowledge, trust within team, and individual knowledge are the three antecedents of knowledge sharing as identified by the TCM. The presence of these antecedents was examined in the sessions with the experts. Results show support for the role of trust in sharing knowledge. It was obvious in most sessions that trust played a significant role in establishing an environment where the experts could share their knowledge easily. Many experts commented to that effect. For example, one of them said "The fact is that we belong to the same company helps to build mutual trust… sharing was almost natural". Another expert from the Quality department mentioned, "An innovation process goes smoothly and quickly when people are in good agreement, have trust in each other." However, initial levels of trust may have been relatively lower. The team appeared to have built trust over time and they seemed to be cognizant of the changes that took place in the team. Said one of the experts from the telecom department "Gradually,

as the work progresses during the sessions, a team spirit was established and trust took place between us from different departments."

There was also support that a propensity to share knowledge is required for knowledge sharing. This was evidenced in the following comment "Creativity may seem like an individual concern but eventually it's well done in teams if people are willing to share/combine their knowledge and expertise." Interestingly, the experts mostly commented on the propensity to share knowledge in combination with the importance of possessing individual knowledge. For example, one of the experts mentioned, "It is not only the matter of willingness to share…but also having something to share and being able to do it". This sentiment was echoed in another session where it was said that "The willingness to share comes first and you must have knowledge and be able to share it ... the method helps a lot for that".

A number of experts also appeared to suggest an association between individual knowledge and knowledge sharing. Experts felt that team creativity requires knowledge sharing when people combine their individual knowledge: "In our business, creativity is no longer an individual concern but the collaboration/pooling of several specialties and skills." As evidenced by a comment from an expert from the IT Department, for knowledge sharing to take place, it does require the presence of individual knowledge: "…having something to share to be able to share." In fact, some experts believed that individual knowledge would only become valuable in a team project if it is shared. Said one expert from the R & D Department, "Knowledge is the only resource which has more value when you share it".

Antecedents to individual creativity

Propensity to be creative and individual knowledge were also assessed in the sessions as antecedents to individual creativity. We found support for the propensity to be creative and moderate support for individual knowledge being an antecedent for creativity at an individual level. While talking about creativity, one of the experts mentioned, "To get out of the box, one must be ready for that" meaning that they should also have the attitude to be creative. Another expert from a different department had something similar to say about his department, "We can say that people were willing and predisposed to create an innovative solution". The experts not only mentioned the predisposition to be necessary, they also noticed the importance of propensity to be creative after the sessions were over: "This is a difficult but exciting exercise ... when we see the outcome, it's impressive for a short time ... eventually one have to be predisposed to it." Support for individual knowledge as an antecedent to individual creativity came from comments such as "In this creativity session, trust and challenge were always our Leitmotiv [i.e. guiding theme] ...we also should have the good knowledge to share". They also commented on the selection of the experts as they contributed to the knowledge pool. For example, one of the experts said, "The choice of experts was important ...".

Individual creativity as an antecedent to team creativity

Individual creativity was named as one of the antecedents to team creativity in the TCM. This was illustrated on several occasions during the creativity sessions providing support to the model. One of the experts from the sales department mentioned that "1+1=3… it's more than the sum of everyone's knowledge" to point out the fact that the combination of individual creativity produced a synergy when working in teams. The experts from different sessions were also in agreement that creativity has become more than an individual endeavor. Comments like "In our business, creativity is no longer an individual concern but the collaboration/pooling of several specialties and skills" illustrate this fact. Some experts also mentioned their support of combining individual creativity to achieve a team outcome: "Creating is always easier collectively."

The participants also seemed to be impressed with the amount of creative ideas that were generated by the use of experts from different departments. For example, one of the participants mentioned "The choice of experts was important ... I believe that they are all creative and sensitive to knowledge sharing and creativity". Other comments that were received in support of this attitude were "I'm not sure we could be able to achieve the same result with a single point of view, in the same department ... especially in such a short time" and "The fact that the group is mixed... Views are complementary and rich that helps to innovate", and "The team was balanced ... each of us brought a stone to the edifice".

Knowledge sharing and shared mental models as antecedents to team creativity

Knowledge sharing and shared mental models are the antecedents to team creativity in the TCM. Shared mental models directly influence team creativity, while the shared knowledge influences team creativity through shared mental models. We found support for both relationships. The experts involved in the creativity sessions acknowledged the importance of knowledge sharing in the creation of an SMM. To this effect, one of the experts commented "This (creativity sessions) strengthens our interest to sharing our knowledge capital and create mental schemas". Another expert mentioned "I did not think that people would be able to share their knowledge so easily and with a goodwill... converging their so different points of view". The participants were aware of the process that took place in order to create a shared mental model. They expressed sentiments like "In the beginning, there was some uncertainty in the project... but once we have advanced in the process of the method, there was a reconciliation (shared understanding) ... the method proved useful and credible for everybody". They were also impressed by the effect of knowledge sharing on team creativity. Said one expert, "... sharing our knowledge and knowhow was the only way for us to be able to create together." One of the experts expressed his surprise at the degree of creativity resulting from the knowledge sharing exercise by saying "I've never imagined the usefulness of this method by constraining us to surpass and force ourselves to be creative together despite our differences".

The experts also acknowledged the importance of shared mental models in the team creativity outputs. Their understanding of the influence of shared mental models on team creativity was reflected by comments such as "Innovation process goes smoothly and quickly when people … have the same aspirations and perceptions" and "In this exercise, everyone must be on the same wavelength ("the same tempo")". The experts even accepted and welcomed the challenges of working with people from different departments so that they could benefit from the rewards of team creativity once a shared mental model was established. Comments like "We do not have the same vocabulary, the same concerns and the same points of view… but we were able to work together and innovate" and "The time we spent together was very challenging to overcome our differences ... yet it was the source of our strength to be able to create, illustrate that point.

Discussion and conclusions

This paper advances the Team Creativity Model (TCM) as an explanation to understand the antecedents of team creativity. TCM posits that both individual creativity and SMMs contribute to team creativity. SMMs acts as a mediator between knowledge sharing and team creativity. Antecedents to individual creativity include an individual's propensity to be creative and individual knowledge. Individual knowledge also is an antecedent to knowledge sharing, as are an individual's propensity to share knowledge and trust within the team. During an exploratory study in a telecom company, experts that participated in four creative design sessions provided initial support for the TCM constructs and their relationships. These findings suggest that further exploratory research is merited. They also allow us to tentatively present a number of implications for research and practice.

TCM is based on a synthesis of research efforts on various aspects of individual and team creativity. The model and related field data suggest that studying team creativity requires careful consideration of a complex set of constructs and relationships. To empirically study team creativity, we need to consider not only individual creativity, but also important antecedents to individual creativity and the development of SMMs during the creative team process. Further, it can be argued that a deeper understanding of nature of relationships between constructs required. For example, what is the exact nature of the relationship between SMMs and team creativity? Do SMMs just lead to higher idea generation productivity in teams? Or, do SMMs just facilitate the selection of the most creative ideas? Or do they support both idea generation and selection? In fact, it has been suggested that SMMs may lead to too much similarity and therefore conformity [START_REF] Cannon-Bowers | Shared mental models in expert team decision making[END_REF]. It is possible that some degree of dissimilarity in SMMs or less sharedness would be beneficial especially for team creativity and innovation. The same questions can be raised with respect to knowledge sharing: What is the optimal level of knowledge sharing to support the formation of SMMs?

Our model and findings also provide insights for organizations that employ teams in creative problem solving activities. In particular, it can be argued that if the logic of TCM holds, then organizations must investigate strategies to increase the formation of SMMs and the amount of knowledge that is shared. Several such strategies have been suggested. For example, discussion and planning have been proposed as a strategy to facilitate the formation of SMM. Through discussion and planning, team members become aware of more aspects on how the creative task has to be executed, the role of different members, and possible difficulties each person may need to address. Indeed, Stout, Cannon-Bowers, Salas, and Milanovich [START_REF] Stout | Planning shared mental models, and coordinated performance: An empirical link is established[END_REF] have found that planning is a way to develop shared mental models that facilitate team performance. Organizations can also structure interventions to bring about social dynamics that increase the level of knowledge sharing [START_REF] Cabrera | Knowledge-Sharing Dilemmas[END_REF]. One such intervention would be to restructure group rewards such that there are greater payoffs for contributing. Another intervention can be to increase the sense of group efficacy as well as the perception of the impact that individuals believe their contributions have on the group. To illustrate this point, Kerr [START_REF] Kerr | Group decision making at a multialternative task: Extremity, interaction distance, pluralities, and issue importance[END_REF] showed that the knowledge sharing in a group reduced when the group size increased and group member's perception of their contribution decreased. A final intervention focuses on establishing a sense of group identity through increased communication among team members [START_REF] Dawes | Social Dilemmas, economic selfinterest, and evolutionary theory[END_REF]. When individuals feel that they are part of a group, they are more likely to share information [START_REF] Bonacich | Communication networks and collective action[END_REF].

The following limitations have to be taken into account when interpreting the results of this study. First, the support for the TCM proposed in this paper originated from limited field observations and expert feedback from a single organization. While no data appeared to challenge the TCM or hint at other constructs, further data collection from organizations in different sectors or from cross-organizational collaborations is required to more broadly examine the extent to which the constructs of TCM manifest in the field. Such data collection should also focus on the development of constructs over time, e.g. adjustments in participants' SMMs. Second, although the field data offers support that the TCM constructs are related, the nature of exploratory study is such that it cannot assert causality. Therefore, further theoretical research and empirical studies are required to find support for causal relationships between TCM constructs. Finally, it was not possible through this field study to actually measure the individual TCM constructs, in particular individual creativity and team creativity. Such measurements would have been useful to gain a deeper understanding of the relationship between individual and team creativity as well as between other TCM constructs.

We envision a number of avenues for further research. First, given the findings reported in this paper, it will be necessary to explore the TCM more fully through additional qualitative studies with different creative problem solving or design teams. Second, some initial confirmatory quantitative studies can also be designed to use Structural Equation Modeling [START_REF] Bollen | Structural Equations with Latent Variables[END_REF] to investigate the relationships between TCM constructs and the mediation relationship between knowledge sharing and SMMs. Finally, it would be useful to examine the fundamental assumption that give rise to the development of the TCM, i.e. that team creativity ultimately leads to higher innovation quality.

Figure 1 .

 1 Figure 1. The Team Creativity Model.

Table 1 :

 1 Participants' demographic data.

	ID Area	Dept.	Age Sex Background Degree Work
						exp.
	E1 Marketing Strategic	35-39 F	Marketing &	MBA 10-14
		Marketing		Sociology
	E2 Interface	R & D	40-44 M Industrial	MSc 15-19
	Design			Design
	E3 Electronic	Telecom	40-44 M Engineering MSc 15-19
	engineering	Solutions
	E4 Telecom	Telecom	40-44 M Telecoms	PhD	15-19
	Engineering	Solutions
	E5 Software	IT	40-44 M Comp.Sci. PhD	15-19
	E6 Network	IT	40-44 M Telecoms	MSc 15-19
	architecture
	E7 Quality of	Quality	40-44 F	Telecoms &	PhD	15-19
	Service			Ergonomics
	E8 Sales and	Sales	45-49 F	Management MBA 20-24
	CRM

Table 2 .

 2 Details of the creativity sessions.

	Session Focus
	1	Recent cell phone trends and evolutions in
		telecom industry. Key cell phone adoption and
		marketing drivers.
	2	Removal of SIM card concept. Expansion of
		remaining concepts. Technical feasibilities
		and constraints related to design, cost, legal,
		health, and market concerns.
	3	Development of solutions and scenarios based
		on technical possibilities, market situations,
		and competitors.
	4	Evaluation of previously formulated solutions
		and scenarios. Selection of final solution.
		Project debrief.


Takefumi Negishi

Hitoyoshi Yasuo
email: yasuo@obs-vlfr.fr

Distinct modes of mitotic spindle orientation align cells in the dorsal midline of ascidian embryos

The orientation of cell division can have important consequences on the choice of cell fates adopted by each daughter cell as well as on the architecture of the tissue within which the dividing cell resides.

We have studied in detail the oriented cell divisions that take place in the dorsal midline of the ascidian embryo. The dorsal midline cells of the ascidian embryo emerge following an asymmetric cell division oriented along the animal-vegetal (A-V) axis. This division generates the NN (Notochord-Neural) cell at the margin and the E (Endoderm) cell more vegetally. Deviating from the default mode of cell division, these sister cells divide again along the A-V axis to generate a column of four cells. We describe these cell divisions in detail. We show that the NN cell mitotic spindle rotates 90° to align along the A-V axis while the E cell spindle forms directly along the axis following the asymmetric migration of its centrosomes. We combine live imaging, embryo manipulations and pharmacological modulation of cytoskeletal elements to address the mechanisms underlying these distinct subcellular behaviours. Our evidence suggests that, in E cells, aster asymmetry together with the E cell shape contribute to the asymmetric centrosome migration. In NN cells, an intrinsic cytoplasmic polarisation of the cell results in the accumulation of dynein to the animal pole side. Our data support a model in which a dynein-dependent directional cytoplasmic pulling force may be responsible for the NN cell spindle rotation.

Introduction

The orientation of the cell division axis has important consequences for tissue architecture and cell fate. The cell division axis is defined by the orientation of the mitotic spindle, with the cleavage furrow always forming at right angles to the spindle (reviewed in [START_REF] Conrad | Mechanism of cytokinesis in animal cells[END_REF].

According to Sachs's rule, which was originally proposed based on his studies on plant cells, each new cell division plane intersects the preceding plane at a right angle so that the "default" orientation of a spindle would be 90° to the previous spindle (reviewed in [START_REF] Wilson | Growth, cell division and development[END_REF][START_REF] Strome | Determination of cleavage planes[END_REF]. The 90° default orientation can be explained by the movements of the duplicated centrosomes, which migrate 90° in opposite directions around the nucleus and serve as the asters of the next mitotic spindle.

Hertwig formulated another important rule stating that spindles align with longest axis of the cell, such that spindle orientation can be controlled by cell shape. [START_REF] Minc | Influence of cell geometry on division-plane positioning[END_REF] further refined Hertwig's rule by proposing a computational model that fully predicts the orientation of a mitotic spindle of sea urchin eggs imposed into various shapes. The key element of this computational model is the assumption that a cytoplasmic pulling force generated on the spindle pole by each microtubule is proportional to its length, as initially proposed by [START_REF] Hamaguchi | Analysis of the role of astral rays in pronuclear migration in sand dollar eggs by the colcemid-UV method[END_REF]. However, in vivo evidence for the presence of such a pulling force in the context of cell shape-dependent spindle orientation is lacking. Most of our current knowledge on the molecular control of spindle orientation in metazoans is based on asymmetric cell divisions of highly polarised cells such as the Drosophila neuroblasts (NBs) and sensory organ precursors (SOPs) and the C. elegans zygote (reviewed in [START_REF] Morin | Mitotic spindle orientation in asymmetric and symmetric cell divisions during animal development[END_REF]. In these systems, distinct polarising cues converge on cortical NuMA proteins.

The NuMA family of proteins regulate the activity of the dynein/dynactin motor complex to exert a pulling force on astral microtubules from the cell cortex [START_REF] Morin | Mitotic spindle orientation in asymmetric and symmetric cell divisions during animal development[END_REF]. Importantly, the cortical NuMA-based mechanism is likely to be evolutionary-conserved since it also operates in chick neuroepithelium [START_REF] Peyre | A lateral belt of cortical LGN and NuMA guides mitotic spindle movements and planar division in neuroepithelial cells[END_REF], mouse embryonic skin progenitors [START_REF] Lechler | Asymmetric cell divisions promote stratification and differentiation of mammalian skin[END_REF] and mammalian cell culture systems [START_REF] Zheng | LGN regulates mitotic spindle orientation during epithelial morphogenesis[END_REF][START_REF] Kiyomitsu | Chromosome-and spindle-pole-derived signals generate an intrinsic code for spindle position and orientation[END_REF]. However, it is not yet clear whether the NuMA-based system is applicable to all oriented cell divisions.

The embryogenesis of many invertebrates proceeds with an invariant cell division pattern [START_REF] Wilson | Growth, cell division and development[END_REF][START_REF] Freeman | The role of egg and organization in the generation of cleavage patterns[END_REF]). An invariant cell division pattern implies a precise control of cell division orientation and timing. In many animal groups, the mechanisms controlling these precise cell division patterns have not been addressed. In this study, we use ascidian embryos, which develop with an invariant bilateral holoblastic cleavage pattern [START_REF] Conklin | The organization and cell lineage of the ascidian egg[END_REF], as a model to explore oriented cell divisions. Ascidians are invertebrate chordates of the phylum Tunicata, a sister group of the vertebrates [START_REF] Delsuc | Tunicates and not cephalochordates are the closest living relatives of vertebrates[END_REF]; Vienne and Pantarotti, 2006;[START_REF] Satoh | Chordate evolution and the three-phylum system[END_REF]. The blastula fate maps of ascidians and vertebrates reveal a high degree of topological similarity and ultimately give rise to tadpole larvae with a central notochord and dorsal hollow nervous system [START_REF] Lemaire | Ascidians and the plasticity of the chordate developmental program[END_REF]. In this study, we focus on two oriented cell divisions that take place in the dorsal midline of ascidian blastula stage embryos (Fig. 1A). These oriented cell divisions occur in a sister cell pair, named NN (Notochord/Neural) and E (Endoderm) cells. Their mother cell, the NNE cell (Notochord/Neural/Endoderm), divides along the animal-vegetal (A-V) axis to generate one NN cell and one E cell. Contrary to Sachs's rule, these sister cells then divide again along the A-V axis to form a column of four cells on the dorsal side of the 44-cell "blastula" stage embryo (Fig. 1A). The molecular mechanisms that determine the differential fates of daughter cells following each of these asymmetric divisions are well documented (Hudson et

Results

Oriented cell divisions of the medial NN and E cells

All experiments described in this study were conducted using the ascidian, Phallusia mammillata.

Phallusia embryos are transparent and injected mRNA is efficiently translated in eggs, thus enabling the visualisation of tagged proteins and subcellular structures in living eggs and embryos [START_REF] Prodon | Dual mechanism controls asymmetric spindle position in ascidian germ cell precursors[END_REF]. Importantly, all cell divisions up to the gastrula stage take place within the planar plane. As described above, contrary to Sachs's rule, NN and E cells both divide along the A-V axis as their mother cell (NNE) did. We addressed whether Hertwig's rule could instead be implicated in the NN and E cell divisions. To do this, we conducted live-imaging of embryos in which microtubules and cell membranes were labelled with ensconsin fused with three GFPs in tandem (ensconsin::3xGFP) (von [START_REF] Von Dassow | Action at a distance during cytokinesis[END_REF][START_REF] Negishi | Practical tips for imaging ascidian embryos[END_REF]) and FM4-64 [START_REF] Prodon | Dual mechanism controls asymmetric spindle position in ascidian germ cell precursors[END_REF], respectively. NN and E cells are quadrilateral (=tetragon) with one side corresponding to the embryonic midline (Fig. 1B). To describe length, we measured the line that is parallel to the embryonic midline and crosses the centre of the nucleus or of the spindle (Fig. 1B). For the width, the line that is perpendicular to the length and crosses the centre of the nucleus or spindle was measured (Fig. 1B). During prophase and prometaphase, the length/width ratio of NN cells remains below 1.0, but increases during metaphase to reach 1.3 ± 0.16 by anaphase (n=7 cells) (Fig. 1C, red line). The E cell on the other hand is elongated throughout the entire cell cycle (Fig. 1C, blue line). Thus, for both NN and E cells, the cell division axis corresponds to the final long axis of the cell.

While both NN and E cells divide along the long axis of the cell, our observations of centrosome and spindle dynamics revealed that their spindles align via distinct mechanisms (Fig. 1D; Movie 1). In the NN cell, the duplicated centrosomes are located on the animal pole side of the cell.

During prophase, the centrosomes migrate 90° in opposite directions within the plane of the apical contour (Fig. 1D). During metaphase, the mitotic spindle undergoes a 90° rotation to align along the A-V axis. This rotation can also be observed with DIC microscopy (Movie 2). From spindle formation to A-V alignment takes 7.2 (±1.1) minutes (n=12). The direction of spindle rotation is not biased with both clockwise and counter-clockwise directions observed equally (n=54) (Fig. 2A). However, within individual embryos, the mitotic spindles of NN cell pairs tend to rotate with bilateral symmetry (72.7%) (Fig. 2A). The E cell exhibits a different sequence of centrosome/aster movements. In E cells, following centrosome duplication, one centrosome becomes preferentially migratory and moves approximately 180° around the nuclear envelope such that when the mitotic spindle forms it is directly aligned with the A-V axis (Fig. 1D; Movie 1). From centrosome duplication to A-V spindle formation takes 14.4 (±2.5) minutes (n=18).

In summary, the spindles in NN and E cells align along the long axis of the cells by distinct mechanisms. In the following sections, we describe in further detail, first, the centrosome positioning mechanism in E cells and, subsequently, the spindle rotation in NN cells.

Correlation between asymmetric centrosome migration of the E cell and cell shape

Differential migratory behaviour of duplicated centrosomes has been reported to underlie the oriented cell divisions of Drosophila stem cells, namely neuroblasts and male germ line stem cells [START_REF] Rebollo | Spindle alignment is achieved without rotation after the first cell cycle in Drosophila embryonic neuroblasts[END_REF][START_REF] Yamashita | Orientation of asymmetric stem cell division by the APC tumor suppressor and centrosome[END_REF]. In these stem cell systems, the duplicated centrosomes exhibit differential microtubule organizing (MTOC) activities with the immobile centrosome showing a more robust astral microtubule array [START_REF] Rebollo | Spindle alignment is achieved without rotation after the first cell cycle in Drosophila embryonic neuroblasts[END_REF][START_REF] Yamashita | Asymmetric inheritance of mother versus daughter centrosome in stem cell division[END_REF]. When the centrosome of the E cell first duplicates, the duplicated centrosomes align along the A-V axis and, in a majority of cases (89.5%), the centrosome proximal to the nucleus starts to migrate away from its distallypositioned counterpart (Figs. 1D and2C,D; Movie 1). The motile centrosome migrates around the nucleus, preferentially taking the route facing to the embryonic midline (88.5%) (Figs. 1D and2C,D; Movie 1). Finally, we observed a difference in size between proximal and distal asters. Using images of embryos immunostained for -tubulin, we counted the number of pixels in the proximal and distal asters of E cells and found that the distal aster was larger in size to the proximal aster (in 80.0% of cases) (Fig. 2B,D,E). This could indicate an asymmetry in MTOC activity or differential microtubule stability between the two asters.

The degree of asymmetry in E cell centrosome migration appeared variable (Fig. 3A). To express this variation quantitatively, we measured the angle between the position of the newly duplicated centrosomes and the position of the distal-most centrosome when migration was complete (centrosomes 180° apart) (Fig. 3C). We refer to this angle as the centrosome migration angle (θ) (Fig. 3C). In 10/18 of E cells, the centrosome migration angle falls between 150° and 180°, which we classed as 'asymmetric' (Fig. 3A,D). However, the remaining E cells analysed exhibited either 'moderate asymmetry' (120°<θ<150°; 4/18) or 'symmetry' (90°<θ<120°; 4/18) in their centrosome migration. Even in the cases where centrosome migration was classed as 'moderate asymmetry' or 'symmetry', the spindle ultimately aligns with the A-V axis (Fig. 3A). This adjustment takes place either via rotation of the centrosome-nucleus complex or via spindle rotation.

We first addressed whether the E cell asymmetric centrosome migration was an intrinsic property of E cells. Individual E cells were isolated at the 24-cell stage from embryos expressing ensconsin::3xGFP to observe centrosome dynamics. In isolated E cells, we did not observe any centrosome migration classed as 'asymmetric' (0/9), suggesting that contact with other cells may be required for asymmetric centrosome migration (Fig. 3B,D). In Drosophila male germ line stem cells (GSCs) that exhibit a similar asymmetric centrosome migration, an adherence junction-mediated contact site with a stem cell niche defines the position of the immotile centrosome [START_REF] Yamashita | Orientation of asymmetric stem cell division by the APC tumor suppressor and centrosome[END_REF]. The symmetrical centrosome migration observed in isolated E cells might thus be due to a loss of contact with specific neighbouring cells. The B6.1 cell would be a good candidate for a cell that provides such a contact site since it is positioned on the vegetal side of the E cell, where the immotile centrosome is predominantly observed (Fig. 3B, middle panels). In order to test whether the asymmetric centrosome migration requires contact with B6.1, we ablated one B6.1 cell from early 24cell stage embryos and observed centrosome dynamics (ensconsin::3xGFP) in the E cell pair (Fig. 3B,D). To our surprise, in the E cell that no longer had contact with B6.1, asymmetric centrosome migration was not lost, with 6/8 cells classed as asymmetric (green cell in Fig. 3B,D). On the other hand, in the E cell on the non-ablated (=contralateral) side, that is, in the E cell that retained contact with B6.1, the frequency of asymmetric centrosome behaviour appeared somewhat reduced with 2/8 E cells classed as asymmetric (Fig. 3D). Thus, contact with B6.1 is unlikely to play a critical role in the asymmetric centrosome behaviour of a neighbouring E cell. Ablation of an E cell also appeared to affect the centrosome behaviour of its contralateral neighbour. When one E cell at the early 24-cell stage was ablated, the frequency of asymmetric centrosome behaviour in the contralateral E cell appeared to decrease, with no migrations classed as asymmetric (0/7) (Fig. 3B,D). Altogether, we found no evidence to suggest that the asymmetric centrosome migration of the E cell was under the influence of a specific neighbouring cell. Rather, we suppose that ablation of neighbouring cells altered the E cell shape and that this change in E cell shape may be affecting the behaviour of the centrosomes. In order to test this possibility, we first measured the width of each E cell analysed in Fig 3D and found that indeed, the cell ablations had an impact on the shape of E cells (Fig. 3E). Finally, we tested if there was a correlation between cell shape and centrosome migration angle (Fig. 3F). We found a moderate correlation between the centrosome migration angle and the width of the E cell, which was supported by statistical analysis (r=-0.63, p<0.0001). Thus, the narrower the E cell shape becomes, the more asymmetrically the two centrosomes tend to migrate.

We conclude that the asymmetric centrosome migration of E cells is influenced by cell shape.

In the following sections, we analyse in detail the NN cell division.

NN cell spindle rotation requires both microtubules and actomyosin

The NN cell division is oriented by rotation of the mitotic spindle. In other systems, astral microtubules play a central role in controlling spindle orientations relative to cell shape and cortical cues (reviewed in [START_REF] Théry | Cell shape and cell division[END_REF][START_REF] Morin | Mitotic spindle orientation in asymmetric and symmetric cell divisions during animal development[END_REF]. Microfilaments (F-actin) are also implicated in spindle orientation and are thought to participate in the establishment of cellular polarity (reviewed in Nance and Zallen, 2011). In addition, F-actin may itself be an integral component of the cortical cues required to orient the spindle [START_REF] Théry | The extracellular matrix guides the orientation of the cell division axis[END_REF][START_REF] Castanon | Anthrax toxin receptor 2a controls mitotic spindle positioning[END_REF].

Our data show similarly that the NN cell spindle rotation is dependent on these cytoskeletal elements.

The role of microtubules was addressed by observing ensconsin::3xGFP labelled spindles in embryos treated with a low dose of nocodazole (5 nM) from the beginning of the NN cell cycle. This dose was sufficient to reduce the size of asters but did not destroy the spindles, although spindles appeared somewhat smaller in size. This treatment resulted in inhibition of NN spindle rotation (Fig. 4).

Similarly, treatment of embryos either with cytochalasin B, an inhibitor of actin filament polymerization, or blebbistatin, an inhibitor of myosin II, prevented spindle rotation (Fig. 4). Thus, the spindle rotation of NN cells is dependent on an intact microtubule and actomyosin system, although these experiments did not allow us to discriminate whether the effects are cell autonomous or non-cell autonomous.

Cytoplasmic asymmetries associated with spindle rotation in NN cells

During the two successive cell divisions of the NNE and NN cells in Phallusia mammillata, mitochondria are differentially segregated, such that they accumulate in the animal pole side (marginal) cell [START_REF] Zalokar | Tracing of cell lineage in embryonic development of Phallusia mammillata (Ascidia) by vital staining of mitochondria[END_REF]. We addressed when this cytoplasmic polarisation takes place relative to the NN cell spindle rotation by examining live embryos in which mitochondria were labelled with MitoTracker and microtubules were labelled with ensconsin::3xGFP. This analysis showed that the mitochondria, which initially localise around the nucleus (Fig. 5A, -9:00), start to segregate towards the animal pole at the onset of nuclear envelope break down (NEBD) (Fig. 5A, -7:00). By the time the mitotic spindle starts to rotate, the mitochondria have already formed a mass on the animal pole side (Fig. 5A, -5:00 time point; and Movies 3 and 4). The astral microtubules of the rotating spindle penetrate the mitochondria mass. A sagittal view (YZ view) of NN cells reveals that mitochondria remain in the sub-apical area during both segregation and spindle rotation (Fig. 5A).

Throughout the entire process, the nucleus and the spindle remain at a similar position relative to the apical contour of NN cells. In addition to the cytoplasmic asymmetry associated with mitochondria segregation in NN cells, we found that the spindle itself exhibits asymmetry (Fig. 5A). Soon after the spindle forms, it bulges on the side of the mitochondria mass (Fig. 5A, -5:00; Movies 3,4). As it rotates, the NN spindle takes on either a boomerang shape as in Fig. 5A, -4:00 (n=7/24), or a form that bulged on the side of the mitochondria mass (n=13/24); such that 20 out of 24 rotating spindles exhibited clear asymmetry. In order to quantify the asymmetry of the metaphase spindle before rotation, we carried out immunostaining of mitochondria and microtubules (Fig. 5B). Max intensity projections of the spindles were divided into mitochondria and contra-mitochondria side halves and the area (in pixels) of each half was measured (Fig. 5C). In this analysis, all spindles examined (n=7/7) exhibited asymmetry with a clearly visible bulge on the side of the mitochondrial mass (Fig. 5C). The bulged form of NN spindles is also clearly visible in a 3D reconstruction of an NN cell immunostained for microtubules and mitochondria (Movie 5). Thus, subcellular asymmetries precede spindle rotation in NN cells. The asymmetry of the spindle itself suggests the presence of a cytoplasmic microtubule-pulling force or stabilising activity on the side of the mitochondria mass.

Microtubules and the actomyosin system are involved in distinct processes during NN cell cytoplasmic segregation and spindle rotation

Both microtubules and the actomyosin system are required for the spindle rotation of NN cells (Fig. 4).

We similarly addressed whether the segregation of mitochondria in NN cells depends on these cytoskeletal elements by observing spindle and mitochondria dynamics in embryos treated with cytochalasin B, blebbistatin or low doses of nocodazole. These treatments revealed that the segregation of mitochondria requires microfilaments and myosin activity but is not affected by treatment with low concentrations of nocodazole (Fig. 6A). Following nocodazole treatment, we also observed a pronounced displacement of the spindle, with its reduced microtubule asters, towards the mitochondria mass (Fig. 6B,C and Movie 6). Together with the structural asymmetries observed in the NN spindle (Fig. 5), this result supports the existence of a pulling force towards the mitochondriaenriched cytoplasmic domain. It also suggests that intact microtubules are required to maintain the spindle position at the centre of NN cell apical contour. [START_REF] Bouchard | erythro-9-[3-(2-Hydroxynonyl)]adenine is an inhibitor of sperm motility that blocks dynein ATPase and protein carboxylmethylase activities[END_REF]. Embryos were treated with ciliobrevin A or EHNA just after mitochondria segregation and just before spindle rotation started (Fig. 7A). Treatment with these dynein inhibitors blocked NN spindle rotation. Furthermore, spindle asymmetry was no longer visible in the majority of inhibitor-treated embryos (4/6 ciliobrevin A-and 7/8 EHNA-treated embryos no longer exhibited clear asymmetry). Next, we addressed the subcellular localisation of dynein in NN cells using double immunofluorescence visualisation of dynein (antibodies against the dynein heavy chain, DHC) and mitochondria (anti-HSP60). This revealed that dynein is enriched in similar, but broader, cytoplasmic domain to the mitochondria (Fig. 7B). While it remains to be shown which subcellular organelles dynein proteins associate with, our results support the hypothesis that a dynein-dependent pulling force from the mitochondria-rich cytoplasmic domain acts on the NN cell mitotic spindle.

Dynein is localised in the mitochondria-rich cytoplasmic domain in NN cells

Mitochondria segregation and spindle rotation take place in isolated NN cells

Finally, we tested if the cellular environment influences the spindle rotation of the NN cell. The cell division of NN cells is coupled with differential fate specification of the daughter cells. In Ciona, this fate specification is controlled by a cell contact-dependent directional ephrin signal [START_REF] Picco | Ephrin-Eph signalling drives the asymmetric division of notochord/neural precursors in Ciona embryos[END_REF].

NN cells isolated at the beginning of their cell cycle undergo a symmetrical cell division in terms of the cell fate with both daughter cells adopting a notochord fate [START_REF] Picco | Ephrin-Eph signalling drives the asymmetric division of notochord/neural precursors in Ciona embryos[END_REF]. In similarlyisolated NN cells of Phallusia, the spindle still rotates, albeit with a reduced rotation angle (57.8 ± 11.8˚) compared to NN cells of intact embryos (74.0 ± 6.3˚) (Fig. 8A,B). Segregation of mitochondria also takes place in isolated NN cells with one spindle pole closely associated to the mitochondria mass (n=6/7) (Fig. 8A andMovie 7). This data suggest that the mitochondria segregation and the spindle rotation of NN cells can take place in the absence of cell-cell contacts, including the inductive signals that influence cell fate, as well as any physical constraints on the cell shape. However, it should be noted that the current experimental design did not allow us to determine whether the segregation and rotation took place along a specific axis.

Discussion

In this study, we analyse in detail the division of two sister cells, the NN and E cells, in the ascidian embryos of Phallusia mammillata. Both of these cells reorient their division axis, such that they divide along the same axis as their mother cell did. Our data provides evidence that the mitotic spindles of NN and E cells are oriented via distinct mechanisms. We propose the following model to describe the mechanisms of spindle orientation in these cells (Fig. 9). In the NN cell, the duplicated centrosomes arise on the animal pole side of the nucleus. The centrosomes migrate 90° in opposite directions such that the mitotic spindle forms orthogonal to the embryo midline (Fig. 1D). In this cell, cytoplasmic dynein and mitochondria, which are initially localised around the nucleus, displace towards the animal pole side of the cell in an actomyosin-dependent manner (Figs. 5, 6A and 7B). This displacement begins at the onset of NEBD and is completed by the time the mitotic spindle forms (Fig. 5A). We propose that this enrichment of cytoplasmic dynein exerts a net directional pulling force on both the polar and astral microtubules of the NN spindle, resulting in spindle asymmetry and rotation at metaphase. The elongated shape of the NN cell at metaphase may also contribute to the robust alignment of the mitotic spindle along the A-V axis, since spindle rotation is less pronounced in isolated cells compared to in intact embryos (Figs. 1C and8). In the E cell, the duplicated centrosomes are positioned on the vegetal pole side of the nucleus (Fig. 1D). From the onset of the cell cycle, the E cell is elongated along the A-V axis (Fig. 1C) and our data show a correlation between the elongated shape of the cell and the behaviour of the centrosomes (Fig. 3E). The duplicated centrosomes align along the A-V axis soon after their formation with the vegetal-side centrosome exhibiting a larger aster (Fig. 2B). While for the moment this remains entirely speculative, we propose that the cell cortex captures the astral microtubules of the distal aster, resulting in immobilisation of the distal centrosome.

In contrast, the proximal centrosome, with its smaller aster, migrates around the nucleus towards the animal pole so that the mitotic spindle forms directly along the A-V axis (Fig. 1D).

NN cell spindle rotation and cytoplasmic displacement

Orientation of the NN cell division appears to begin with the enrichment of mitochondrial and dynein to the animal pole side of the cell, although it is currently unclear if there is any causal relationship between these two observations. We propose that this movement creates a net-asymmetric cytoplasmic pulling force. Mitochondria displacement and segregation also take place in the NN cells of Ciona intestinalis [START_REF] Kobayashi | Differential gene expression in notochord and nerve cord fate segregation in the Ciona intestinalis embryo[END_REF] [START_REF] Takatori | Segregation of germ layer fates by nuclear migration-dependent localization of Not mRNA[END_REF]. In the NN cell division described here, however, the segregation of mitochondria and dynein appears to take place via a distinct mechanism. The nucleus and mitotic spindle of NN cells are not displaced towards the animal pole, but remain approximately at the centre of the apical contour (Fig. 6C). In addition, this cytoplasmic segregation is actomyosin-dependent but not affected by low doses of nocodazole. In both cases, the polarity cues directing cytoplasmic segregation remain unknown. Our ex-vivo experiment indicates that the mitochondria segregation in NN cells is a cell intrinsic property, although the experimental conditions used did not allow us to determine whether the segregation takes place along a specific axis.

While a causal link between the cytoplasmic concentration of mitochondria and dynein and the rotation of the NN cell spindle remains to be formally demonstrated, our data support the existence of dynein-dependent net-asymmetric pulling force from the cytoplasmic domain enriched with mitochondria. Asymmetry in the NN spindle itself suggests a microtubule pulling or stabilising force within the cytoplasm from the side of the mitochondrial mass (Fig. 5). Furthermore, the pronounced displacement of the spindle in nocodazole-treated embryos strongly suggests a pulling force towards the mitochondria side of the cell (Fig. 6). The preferential accumulation of dynein to the mitochondrial side of the cell, and the dependency of spindle rotation on dynein activity further support this hypothesis (Fig. 7). Co-immunodetection of mitochondria and dynein shows that dynein proteins follow similar temporal and spatial dynamics to mitochondria, although the two were not perfectly colocalised, suggesting that dynein proteins may (also) be associating with other organelles/structures (Fig. 7B). Motor proteins, including kinesin, dynein and myosin, are known to associate with intracellular organelles, including mitochondria, in eukaryotic cells and are important for transport of these organelles (reviewed in [START_REF] Rogers | Membrane trafficking, organelle transport, and the cytoskeleton[END_REF][START_REF] Vale | Myosin V motor proteins: marching stepwise towards a mechanism[END_REF]. In NN cells, the mitochondriaenriched cytoplasmic domain forms in an actomyosin-dependent manner and we propose that this cytoplasmic domain acts as a scaffold from which dynein exerts a pulling force onto both the astral and polar microtubules of the spindle. An organelle-mediated dynein-dependent cytoplasmic pulling force was shown to position the male pronucleus-centrosome complex in the centre of the C. elegans zygote, by exerting a net symmetric pulling force from evenly-distributed intracellular organelles, such as endosome, lysosome and yolk granule, onto the astral microtubules [START_REF] Kimura | Intracellular organelles mediate cytoplasmic pulling force for centrosome centration in the Caenorhabditis elegans early embryo[END_REF]. However, our model for the NN cell division differs importantly from this since the cytoplasmic segregation would lead to a net-asymmetric pulling force.

The subcellular organisation of the mitochondria and dynein-rich cytoplasmic domain as well as the mechanisms of the cytoplasmic segregation itself remain to be addressed. In addition, questions remain concerning the mechanisms whereby the NN spindle rotates. It is not yet clear how the spindle remains central despite the net-asymmetric pulling force. The displacement of the spindle towards the mitochondria-rich side of the cell following treatment of embryos with low concentrations of nocodazole suggests that microtubules are involved (Fig. 6; Movie 6). Astral microtubules could be important in maintaining the central position of the spindle by mediating isotropic cytoplasmic pulling forces or by mediating compressive forces generated by astral microtubule buckling at the cortex (reviewed in Minc and Piel, 2013). In the case of NN cells, in which the cytoplasmic pulling force is asymmetrical, it is easier to envisage that astral microtubules are mediating isotropic compressive forces to counterbalance the directional cytoplasmic pulling force.

We also do not understand how the proposed directional cytoplasmic pulling force is converted into the rotational movement of the NN spindle. In order to convert pulling forces into torques in the context of spindle rotation, an imbalance in the pulling forces acting on two spindle poles is envisaged (reviewed in [START_REF] Morin | Mitotic spindle orientation in asymmetric and symmetric cell divisions during animal development[END_REF]. For the NN cells, the mitotic spindle is first set perpendicular to the force-generating cytoplasmic domain situated on the animal pole side.

Assuming that the cytoplasmic pulling force is proportional to the length of astral microtubules [START_REF] Hamaguchi | Analysis of the role of astral rays in pronuclear migration in sand dollar eggs by the colcemid-UV method[END_REF][START_REF] Minc | Influence of cell geometry on division-plane positioning[END_REF], we propose two scenarios to account for the conversion of the cytoplasmic pulling force to a rotational movement. In the first, the two asters of NN spindle are similar in size and a slight tilt of the spindle axis relative to the axis of the pulling force generates the required imbalance. In the second, the two spindle poles exhibit distinct aster sizes, resulting in an imbalance in the pulling forces exerted on the poles. In either case, it remains a mystery how the spindles of a bilateral pair of NN cells have a tendency to rotate with bilateral symmetry (Fig. 2A).

Asymmetric centrosome migration in E cells

In contrast to the spindle rotation observed in NN cells, asymmetric centrosome migration results in the direct formation of the mitotic spindle along the A-V axis of E cells. A similar asymmetric centrosome migration has been shown in Drosophila, in male germline stem cells (mGSCs) [START_REF] Yamashita | Orientation of asymmetric stem cell division by the APC tumor suppressor and centrosome[END_REF] and in both larval and embryonic neuroblasts (NBs) [START_REF] Rebollo | Functionally unequal centrosomes drive spindle orientation in asymmetrically dividing Drosophila neural stem cells[END_REF]2009). In the cell divisions described in Drosophila, the centrosome exhibiting a larger aster associates with cortical polarity cues, such as a contact site with the stem cell niche (mGSCs) or the apical cortex (NBs), and becomes immobilised. Importantly, in the cell division of mGSCs and NBs, there is a strong correlation between centrosome age and behaviour, such that the mother centrosome becomes immotile in mGSCs and the daughter centrosome becomes immotile in NBs [START_REF] Yamashita | Asymmetric inheritance of mother versus daughter centrosome in stem cell division[END_REF][START_REF] Conduit | Cnn dynamics drive centrosome size asymmetry to ensure daughter centriole retention in Drosophila neuroblasts[END_REF][START_REF] Januschke | Drosophila neuroblasts retain the daughter centrosome[END_REF]. In E cells, the centrosome with a larger aster becomes immotile, similar to the examples in Drosophila, although we were unable to distinguish mother from daughter centrosomes in our experiments. One notable feature of the E cell centrosome behaviour is that the centrosome duplication appears to take place along the A-V axis of the cell, resulting in an alignment of the nucleus and two centrosomes along the axis. The larger distal (vegetal-side) centrosome becomes immobilised (Fig. 2D). We propose a model in which the larger size of the distal aster, together with the elongated shape of the E cell, may facilitate the capturing of the distal aster by the E cell cortex, resulting in its immobilisation (Fig. 9).

In conclusion, we analysed in detail the division of a pair of embryonic sister cells in ascidian embryos. Our data are consistent with our proposed model whereby: 1) in E cells a cell shapedependent mechanism directs asymmetric centrosome migration; 2) in NN cells a dynein-enriched cytoplasmic domain generates a net-asymmetric pulling force that leads to spindle rotation. It will be imperative in future study to address the molecular mechanisms controlling these oriented cell divisions. Additional works focusing on diverse cellular and developmental contexts are needed to understand the diversity of mechanisms that may control oriented cell division.

Materials and methods

Phallusia Embryos

Adult Phallusia mammillata were collected at Sète (Etang de Tau, Mediterranean coast, France).

Embryo culture, cell isolation, cell ablation and microinjection are previously described [START_REF] Sardet | Embryological methods in ascidians: the Villefranche-sur-Mer protocols[END_REF]). Ensconsin::3xGFP and EB3::GFP RNA were synthesized from pRN3 plasmid [START_REF] Lemaire | Expression cloning of Siamois, a Xenopus homeobox gene expressed in dorsal-vegetal cells of blastulae and able to induce a complete secondary axis[END_REF] using T3 mMESSAGEmMACHINE kit (Life Technologies), and injected at 2 µg/µl into unfertilized eggs. The cDNA encoding EB3::GFP was a gift from Dr A. Akhmanova. FM4-64 (Molecular Probes) was used at 20 µg/ml to observe cell membranes. Embryos were treated with 1 µM Mitotracker Red 580 (Molecular Probes) for 20 minutes to label mitochondria.

Pharmacological inhibitors

DMSO (Sigma), as control, was diluted 1:1000. Nocodazole (Sigma), cytochalasin B (Sigma) and blebbistatin (Sigma) were used at 5 nM, 2.5 µg/ml and 100 µM, respectively. Embryos were treated with inhibitors from the onset of the 32-cell stage. To block dynein function after mitochondrial segregation at the 32-cell stage, embryos were treated with ciliobrevin A or EHNA when vegetal cells started to show "decompaction" [START_REF] Nakatani | Induction of notochord during ascidian embryogenesis[END_REF]. ciliobrevin A (Sigma; as HPI-4) was used at 50 µM and EHNA (erythro-9-(2-Hydroxy-3-nonyl) adenin-e hydrochloride) (Tocris Biocience) at 0.5 mM. All stocks of inhibitors, with the exception of cytochalasin B and EHNA, were prepared in DMSO and diluted to at least 1:1000 in seawater. The stocks of cytochalasin B and EHNA were prepared in ethanol and diluted to 1:1000 and 1:100 in seawater.

Immunostaining

For microtubule immunostaining, embryos were fixed in cold methanol overnight and washed with PBS containing 0.1 % Triton-X (PBSTr). DM1A (Abcam) was diluted 1:100 in PBSTr containing 1 % BSA (BSA/PBSTr). Alexa Fluor 594 Goat Anti-Mouse IgG (Molecular Probes) was diluted 1:200 in BSA/PBSTr. For co-immunostaining of microtubules and mitochondria, anti-HSP60 antibody [START_REF] Chenevert | Purification of mitochondrial proteins HSP60 and ATP synthase from ascidian eggs: implications for antibody specificity[END_REF] and Alexa Fluor 488 Goat Anti-Rabbit IgG (Molecular Probes) were added at 1:50 and 1:200 dilution to the primary and secondary antibody reactions, respectively. For immunostaining of dynein heavy chains, embryos were fixed in artificial seawater containing 4% PFA overnight at 4 ˚C and then washed with PBSTr. Dynein HC antibodies (R-325) (Santa Cruz Biotechnology) were diluted at 1:50 in BSA/PBSTr. As secondary antibody, the MaxPO system (Nichirei Bioscience) was used at a dilution of 1:20 and signals were amplified using a TSA system, as described previously [START_REF] Hudson | β-Catenin-driven binary fate specification segregates germ layers in ascidian embryos[END_REF]. For co-immunostaining of mitochondria and dynein heavy chains, NN18 (ICN) [START_REF] Chenevert | Purification of mitochondrial proteins HSP60 and ATP synthase from ascidian eggs: implications for antibody specificity[END_REF] and Alexa Fluor 488 Goat Anti-Mouse IgG (Molecular Probes) were added at 1:50 and 1:200 dilutions, respectively. Immunostained embryos were mounted with VECTASHIELD Mounting Medium with DAPI (Vector Laboratories).

Imaging and quantification

Images of fixed samples and time-lapse movies of live samples were acquired using a Leica SP5 confocal microscope system with a Leica HCX PL APO 40x/1.3NA objective lens [START_REF] Negishi | Practical tips for imaging ascidian embryos[END_REF]. DIC time-lapse movies were acquired with a Zeiss Axiovert 200 microscope with a Zeiss plan achromat 40x/0.95NA objective lens operated using MetaMorph software (Universal Imaging Cooperation). For all image acquisitions, embryos were titled so that the apical contour of NN or E cells was as parallel as possible to the imaging plane with the embryonic median plane perpendicular to the coverslip [START_REF] Negishi | Practical tips for imaging ascidian embryos[END_REF]. All image processing and analyses were conducted using ImageJ software (National Institutes of Health). All statistical analyses were conducted using GraphPad Prism 6 software (GraphPad Software).

For all image analyses, if necessary, image stacks were reoriented so that the apical contour faces parallel to the screen. To quantify the cell shape of NN and E cells (Fig. 1B,C), we measured the length and width of their apical contour based on 4D image stacks of embryos labelled for microtubules (ensconsin::3xGFPs) and membrane (FM4-64) (Fig. 1B). For the length, we measured the line that is parallel to the embryonic midline and crosses the centre of the nucleus or of the spindle.

For the width, the line that is perpendicular to the length and crosses the centre of the nucleus or of the spindle was measured.

To quantify the size difference of two asters in E cells (Fig. 2B,D), 3D image stacks of embryo immunostained for microtubules were converted into binary images and the size of the asters, in pixel counts, in each z-section was measured.

To quantify the degree of the asymmetry of centrosome migration in E cells (Fig. 3D), we measured the angle of two lines crossing the centre of the nucleus (Fig. 3C). The first line is from the site in which centrosome duplication took place to the centre of the nucleus, while the second line joins the positions of the two centrosomes when migration is complete. In this quantification system, 90° would indicate that two centrosomes migrated equal distance, thus the centrosome migration was symmetrical, whereas 180° would indicate complete asymmetric migration. For this quantification, we selected two time points, at the beginning and end of centrosome migration, from 4D image stacks of ensconsin::3xGFPs-expressing embryos. If necessary, 4D image stacks were first reoriented so that the apical contour of the cell faced parallel to the screen.

To quantify the width of E cells (Fig. 3E), we measured the line perpendicular to the embryonic midline crossing the centre of nucleus as in Fig. 1B.

To quantify the degree of spindle rotation in NN cells (Figs. 4B and8B), an angle corresponding to the difference in the spindle axes at the onset of metaphase and at the onset of anaphase was measured based on corresponding 3D stacks selected from 4D image data of embryos expressing ensconsin::3xGFPs.

To quantify the area of mitochondria side half and contra-mitochondria side half of NN spindles (Fig. 5C), 3D image stacks of embryos immunostained for microtubules were analysed as follows. Using max intensity projection, 3D stacks were then projected into 2D images, which were subsequently converted into binary images. The number of pixels covering mitochondria side half and contra-mitochondria side half of NN spindles was counted as the area representing the NN spindle halves.

To quantify the distance of the displacement of NN spindles in the animal pole direction following treatment of cytoskeletal inhibitors (Fig. 6B,C), first the 4D image stacks of ensconsin::3xGFPs-expressing embryos were reoriented so that the embryonic midline run vertically. YZ-view images were then reconstructed from selected 4D image stacks. In these YZ images, the cell centre (Pc) was defined at the midpoint of the line connecting the two apical edges (Fig. 6B). In order to define the position of the prometaphase nucleus, Pn, the nucleus centre was projected onto the apical-edge line at a right angle (Fig. 6B). The spindle position at the onset of anaphase, Ps, was defined in the same manner as for Pn. The distances between Pc and Pn (dn) and between Pc and Ps (ds) were measured in order to quantify the relative position of these organelles along the A-V axis of NN cells. The animal side of the apical centre was measured with positive numbers and the vegetal side with negative numbers. The distance of the spindle displacement was obtained by subtracting dn from ds (ds-dn). All dn values were within ±3.2 m of the cell centre at prometaphase. The NN cell appears in the upper half of the frame and the E cell in the lower half. Animal pole side is up and embryo midline on the right. This movie was processed with max intensity z-projection of selected confocal planes. Frames were collected at 1-min intervals and the movie was generated at 4 frames/second.

  al, 2013; Takatori et al, 2010; Kim et al, 2007; Minokawa et al, 2001; Picco et al, 2007). Here we provide a detailed description of the oriented cell division of NN and E cells along the A-V axis and present our evidence showing that distinct mechanisms govern spindle orientation in these two cell types.

  Our evidence for a microtubule-pulling force prompted us to address the potential involvement of dynein motor proteins during NN cell spindle rotation (reviewed in Morin and Bellaïche, 2011; Mitchison et al, 2012). We used two pharmacological inhibitors of dynein: ciliobrevin A, a small molecule inhibitor of the AAA+ ATPase motor cytoplasmic dynein (Firestone et al., 2012) and EHNA (erythro-9-[3-(2-hy-droxynonyl)]adenine)

  as well as in the NNE cell (mother cell of NN and E cells) in three solitary ascidian species, Phallusia, Ciona and Halocynthia (Zalokar and Sardet, 1984; Hudson et al, 2013; Takatori et al, 2010). In Ciona and Halocynthia, the segregation of mitochondria to the NN cell is accompanied by segregation of newly-synthesised zygotic transcripts (Hudson et al, 2013; Takatori et al, 2010). In Halocynthia NNE cells, mitochondria and zygotic transcripts are displaced in association with the nucleus and retained at the animal pole in a Wnt5-dependent manner (Takatori et al, 2010). The mitotic spindle then migrates back to the centre of the apical contour just before initiation of cytokinesis. Nuclear migration and thus cytoplasmic segregation are microtubuledependent

Figure 1 .

 1 Figure 1. Oriented cell division of the medial NN and E cells

Figure 2 .

 2 Figure 2. Description of oriented cell divisions of NN and E cells

Figure 3 .

 3 Figure 3. Correlation between cell shape and asymmetric centrosome migration in E cells

Figure 4 .

 4 Figure 4. Role of cytoskeletal elements in NN spindle rotation

Figure 5 .

 5 Figure 5. Spindle asymmetry in NN cells

Figure 7 .

 7 Figure 7. Dynein is localised in the cytoplasmic domain enriched with mitochondria in NN cells

Figure 8 .

 8 Figure 8. Mitochondrial segregation and spindle rotation in isolated NN cells

Figure 9 . 1 .

 91 Figure 9. Model proposing a mechanism for spindle orientation in NN and E cells

Movie 2 .

 2

Acknowledgements

We thank Clare Hudson, Cathy Sirour and Evelyn Houliston for helpful discussions and comments on the manuscripts, Alex McDougall and members of his group for assistance with Phallusia, Janet Chenevert for providing anti-HSP60 and NN18 antibodies and Laurent Gilletta and Sophie Collet for animal husbandry. TN was a recipient of a postdoc fellowship from the Agence Nationale de la Recherche. HY is a CNRS investigator. The current study was supported by the CNRS, the Université Pierre et Marie Curie, the ARC (1144) and the Agence Nationale de la Recherche (ANR-09-BLAN-0013-01).

Mean pixel sizes for the mitochondria-and contra-mitochondria-side halves are 177 (SD=33.8) and 123 (SD=29.5), respectively.