Sandra Cortijo
email: sandra.cortijo@cnrs.fr

Mirela Domijan
email: mirela.domijan@liverpool.ac.uk

Varodom Charoensawan
email: varodom.cha@mahidol.ac.th

Sónia Negrão
email: sonia.negrao@ucd.ie

Multi-Disciplinary Approaches to Plant Responses to Climate Change

Keywords: climate change, systems biology, omics analyses, interdisciplinary research, bioinformatics

In August 2021, the Intergovernmental Panel on Climate Change (IPCC, 2021) released a new report on the anthropogenic effects of climate change. Climate change is causing a steady rise of global surface temperature and atmospheric CO 2 concentration, increasing the frequency and intensity of temperature extremes, causing heat waves, heavy precipitation, and, in some regions, agricultural and ecological droughts [START_REF] Schiermeier | Droughts, heatwaves and floods: how to tell when climate change is to blame[END_REF]. Such knock-on effects of climate change have a dire impact on plants, affecting plant development and reproduction, impacting natural vegetation and agriculture. Hence, to overcome changes in their environment, plants must adapt swiftly and respond to changes in their natural settings or else face extinction.

Environmental cues, such as progressive changes in photoperiod, or in day and nighttime temperatures, allow the plant to make intricate developmental decisions to negotiate diverse environmental conditions through different seasons [START_REF] Wilczek | Effects of genetic perturbation on seasonal life history plasticity[END_REF][START_REF] Zhao | Temperature-dependent growth contributes to long-term cold sensing[END_REF]. On the other hand, severe and sudden environmental changes lead to stresses that the plants must quickly respond and adapt to, in order to survive [START_REF] Balasubramanian | Potent induction of Arabidopsis thaliana flowering by elevated growth temperature[END_REF][START_REF] Berg | An examination of climate-driven flowering-time shifts at large spatial scales over 153 years in a common weedy annual[END_REF]. Thus, it is crucial for us to continue to advance our overall understanding of how plants respond and adapt to environmental changes, and to work toward improving their resilience in face of the unpredictability of climate change.

Several decades of studies, especially in model plants such as Arabidopsis, have demonstrated the complexity of molecular mechanisms, that plants employ to perceive and respond to the ever-changing environments, highlighting the need for large-scale data analysis, integration, and comparison of multiple related data sets, as well as for the combination of multidisciplinary approaches to address such complex problems, e.g., [START_REF] Jung | Phytochromes function as thermosensors in Arabidopsis[END_REF], [START_REF] Cortijo | Transcriptional regulation of the ambient temperature response by H2A.Z nucleosomes and HSF1 transcription factors in arabidopsis[END_REF], [START_REF] Awlia | Genetic mapping of the early responses to salt stress in Arabidopsis thaliana[END_REF], and [START_REF] Sriden | Large-scale comparative transcriptomic analysis of temperature-responsive genes in Arabidopsis thaliana[END_REF]. More recently, latest technological developments and improvements have allowed the acquisition of phenomics, physiological, transcriptomic, epigenomic, and microscopic data, as well as comprehensive data integration, in non-model crops and agricultural plants, as summarized in Zenda et al. in this Research Topic, and also other recent reviews [START_REF] Wee | Tools for high-spatial and temporalresolution analysis of environmental responses in plants[END_REF][START_REF] Scossa | Integrating multiomics data for crop improvement[END_REF][START_REF] Yang | Applications of multi-omics technologies for crop improvement[END_REF].

This Research Topic highlights multidisciplinary efforts that tackle responses to a wide variety of environmental changes associated with climate change, including cold (Wang et al.), drought (Wang et al.; Liu et al.) and even mineral deficiency (Zhang et al.). Interestingly, all the articles presented in this issue focus on non-model crop plants, highlighting the importance of applying omics and multi-omics techniques beyond conventional model plant species, as well as showing that omics techniques have been fully developed to be successfully translated in a wide variety of crop species.

In this Research Topic, Wang et al. combined transcriptomics and metabolomics to explore genes and metabolites that are involved in responses to cold and drought stresses in Poa crymophila Keng, a variation of gramineous forage grass widely distributed in the Qinghai-Tibet Plateau. The authors were able to identify the phenylpropanoid pathway, among a few others, as the key mechanism that allows the plant to adapt to these harsh environments. Using transcriptomics in combination with physiological analyses, Liu et al. have characterized morphological and physiological responses against chilling stress in 14 pumpkin (Cucurbita moschata) varieties, and identified differentially expressed genes in the leaves and the roots, notably, several in the α-linolenic acid biosynthesis pathway. Zhang et al. used a combination of phenomics, statistical methods and physiological measurements, including photosynthetic indicators to monitor the root architecture phenotypes of cotton (Gossypium hirsutum) under Phosphorus deficiency. The authors presented RhizoPot, an improvised in situ root phenotyping observation device, and found that several changes in root surface area are important indicators of cotton root phenotypes under low phosphorus. Finally, Zenda et al.

reviewed the recent developments in omics that can help us toward crop improvement and breeding more resilient plants.

In summary, the articles presented in this Research Topic further reiterate that multidisciplinary approaches have already shown their importance, and are on their way to becoming indispensable in our quest to understand how the model plants as well as non-model crops respond to a wide range of stress conditions. This will in turn pave the way for the rational design and development of crops that are resilient to various stresses associated with climate change.

Frontiers in Plant Science | www.frontiersin.org

March 2022 | Volume 13 | Article 876432

FUNDING

Work in the laboratory of VC was supported by the midcareer researcher grant from National Research Council of Thailand (NRCT) and Mahidol University (NRCT5-RSA63015-24), and Mahidol University's Postdoctoral Fellowship (MU-PD_2021_09). SN acknowledges the funding from Science Foundation Ireland through the SFI President of Ireland Future Research Leaders under Grant No. 18/FRL/6197.

AUTHOR CONTRIBUTIONS

VC, SC, MD, and SN wrote and revised the article. All authors contributed to the article and approved the submitted version.

Conflict of Interest:

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Publisher's Note: All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Anais Ogrizek
email: anais.ogrizek@gmail.com

Marie Rose Moro

Jonathan Lachal

Incarcerated mothers' views of their children's experience: A qualitative study in French nurseries

Keywords: Incarcerated mothers, prison nursery, mother-child interactions, family relations, social adaptation, child development, psychological outcome, Qualitative research

Context

In France, young children of incarcerated women live with their mothers in prison in specific units called nurseries, up to the age of 18 months. Only a few studies have examined the impact of this environment on these children. This study sought to explore through mothers' narratives how they perceive their children to experience this environment and how it affects their development.

Method

We used semi-structured interviews to collect the perceptions of 25 mothers about their children's experience of daily life in 12 different prison nurseries in France. Interpretative phenomenological analysis was used to explore the data.

Results

According to mothers, prison is a sensorially aggressive environment for children that may impair the children's sensory abilities. The children's and mothers' emotional perceptions of anger, sadness, insecurity, and imprisonment intertwine. Prison is home for these children, which leads them to establish strong bonds with this inside world -prison staff, other inmates-but to develop reluctances and concern about exploring the outside world.

Conclusion

The prison's sensorial environment may overlay the sensorial environment created by the mother, interfering with early mother-child interactions and leading to emotional misattunement. The gap between what the mother and the child each feel to be their home, and the social group they feel they belong to, causes pain to the mothers. This could alter their mandate of being a protective shield that could reduce the traumatic potential of the carceral environment on their babies. These mothers' sentences must be reconsidered for the sake of their children.

Introduction

Every year in Europe, 10,000 children live with their mothers in prison [START_REF]World Health Organization Europe -Data and statistics[END_REF], in accordance with the International Convention for Children's Rights, which notes that "a child benefits from living with his mother during his first months of life" [START_REF]La Convention internationale des droits de l'enfant (CIDE)[END_REF]. According to the latest data available from the French national prison administration, on average each month, 42 children were incarcerated along with their mothers in 31 different prisons from January through August 2019 [START_REF]Etablissements pénitentiaires disposant de quartier femmes[END_REF]. These children are not inmates from a legal perspective (4) and accordingly are not in custody [START_REF]Code de procédure pénale -Article D149[END_REF]. Rather, in France as in many other countries, young children up to the age of 18 months (24 months in exceptional circumstances) stay with their mothers in prison [START_REF] Staff | Laws on Children Residing with Parents in Prison[END_REF]. These mothers and children reside in specific units called "nurseries", together with pregnant women from their 6 th month of gestation. They are separated from the other women inmates.

The organization of these nurseries in France is defined by a ministerial circular [START_REF][END_REF] providing that the mother and her child must be kept in an individual cell, of 15 m2, usually open 8 hours a day. They are to have access to a kitchen, living room, and courtyard, shared with other mothers and children, if not for them alone. It also requires the prison to provide the mother with all appropriate supplies for the baby's care. Unfortunately, hosting conditions in nurseries can differ substantially between establishments and do not always comply with these guidelines [START_REF] Sannier | La maternité en détention[END_REF].

The residence of these children is the object of many recommendations across Europe [START_REF] Vis | Mères et bébés en prison[END_REF] and worldwide [START_REF] Shlonsky | Literature Review of Prison-based Mothers and Children Programs: Final Report[END_REF][START_REF] Elmalak | Babies Behind Bars: An Evaluation of Prison Nurseries in American Female Prisons and Their Potential Constitutional Challenges[END_REF][START_REF]Babies and small children residing in prisons -OHCHR Library Catalogue[END_REF]. Child psychiatrists agree that the evidence is clear that mothers and children should not be separated in the children's first years of life (13) Nevertheless, only a few studies have explored the impact of this environment on these children [START_REF] Ferrara | Health care of children living with their mother in prison compared with the general population[END_REF], on the mother-child relationship [START_REF] Cassidy | Enhancing attachment security in the infants of women in a jail-diversion program[END_REF] and motherhood.

We seek to explore this question through mothers' narratives of what they believe to be their children's experience of this environment and its impact on their development.

Method

This is an observational phenomenological study based on the analysis of interviews with mothers in French nurseries in different regions. This study was approved by the ethics committee CER-PARIS DESCARTES (CER 2018-108, 19/01/2019).

Participants

Our population was composed of women incarcerated in French prisons with a nursery unit. The criteria for inclusion were: adult women, currently or within the past year incarcerated in a French prison nursery and living with her child there. Women with a psychiatric or physical disorder possibly disruptive for the interview were excluded.

Data collection

Participants were recruited from February 2019 through February 2020.

After obtaining administrative consents,1 a visit by the principal investigator to the nursery was organized, and individual oral and written information about the study was provided to the mothers before requesting and receiving their written consent.

A semi-structured individual interview was then set up, based on questions developed by the research group (Table 1). The principal investigator began with a few open-ended questions that led to a dialogue with the mothers around the research themes. Each interview was around 60 to 90 minutes in length; they were audio recorded and fully transcribed.

Analysis

The study used interpretative phenomenological analysis (IPA) [START_REF] Smith | Interpretative phenomenological analysis[END_REF], because of the access it enables to the mothers' feelings and perceptions of their experiences. The analytic process for this method involves the following steps: interviews are read and reread repeatedly by the researcher, who annotates the transcripts during each reading. The comments are then assembled into different themes that reflect the major ideas identified by them. Links are then identified and developed between the themes to produce a logical and fluid organization.

Next, metaconnections are determined between these themes, and the themes then assembled into different metathemes that describe global aspects of the subjects' experiences. Therefore, each and every metatheme attracts different themes, themselves composed of the different comments to the interview transcript. At each step in analyzing each interview, the researcher must pay attention to the consistency of the links made between the different themes, by a constant back-and-forth analysis of the different interviews.

Two researchers (AO and JM) conducted the analysis, using NVIVO 11 software as a tool.

Sticking points were discussed during research staff meetings.

Results

Twenty five women were recruited in 12 detention centers: 20 women with children born during their incarceration in the nursery, 2 women with small children born outside of the nursery initially placed there with the baby at the outset of their incarceration, and 3 women still in the prison but whose children had been moved out of it in the past year. Table 2 describes the characteristics of both the mothers and the children. Four women refused to participate: two because of a language barrier, and two who did not specify their reason.

Our results are organized around 4 themes: Prison as a sensorially aggressive environment,

The children's emotional reaction to this specific environment, Children as prisoners, and

Prison as a home.

Prison as a sensorially aggressive environment

Prison must be considered a fiercely aggressive acoustic environment. It exposes the children to constant, brutal, and high intensity noises that may, according to their mothers, interfere with their biological rhythms.

"The thing I am the most worried about for my child is the noise. The other inmates' music; the screaming, when they shout, the doors slamming; I think that it is not appropriate for children." C2

"We hear noise all the time, there is this inmate banging over there, this other woman who yells all the time." B1 "Every day is the same: fighting in the courtyard, yelling, shouting in the evening, so the babies cannot sleep, they keep on crying." F3

Habits of both prison guards and some inmates sometimes inflict visual stimuli on babies that they cannot handle. Mothers worry that living in the nursery might also affect their adult visual capacities.

"

Children as prisoners

Children kept with their mothers in prison are inevitably kept away from the outside world, which makes it difficult, even impossible, for them to explore it.

"I put her at the window to show her the sky, I do as I much as I can to show her the outside world." F2

"The child needs to go outside more often, to discover the outside world, because here, he can only see the wall and the birds, that's it." H1 "Here she can only see women; she's never seen a man." M1 "He's 17 months old, he's never been out of here, never." A1

While prisoners know it will be difficult to adapt back to their "normal" life once they are released, these mothers feel that it will be even harder for their children, when they are transferred out of prison, without their mother, to adapt to their new environment, so different from prison.

"Car horns, animals, trees, my [former] day-to-day life… I think it might be a long-lasting trauma for her, it might give her disabilities, or I don't know…

Prison as a home

These children born in prison have never known any other living environment. Most children consider home to be the place where they were born and where they live. The mothers report that the children become used to their life in prison; the prison's routine becomes their dayto-day life, and this worries their mothers.

DISCUSSION

In prison, mothers and children are kept together locked inside a very small cell most of the day, with rare contact with the outside world. Although this condition may appear to forecast a very close mother-child relationship, our results indicate that it may actually impede emotional and affective attunement between the mother and her child.

First, the children are exposed to a harsh living environment. This could affect the development of sensory abilities [START_REF] Black | Early childhood development coming of age: science through the life course[END_REF][START_REF] Pedrosa | Factors predicting sensory profile of 4 to 18 month old infants[END_REF]. Sensory abilities appears very early in the fetus development, during pregnancy [START_REF] Clark-Gambelunghe | Sensory development[END_REF]. These abilities actually constitute the baby's prelanguage skills, that he uses to distinguish his mother [START_REF] Hugill | The senses of touch and olfaction in early mother-infant interaction[END_REF] and to interact with her [START_REF] Bowlby | Attachment and loss: Retrospect and prospect[END_REF][START_REF] Brazelton | The Earliest Relationship: Parents, Infants, And The Drama Of Early Attachment[END_REF][START_REF] Winnicott | Babies and Their Mothers[END_REF]. The mother also uses hers to create a reassuring environment for her child's healthy development [START_REF] Mellier | The psychic envelopes in psychoanalytic theories of infancy[END_REF]. But the grimness of the prison's sensory environment may overlay that created by the mother and interfere negatively with the emergence and evolution of early mother-child interactions [START_REF] Hugill | The senses of touch and olfaction in early mother-infant interaction[END_REF][START_REF] Doucet | The "smellscape" of mother's breast: effects of odor masking and selective unmasking on neonatal arousal, oral, and visual responses[END_REF] by impeding the mother's reassuring capacities toward her child [START_REF] Nishitani | The calming effect of a maternal breast milk odor on the human newborn infant[END_REF].

As a result, the tonic-emotional dialogue which usually helps mother and child understand their mutual emotional states (28) seems unsettled. Emotional misattunement (29) may lead to communication misunderstandings between the mother and her baby.

Second, there is a gap between what the mother and the child each feel to be their home.

The mothers emphasize their separation from the outside world, their home, their usual life outside the prison walls. On the contrary, their children, raised in prison, consider it their home. As mothers try desperately to introduce their children to their outside home and protect them from the prison environment, the children seem to turn away from that outside world, remaining attached to their inside prison home.

This gap is widened by the social organization in prison. Goffman defined prison as a totalitarian institution, in which all aspects of human life (work, hobbies, nighttime) take place in a single setting, under one single authority, exerted by the prison guards [START_REF] Goffman | Asylums: Essays on the Social Situation of Mental Patients and Other Inmates[END_REF] As a result of the dichotomy between a limited number of powerful guards, and a large number of secluded and disempowered inmates, including the mothers of these children, conflictual relationships arise between these populations. But children do not fit into this organization: they build solid relationships with guards as well as with other prisoners, with no status distinctions. These bonds disrupt the well-established institutional "etiquette" of prisoners on one side, and guards on the other and create an unintentional connection between those two populations.

The children, innocent and free, but residing in prison, do not belong to any of these "clans", and form a population of their own, apart from their mothers.

This complex situation causes suffering among the mothers. Their description of their babies' negative emotions thus appears to us to reflect mainly their own negative perceptions of the prison environment and the intolerability of feeling that their children are attached to this prison environment that they reject.

The interviews showed the mothers' ambivalence about this environment: they consider it threatening, unsafe, stressful, and causing great sadness to their children, but in the meantime they allow their babies to build strong bonds with the staff and other inmates and to share moments of joy and peace. Prison is the only totalitarian institution in which residents are guilty rather than victims of their condition [START_REF] Goffman | Asylums: Essays on the Social Situation of Mental Patients and Other Inmates[END_REF], and feelings of guilt and shame may arise for exposing their child to this environment, resulting in this ambivalence.

CONCLUSION

The mothering abilities of some of these women appear to be affected by their situation and their distress. Mentally healthy mothers can establish a protective shield meant to reduce the potential negative impact of the brutal sensory environment on their child [START_REF] Freud | Beyond The Pleasure Principle[END_REF], especially before the acquisition of intersubjectivity [START_REF] Stern | The Interpersonal World of the Infant: A View from Psychoanalysis and Developmental Psychology[END_REF][START_REF] Golse | Le développement psychique précoce: De la conception au langage[END_REF] But what happens when the mothers' skills are affected by distress? What will happen when the baby leaves the harsh environment that is its home and meets the real world?

The traumatic potential of the carceral environment on mothers and their babies should be considered when they are sentenced. Whenever possible, these mothers and babies should be kept out of prison, under non-custodial sentences. When prison is inevitable in some rare situations, some of their mothers' outside world should be introduced to these children: by bringing some of it inside the prison -through some familiar objects or furniture the woman is attached to, or sensorial vectors of her outside life (food or music) that she could bring inside her cell -but also by giving these children direct access to the outside world, though regular accommodation in outside children nurseries. So that the children can be raised in a setting that has some resemblance to their mothers' social, emotional, and affective environment. This could limit the potential psychological trauma of these children when released from prison into the outside world [START_REF] Bowlby | Attachment and loss: Retrospect and prospect[END_REF] and, paradoxically, create an even closer bond between them than simply living physically very close to one another in prison can.

Incarceration of mothers and their child in prison nurseries creates an emotional, affective and social gap between them that could be difficult to close at their release. The suffering of these mothers, due in part to their guilt for their responsibility for this situation, reinforces the difficulties in attuning mother-child relationship. Alternatives to incarceration must be preferred for these mothers and their children whenever possible.

KEY MESSAGES

• Only a few studies have examined the impact of prison environment on children kept with their mothers in prison nursery.

• This study explores how the mothers perceive their children to experience this environment and how it affects their development.

• Semi-structured interviews wereled, and Interpretative phenomenological analysis was used to explore the data.

• For these children, being raised in prison creates a gap between them and people from the outside world -including their mothers -which might interfere with early mother-child interactions and lead to emotional misattunement.

• These mothers' sentences must be reconsidered for the sake of their children.

Among the 31

 31 such prisons in France (3), authorizations were obtained for 19 of them, located in all 10 French prison interregional administrative areas (16) Unfortunately, mothers were incarcerated with children in only 12 of these establishments, representing only 8 interregional areas. They were recruited in the interregional areas of Bordeaux (1 prison), Lille (3 prisons), Lyon (1 prison), Paris (1 prison), Rennes (3 prisons), Strasbourg (1 prison), Toulouse (1 prison) and French oversea territories (1 prison).

The children's emotional reactions to this specific environment

  Smell is a highly developed sense in early childhood. Children use it to distinguish the different people around them, especially their mothers. Prison's brutal smells might impair these children's sense of smell and olfaction development. It looks like he sleeps very lightly." J1 "He can't sleep on his own. When we were at home he could, alone in his crib. But since we arrived here, he can't anymore. When I put him in the crib, he wakes up straight away..." K5 "I have to take him with me so that he can fall asleep." E4 Mothers say they can feel their children expressing anger and sadness, and they relate it to a

	"I really don't want her to smell the cigarette smoke, so sometimes I put her in her bouncing
	chair and put a scarf over it so that the smoke can't reach her." F2
	"Some inmates smoke cigarettes at their cell window, and sometimes not very legal stuff…
	with an even stronger smell. The smoke comes into the cell." H1 feeling of imprisonment.
	"If you go to the cell right next to mine ... You'll have an awful smell; an old, rotten, musty
	smell, because the flooring made out of wood is very humid and there's mold under it." E1
	t eat
	this, she doesn't eat this either, she hasn't had the time to taste this and this and this'." G1
	"Four to six months is the start of taste discovery, so she should be able to have access to all
	kind of tastes." F2

When you're sleeping, some guards turn on the light, which wakes him up. Then he doesn't sleep anymore." K3 "Some guards leave the lights on, which makes it difficult for us to sleep, so it wakes the baby up." I1 "Since I'm here, I have double vision because our visual field is so limited. It makes it hard for me to read. I hope it won't affect her eyes, or I'll feel even more guilty." E1 Discovering different tastes through dietary diversification is another important aspect of the child's exploration of the world. Mothers worry that prison might impair it. "There aren't a lot of vegetables: there are zucchini, potatoes, but you can't find any turnips, or sweet potatoes… But he has to discover all kind of tastes!" B2 "I don't want to delay everything, so that when she leaves here, I have to say 'She doesn'Prison is inevitably permeated by a feeling of insecurity that, according to these mothers, puts their children into a state of constant altertness. "I feel like she's very stressed, she's always on alert, she's much tenser than when we can go out of the prison for a temporary furlough. Here I can see it on her face, it's not the same face as outside." E1 "He cries for all of this: the anger, the sadness, the stress maybe also, because he can't speak, he can't explain." K5 "My baby is startled whenever he hears the key opening the cell door in the morning or opening the courtyard. He looks frightened. He is moved in some way." K5 "Every morning, they open the door, 'bang, slam, clack'. My daughter is always startled." M4 Several mothers also report that their children have sleeping problems, which they attribute to the prison environment, perceived as threatening. "Sometimes, when he sleeps, he cries in his sleep, and I can see that he's not sleeping well because of the noise, the sadness… all of it… he also has nightmares." K3 ""He often wants to open the window, he holds on to the window bars, he likes to speak, yell, I can feel that he has this need to get out and take a big breath of fresh air." F1 "They have difficulties being locked in. My daughter, as soon as she sees the guards coming… she cries whenever they lock the door, and she kicks the door." E2 "When she's outside, she looks fine… but when we're locked in, she cries a lot, she can feel that we're imprisoned." K4 "When it's time to lock the door, she knows. So when the guard comes to lock the door, she always runs, she runs far away. When she runs, I see, I know that she's not ok." C2

"

  She knows the opening hours of the cell. Sometimes, when guards are late, she kicks the door, and pushes it." G2 "My daughter has done everything here, she grew up here, she knows nothing else.

	"My daughter sees her father 45 min, once a week. It is too short to build a proper
	relationship." G1
	To her
	it's her home, but it's not my home." L1 "He didn't know my sister-in-law, his cousin, he knew none of them, he cried, and didn't want
	to come near anyone." M3
	"She's getting used to it here, I can see that she's getting used to all this, and I don't like it."
	M2
	"I wanted to take him out, for a short day furlough but he wouldn't cross the line, he was
	crying." A1

The prison staff do appear to care for the children as if they were part of their own family, and the children seem to build strong relationships with both prison staff and other inmates.

"For his first birthday, the guards took everything in hand, one of the inmates made him a cake… they made a candle out of cardboard. He was really happy that day, he felt wonderful." A1 "He's in love with one of the guards!" A1 "She grew up here, so the guards have seen everything, her first words, her first laughs, her first steps, her first falls, her first mischief, … as my family would have outside … She calls some of the guards "daddy" … everybody knows her here." L1 "Some of the guards are authorized to bring baby clothes, because the child's not in custody … So he got presents from them." H1 "They gave everything to my son, for all his needs, everything, everything." K1 Some of the bonds established in prison can be sometimes even stronger than the ones with their biological family outside.

"When we go to the parlor, the children, the relatives come to see her. She kisses them because I ask her to, but otherwise she wouldn't even come close to them." G1 "We did all the administrative procedures so that his father could take him home, but my son cried a lot that day. He didn't want to go." A1

From the research and health departments of the central prison administration, each interregional prison administration, and every director of every prison visited.

Data availability statement

All data generated or analysed during this study are included in this published article and tables.


Martijn Kemerink

Christian Müller

Michael L Chabinyc

Martin Brinkmann
email: martin.brinkmann@ics-cnrs.unistra.fr

Organic and hybrid thermoelectrics

published or not. The documents may come   L'archive ouverte pluridisciplinaire

Introduction.

Exactly two centuries ago, in 1821, the Physicist Thomas Johann Seebeck discovered that a voltage appears at the junction between two materials subjected to a temperature gradient. About one decade later, in 1834, Jean Charles Athanase Peltier discovered the mirror effect, namely the presence of heating or cooling at a junction of two different conductors traversed by an electric current. Both these phenomena are exploited in thermoelectric materials. The Peltier effect is widely used to fabricate small cooling elements or fridges whereas the Seebeck effect is essential for the design of radioisotope thermoelectric generators that were for example used to power the Pioneer and Voyager spacecrafts from NASA. [START_REF] Rowe | Thermoelectrics Handbook: Macro to Nano[END_REF] For decades, the field of thermoelectricity has been dominated by inorganic materials. The working horse among inorganic thermoelectric materials, Bi 2 Te 3 , shows one of the highest reported values of the thermoelectric figure of merit ZT beyond 1 near room temperature.

(2) However, the increasing requirements for efficient energy management opened new perspectives for alternative organic/hybrid thermoelectric materials that are abundant and easy to process at low cost over large areas as compared to inorganic materials. They could help recover some usable energy from waste heat, eventually in combination with other energy sources such as light. An alternative application field of these hybrid/organic compounds would be in small-scale power supplies for stand-alone applications where batteries or photovoltaic solutions are problematic. In that short period of time, organic/hybrid thermoelectrics emerged as a new and promising branch in 'plastic electronics' and material science in general.

II. SPECIAL ISSUE.

In this context of emerging new organic/hybrid materials for energy generation, we organized the present Special Issue in Applied Physics Letters entitled "Organic and Hybrid Thermoelectrics."

This special issue provides an overview of state-of-the-art organic/hybrid thermoelectric materials and related physical and physico-chemical phenomena, such as charge transport and doping. This research field is by essence highly interdisciplinary, as it encompasses materials science, physics, chemistry, computer science, and engineering. For this special issue, we have selected articles that give a representative picture of the progress over the past decade in this topical area.

Most of the articles address well identified topics such as:

-Device architectures and systems (ref. [4][5][6] -New hybrid materials for thermoelectrics (ref. [7][8][9][10][11][12][13][14][15] -Material processing and structure-property correlations (ref. [16][17][18][19] -Doping mechanisms in p-and n-type thermoelectric materials (ref. [20][21][22][23][24][25][26] -Charge transport and related electronic phenomena (ref. [27][28][29][30] Beside these dominant topics, some papers also focus on the way to tune the thermopower by designing new polymer architectures or new hybrids and thermoelectric generators. We hope this collection of papers will invigorate this flourishing research field, serve as a source of inspiration and catch the attention of new researchers and students.

In 2011 ,

 2011 figures of merit ZT > 0.1 and reasonable stability. (3) This achievement triggered

ACKNOWLEDGMENTS

We would like to acknowledge all authors who contributed to this Special Topic. M. K. thanks the Carl Zeiss Foundation for financial support. MLC acknowledges support from the National Science Foundation (NSF) under DMR 1808622. MB acknowledges support from ANR under ANR-17-CE05-0012. This project has received funding from the European Union's Horizon 2020 research and innovation programme: Marie Sklodowska Curie project HORATES under grant agreement n° GA955837.


Isabelle Cassar-Malek
email: isabelle.cassar-malek@inrae.fr

Lise Pomiès

Anne De La Foye

Jérémy Tournayre

Céline Boby

Jean-François Hocquette

Transcriptome profiling reveals stressresponsive gene networks in cattle muscles

Keywords: Agricultural Science, Genomics, Molecular Biology, Zoology Pre slaugther stress, Cattle, Transcriptomics, Muscle response, Transcription factors

In meat-producing animals, preslaughter operations (e.g., transportation, mixing unfamiliar animals, food and water deprivation) may be a source of stress with detrimental effects on meat quality. The objective of this work was to study the effect of emotional and physical stress by comparing the transcriptomes of two muscles (M. longissimus thoracis, LT and M. semitendinosus, ST) in Normand cows exposed to stress (n = 16) vs. cows handled with limited stress (n = 16). Using a microarray, we showed that exposure to stress resulted in differentially expressed genes (DEGs) in both muscles (62 DEGs in LT and 32 DEGs in ST, of which eight were common transcription factors (TFs)). Promoter analysis of the DEGs showed that 25 cis transcriptional modules were overrepresented, of which nine were detected in both muscles. Molecular interaction networks of the DEGs targeted by the most represented cis modules helped identify common regulators and common targets involved in the response to stress. They provided elements showing that the transcriptional response to stress is likely to (i) be controlled by regulators of energy metabolism, factors involved in the response to hypoxia, and inflammatory cytokines; and (ii) initiate metabolic processes, angiogenesis, corticosteroid response, immune system processes, and satellite cell activation/quiescence. The results of this study demonstrate that exposure to stress induced a core response to stress in both muscles, including changes in the expression of TFs. These factors could relay the physiological adaptive response of cattle muscles to cope with emotional and physical stress. The study provides information to further understand the consequences of these molecular processes on meat quality and find strategies to attenuate them.

INTRODUCTION

In livestock species, psychological/emotional factors (including unfamiliar environment or social mixing), physical factors (including transportation, noise or vibrations), climatic factors (including temperature and humidity associated with transfer to the slaughterhouse), and deprivation of food and water are sources of emotional and physical stress. The exposure of animals to stressful conditions has several adverse impacts, including metabolic and health problems and poor welfare. Most of the above conditions often occur during farm management, during preslaughter operations [START_REF] Deters | Invited review: linking road transportation with oxidative stress in cattle and other species[END_REF] or during slaughter, and they have detrimental effects on nutritional and organoleptic meat quality, as reported in pigs and poultry [START_REF] Debut | Behavioural and physiological responses of three chicken breeds to pre-slaughter shackling and acute heat stress[END_REF][START_REF] Debut | Variation of chicken technological meat quality in relation to genotype and preslaughter stress conditions[END_REF][START_REF] Monin | Pork of low technological quality with a normal rate of muscle pH fall in the immediate post-mortem period: the case of the Hampshire breed[END_REF]. There is also compelling evidence that preslaughter stress has an undesirable effect on meat quality traits (e.g., low tenderness and juiciness) in both beef and lamb [START_REF] Ferguson | Have we underestimated the impact of pre-slaughter stress on meat quality in ruminants[END_REF][START_REF] Muchenje | Relationship between pre-slaughter stress responsiveness and beef quality in three cattle breeds[END_REF][START_REF] Terlouw | Stress reactivity, stress at slaughter and meat quality[END_REF][START_REF] Terlouw | Pre-slaughter conditions, animal stress and welfare: current status and possible future research[END_REF][START_REF] Terlouw | Understanding the determination of meat quality using biochemical characteristics of the muscle: stress at slaughter and other missing keys[END_REF][START_REF] Reiche | Pre-slaughter stress and horn status influence physiology and meat quality of young bulls[END_REF]. The impact of stress on meat quality has been explained by changes in physiological and metabolic properties of the muscle, which is converted to meat postmortem: higher depletion of glycogen before slaughter, less production of lactic acid (a byproduct of post-mortem glycolysis), and thus insufficient pH decline. Muscles with borderline pH (5.9-6.1) end up being very tough [START_REF] Marsh | Effects of early-postmortem glycolytic rate on beef tenderness[END_REF], leading to a defect known as dark cutting beef or dark, firm, and dry (DFD) meat.

Changes may be related to variations in protein activities, as well as in protein levels driven by changes in gene expression. Some studies have reported alterations in the muscle proteome of farmed pigs [START_REF] Morzel | Proteome changes during pork meat ageing following use of two different pre-slaughter handling procedures[END_REF], chickens [START_REF] Hazard | Functional genomics of the muscle response to restraint and transport in chickens1[END_REF][START_REF] Zanetti | A note on protein expression changes in chicken breast muscle in response to time in transit before slaughtering[END_REF], fishes [START_REF] Silva | Effects of preslaughter stress levels on the post-mortem sarcoplasmic proteomic profile of gilthead seabream muscle[END_REF] and cattle [START_REF] Díaz | What functional proteomic and biochemical analysis tell us about animal stress in beef?[END_REF][START_REF] Sentandreu | Protein biomarkers of bovine defective meats at a glance: gel-free hybrid quadrupole-orbitrap analysis for rapid screening[END_REF]. However, there are few data on the transcriptional response to stress in the muscle of meat-producing animals despite few studies in pigs [START_REF] Davoli | Transcriptome analysis of skeletal muscle tissue to identify genes involved in pre-slaughter stress response in pigs[END_REF] and in steers following surgery [START_REF] Zhao | Functional genomic analysis of variation on beef tenderness induced by acute stress in angus cattle[END_REF]. Herein, we examined the transcriptomic response in two different skeletal muscles from cows exposed to preslaughter stress conditions. We used these responses to infer stress-induced changes in the biological and physiological function of these muscles and discussed the biological functions affected by exposure to stress of psychological and physical origins and their potential impact on meat quality.

MATERIALS AND METHODS

Animals and samples

The experiment was conducted with 32 pure Normand cull cows of 48-60 months of age purchased from different private farms in western France. The cows were not pregnant or lactating and had a medium fatness score. They were housed in the experimental farm of the INRAE research centre (UE Herbipôle-Low Mountain Ruminant Farming Systems Facility; DOI 10.15454/1.5572318050509348E12) as described by [START_REF] Gobert | Pre-slaughter stress and lipoperoxidation: protective effect of vitamin E and plant extracts rich in polyphenols given to finishing cattle[END_REF] and [START_REF] Delosière | Lipid oxidation, pre-slaughter animal stress and meat packaging: can dietary supplementation of vitamin E and plant extracts come to the rescue?[END_REF].

The cows (mean live weight of 642 kg) received a straw (30%) and concentrate (70%) based diet supplemented with lipids (40 g oil/kg diet DM) obtained from extruded linseeds (60%) and rapeseeds (40%) for 101 ± 3 d. For one group of cows, this diet was the control diet described in [START_REF] Delosière | Lipid oxidation, pre-slaughter animal stress and meat packaging: can dietary supplementation of vitamin E and plant extracts come to the rescue?[END_REF]. For the other group, the diet was supplemented with vitamin E (155 IU/kg diet DM) and plant extracts rich in polyphenols (INRA patent #P170-B-23.495 FR; c7 g/kg diet DM, respectively; EP diet). Supplementation with vitamin E (lipophilic antioxidant) and plant extracts rich in polyphenols was used to protect against beef lipoperoxidation in a previous study on sheep [START_REF] Gladine | Bioavailability and antioxidant capacity of plant extracts rich in polyphenols, given as a single acute dose, in sheep made highly susceptible to lipoperoxidation[END_REF]. The cows received a morning and evening meal representing a daily quantity of 1.8 kg of concentrate and 0.8 kg of hay. They had free access to water. The cows received an isoenergetic and isonitrogenous ration for a mean daily body weight gain of 1.6 kg over the 101 ± 3 d finishing period. Animals were housed in groups of four in 6 × 6 m pens with straw bedding, according to a balanced design relative to feeding treatments. The cows were housed in a freestall barn equipped with individual feed bunks and automatic gates. They were individually offered their appropriate allowance of concentrates and straw each day. Feed intake was calculated daily and adjusted periodically to ensure a daily gain of 1,150 g/d. The finishing period of 101 d, slightly longer than the French standards for cattle (approximately 70 d), was chosen to achieve good production conditions and to facilitate experimental organization.

Cows were finished and slaughtered under conditions of limited stress (n = 16) or physical and psychological stress (n = 16) as described in [START_REF] Bourguet | Characterising the emotional reactivity of cows to understand and predict their stress reactions to the slaughter procedure[END_REF]. Groups included the same numbers of animals for each diet. For the limited stress condition, the cows were transported with a nonexperimental conspecific to avoid social isolation stress and were handled calmly. Specifically, for the stress condition, each cow was individually transported in a lorry (social isolation) towards an unfamiliar farm, where it was taken through a labyrinth by two purposefully noisy experimenters over a period of 30 min. The cow was then transported for 15 min to the experimental slaughterhouse. All cows were slaughtered without any electrical stimulation in compliance with INRAE ethical guidelines for animal welfare at the experimental slaughterhouse of INRAE. The cows were stunned by a captive bolt gun and exsanguinated, as performed in French commercial slaughterhouses. Carcasses were stored in a chilling room (4 C) for approximately 45 min following exsanguination. Carcasses were sold for human consumption as in any controlled commercial slaughterhouse. Stress status was evaluated through the plasma and urinary cortisol and heart rate as described in [START_REF] Bourguet | Characterising the emotional reactivity of cows to understand and predict their stress reactions to the slaughter procedure[END_REF]. Muscle samples from the M. semitendinosus (ST) (a himdmuscle involved in locomotion) and the M. longissimus thoracis (LT) (a support muscle for the body) were excised within 15 min after slaughter. They were immediately frozen in liquid nitrogen and stored at -80 C until RNA extraction.

As indicated in [START_REF] Bourguet | Characterising the emotional reactivity of cows to understand and predict their stress reactions to the slaughter procedure[END_REF] and [START_REF] Delosière | Lipid oxidation, pre-slaughter animal stress and meat packaging: can dietary supplementation of vitamin E and plant extracts come to the rescue?[END_REF], experimental procedures and animal holding facilities respected French animal protection legislation, including licensing of experimenters. The procedures were controlled and approved by the French Veterinary Services (agreement B63 345 17). The animal experimental design was described in and registered in the research unit quality management database.

Muscle transcriptome analysis

Transcriptome analyses were carried out with Agilent oligonucleotide microarrays designed with 10,064 probes (including 1,614 control probes) for 4,210 bovine genes, including more than 3,000 specific muscular genes. The microarray was designed for monitoring transcriptional changes in genes involved in muscle growth (including energy and protein metabolism), carcass composition, fat metabolism and beef quality (including marbling). The microarray was first described in Hocquette et al. (2012a) and subsequently used in [START_REF] Costa | Does growth path influence beef lipid deposition and fatty acid composition?[END_REF].

RNA extraction, RNA quality checking and quantification, target amplification and labelling, hybridization with the probes, and extraction of fluorescent hybridization signals were performed as previously described in Hocquette et al. (2012a).

Data were preprocessed by Feature Extraction 10.1 software (Agilent Technologies, Santa Clara, CA, USA) for all samples and probes. The probes not meeting the quality criteria (saturation and uniformity of spots, intensity above background noise, etc.) were filtered out. Each array was normalized by dividing the raw intensity values of its probe by the median intensity of the control probes of the array. Each probe intensity was then normalized by dividing its raw value by the median of the corresponding probes from all arrays. After removing the probes with missing values, a log 2 transformation was applied to the data.

The transcriptomic data were submitted to Gene Expression Omnibus (GEO) under the accession number GSE119912. Differential analyses were conducted via linear modelling with the diet supplementation à period à stress interaction factor to explain the probe levels. An empirical Bayes method was used to moderate the standard errors of the estimated log-fold changes using the R/Limma package (http://bioinf.wehi.edu.au/limma/) as described in [START_REF] Smyth | Statistical issues in cDNA microarray data analysis[END_REF] with a Benjamini and Hochberg multiple testing correction [START_REF] Benjamini | Controlling the false discovery rate-a practical and powerful approach to multiple testing[END_REF]. The genes for which at least 80% of the probes were consistently different at the adjusted p value 10% (i.e., estimated rate of true positives in the probe list of 90%) were retained and considered differentially expressed genes (DEGs). All probe ratios were found to be consistent for each DEG, meaning that for one gene, all probe ratios were lower than 1 or greater than 1.

Gene Ontology enrichment

Functional enrichment according to Gene Ontology Biological Process (GO BP) was assessed by submitting lists of accession numbers (for DEGs) or gene names (for common regulators and targets of the DEGs) to ProteINSIDEv2 [START_REF] Kaspric | ProteINSIDE to easily investigate proteomics data from ruminants: application to mine proteome of adipose and muscle tissues in bovine foetuses[END_REF], https://umrhbioinfo.clermont.inrae.fr/ProteINSIDE_2/). This workflow enables the analysis of lists of protein or gene identifiers from ruminant species and gathers biological information provided by functional annotations, putative protein secretion and protein interactions. It queries the g:Profiler database based on the most complete information available for Bos taurus. The list of array probes was used as a background list for enrichment analysis of the DEG lists. The GO enrichment test was declared significant at a Benjamini-Hochberg FDR < 0.08 (i.e., estimated rate of true positives in the gene list of 92%). The results are expressed as -log 10 (p value) on the graphs.

Identification of cis-transcriptional modules

Promoter sequences were extracted using the program Gene2Promoter (version 3.4.1; Genomatix Software Suite, Munich, Germany, www.genomatix.de) using the default settings, 500 bp upstream and 100 bp downstream of the transcription start site. We selected bovine promoters with at least one relevant transcript and preferentially a high quality level (experimentally verified 5′ transcript or with 5′ end confirmed by PromoterInspector prediction) and for whose number of conserved orthologous promoters was at least 50% of loci. This was performed for DEGs and for all of the genes in the microarray. As the coregulation of mammalian genes usually depends on a combination of TFs rather than individual TFs alone, cis-acting regulatory elements are often organized into frameworks of motifs called cis-transcriptional modules. The selected promoters were submitted to the 'ModelInspector' task of GEMS Launcher (version 4.1; Genomatix Software, Munich, Germany, www.genomatix.de) to search for cis-transcriptional modules. For this purpose, the promoter sequences of the genes were scanned for matches to the Promoter Module 5.4 Library (Vertebrate Module section). Fisher's exact test was then used to identify overrepresented cis-transcriptional modules in the DEG set compared to the total gene set of the microarray.

Construction of interaction networks

Network analysis was performed with Pathway Studio software version 12.0.1.9 using Elsevier's Resnet Mammal DataBase (Ariadne Genomics, Rockville, MD, USA). Gene interaction networks were built with the DEGs targeted by the most represented cistranscriptional modules for each muscle (targeting at least 5 and 4 DEGs for LT and ST, respectively) and with the DEGs targeted by the nine overrepresented cis transcriptional modules common to both muscles, generating two muscle-specific networks and one common network of stress response. For each set of genes, to reconstruct the network, Pathway Studio searches known relations between the genes and adds regulators and the expression targets common to them. Filters were applied to identify only key expression regulators and targets of each network. To be added to the network, target genes must be linked to a minimum of three bibliographic references and have at least six known relations in the Pathway Studio Database. For regulators, three bibliographic references and two known relations are also needed, except for the regulators of the nine common modules for which a cut-off of five relations was chosen.

Finally, Venn diagrams were used to identify the major regulator genes and major target genes among the DEGs targeted by cis-transcriptional modules specific to LT and ST and in both muscles. Subnetworks between DEGs and their major regulator genes and between DEGs and their major target genes were extracted.

Validation of differential expression levels

A RT-qPCR assay was performed on the LT samples of 10 animals/group for 4 genes (ATF3, CEBPD, SMAD7 and FOS) with the StepOne Plus TM Real-Time PCR System using the Power SYBR1 Green master mix (both Applied Biosystems, Foster City, CA, USA). The GeNorm algorithm [START_REF] Vandesompele | Accurate normalization of real-time quantitative RT-PCR data by geometric averaging of multiple internal control genes[END_REF] was used to determine the optimal number of reference genes required to effectively normalize the qPCR data. Four housekeeping genes were selected: UXT, MRPL39, CLN3 and TOP2B. The primer sequences (Table S1) with an annealing temperature of 60 C were designed using Primer3 software. qPCR was performed using a StepOnePlus thermocycler (Applied Biosystems, Foster City, CA, USA). The PCR efficiency of each primer pair was tested with a 10-fold dilution series of purified cDNA. Each reaction was subjected to melting curve analysis to ensure the specificity and integrity of the PCR product. Student's t test was used to test the significance of the difference between the limited stress and stress groups.

Identification of genes corresponding to DEGs in quantitative trait loci (QTLs)

A query of genetic information from the lists of the DEGs and the common regulators and targets of the DEGs was performed with the QTL module included in ProteINSIDEv2. Briefly, each DEG was searched on NCBI to retrieve the location of the corresponding gene on the genome. Then, this location was compared with the QTL positions in the QTL database "AnimalQTLdb". The location on the genome must be included entirely in a QTL to consider that the DEG is mapped in the QTL.

RESULTS

Transcriptomic profiles

We recorded changes in gene expression profiles in the LT and the ST. Individual data are available in the GEO repository under accession number GSE119912. No effect was detected for the EP diet, the stress à EP diet or the stress à diet in either muscle, but an effect of stress was detected (P < 0.1). In the stressed cows compared to cows handled with limited stress, microarray analysis revealed changes in the abundance of 67 transcripts in the LT (including 43 up-and 24 downregulated transcripts; P < 0.1, Table S2) corresponding to 62 DEGs with unique gene names (Fig. 1). In the ST, changes in 36 transcripts were detected (including 33 up-and 3 downregulated transcripts; P < 0.1, Table S2) corresponding to 32 DEGs with unique gene names (Fig. 1). Among the differentially expressed transcripts, 27 were common to both muscles, corresponding to 24 unique gene names (Fig. 1). They included eight known transcription factors (TFs): SMAD7, ETS2, MYOG, ATF3, HES6, CEBPD, HEYL, and FOS (Table S2). In addition, muscle-specific genes were differentially expressed according to the stress status (38 in the LT and 8 in the ST; Fig. 1). These genes included four TFs (MYOD1, MYF6, CEBPB, and HES1) and one transcription cofactor (MED23) in the LT and a transcriptional activator (ZNF750) in the ST. The differential abundance of four TF transcripts (ATF3, CEBPD, SMAD7 and FOS) was checked by qPCR experiments in the LT and confirmed the observed changes, as illustrated in Table 1.

Lists of DEGs according to stress status were submitted to biological information mining through Gene Ontology (GO) term enrichment compared to the background list of the microarray (Data S1). In LT, 9 GO biological process (GO BP) terms were enriched (P < 0.08). In the ST, 26 GO BP terms were enriched (P < 0.08). As illustrated in Fig. 2, 9 of these GO terms identified in both the LT and ST: regulation of gene expression (23 genes in the LT, 13 in the ST), transcription by RNA polymerase II (16 genes in the LT, 12 in the ST), regulation of transcription by RNA polymerase II (16 genes in the LT, 12 in the ST), regulation of biosynthetic process (21 genes in the LT, 14 in the ST), regulation of cellular biosynthetic process (20 genes in the LT, 13 in the ST), regulation of macromolecule biosynthetic process (20 genes in the LT, 13 in the ST), regulation of cellular macromolecule biosynthetic process (20 genes in the LT, 13 in the ST), skeletal muscle cell differentiation (4 genes in the LT, 3 in the ST), and muscle organ development (8 genes in the LT, 5 in the ST). For each considered GO BP, the list of genes included both common and muscle-specific DEGs. There were no significant GO cellular component terms at P < 0.08 in both the LT and ST muscles (data not shown). In the LT, 18 GO molecular factor (MF) terms were enriched (P < 0.08). In the ST, 19 GO MF terms were enriched (P < 0.08). Sixteen of these GO terms were enriched in both the LT and ST. They mainly refer to DNA-binding and transcription regulator activity (Supplemental Dataset).

Cis-transcriptional modules

A promoter analysis was performed with Gene2promoter of the Genomatix Software Suite to identify common TF binding sites in the promoter regions of genes-called cis transcriptional modules-that may account for coregulation among differential transcripts. For 52 of the DEGs in the LT, 168 promoters were retrieved from the Genomatix Promoter Table 1 Validation of some differentially expressed genes following pre-slaughter stress in the Longissimus thoracis muscle. The abundance of some DEGs detected by microarray analysis was quantified by qRT PCR in the Longissimus thoracis muscle of stress cows vs. cows handled with limited stress (2n = 20). Variation of reference genes used for normalization was computed with the GeNorm software package. Student t-test was used to test the significance of the difference between the two conditions. Database, of which 111 were selected for further analysis (Table S3). ModelInspector enabled the retrieval of 288 different cis-transcriptional modules (at 1,378 locations).

For 28 of the DEGs in the ST, 84 promoters were retrieved. Of these, 57 promoters were further analysed with ModelInspector, and 201 cis-transcriptional modules were found (on 675 match positions). The same analysis was performed for all the genes represented on the microarray. As illustrated in Table S3, 24 cis-transcriptional modules were identified as overrepresented DEGs compared to the genes represented on the microarray (P < 0.1) in the LT and 25 in the ST. Nine of the modules were overrepresented in both muscles. The cis-transcriptional modules and the DEGs targeted by these modules in each muscle as identified by ModelInspector are listed in Table 2. Cis-transcriptional modules with binding sites for TFs in the ETS family and SP1 family had a high occurrence in the promoters of the DEGs in both muscles.

Interaction networks and identification of regulators and main targets of DEGs

Finally, with Pathway Studio 2, we constructed interaction networks between the DEGs targeted by the overrepresented cis-transcriptional modules for each muscle and between the DEGs targeted by the 9 overrepresented cis-transcriptional modules common to both muscles. We thus generated muscle-specific networks and one core network of stress responses. Then, using the Pathway Studio 2 database, we searched for the main regulators and the main targets of the 3 networks (Data S2, sheets 1-6). We next identified the similarities among the lists obtained from these datasets to identify the key common regulators and targets (Data S2, sheets 7-8). Ten main regulators of stress-responsive genes were identified: AKT1, EGF, HIF1A, IFNG, IL1B, INS, MAPK1, MAPK14, TGFB1, and TNF. GO mining showed that they were highly related (P < 0.001) to the regulation of oxidoreductase activity and specifically the regulation of monooxygenase activity, inflammatory response, immune system, and carbohydrate metabolism (transport and metabolic processes) (Data S2, sheet 7). A list of 14 main targets of the DEGs was identified: BCL2, BGLAP, CDKN1A, COL3A1, ERBB2, FN1, ICAM1, IL6, MMP2, PPARG, SELE, SLC2A4, TLR4, and VEGFA. GO mining showed that they were highly related (P < 0.001) to response to cytokine, response to oxygen levels, response to glucocorticoids, response to stress, immune system, angiogenesis, and carbohydrate homeostasis (Data S2, sheet 8). The networks of the DEGs and the common regulators and of the DEGs and common targets are presented in Fig. 3. The list of DEGs (LT-specific, ST-specific and common DEGs), the main regulators and the main targets of the DEGs are summarized in Table 3. Twenty-four of the corresponding genes were located in QTLs linked to meat quality: shear force (CDIPT, CEBPD, DNAJB4, GPAM, RAB3IL1, MAPK1, and TLR4), muscle compression (ADRB2), tenderness score (ADRB2, CDIPT, RAB3IL1, and IFNG), muscle pH (DLL4, ERBB2), juiciness (ATP1B1, DFFB, RAB3IL1, SELE), and marbling (CEBPD, DLL4, ERBB2, GADD45, ICAM1, IL1B, IL6, LEAP2, MYF6, PDK4, PMP22, SMAD7, TNF). Some genes were also mapped in QTLs associated with marbling score (CEBPD, DLL4, DNAJB4, GADD45A, LEAP2, MYF6, PDK4, PIGM, PMP22, RAB3IL1, and SMAD7) and with lipid class contents (e.g., monounsaturated fatty acid content and conjugated linoleic acid content: ADRB2; omega-6 to omega-3 fatty acid ratio and palmitic acid content: LCAT; trans-11, cis-15-C8:2 fatty acid content: GADD45A).

DISCUSSION

Transcriptional response to stress

Understanding how preslaughter stress impacts muscle physiology would provide information for the management of beef quality, especially tenderness. In this study, we compared the muscle transcriptional profiles of cows exposed to preslaughter emotional and physical stress with those of control cows handled with limited stress. We hypothesized that this approach may be useful for investigating the molecular mechanisms of the stress response and the potential impact on meat quality. We identified changes in the abundance of several gene transcripts in two muscles of the cows exposed to stress. We found evidence of a common transcriptional response in both muscles, albeit with different metabolic types and activities, even though some muscle-specific DEGs were detected. Notably, there was a core stress response in both muscles, as shown by common DEGs and common GO terms (mainly related to the regulation of gene expression and muscle development). The highest number of DEGs was detected in the LT. This may be related to the high oxidative metabolism of the LT (Hocquette et al., 2012b), which makes it more prone to changes in oxidative status and therefore susceptible to stress. Muscle gene expression in response to stress likely also depends on sex [START_REF] Oster | Transcriptional responses of PBMC in psychosocially stressed animals indicate an alerting of the immune system in female but not in castrated male pigs[END_REF], the nature and intensity or duration of the stress, and breed, which may explain some differences in the results of our study and a previous study on Angus animals [START_REF] Zhao | Functional genomic analysis of variation on beef tenderness induced by acute stress in angus cattle[END_REF]. Nevertheless, the regulation of genes involved in carbohydrate, lipid, and protein metabolism is likely to occur in many cases, as observed in this study, as well as in other studies on cattle [START_REF] Sporer | Transportation stress in young bulls alters expression of neutrophil genes important for the regulation of apoptosis, tissue remodeling, margination, and anti-bacterial function[END_REF][START_REF] Zhao | Functional genomic analysis of variation on beef tenderness induced by acute stress in angus cattle[END_REF] and pigs [START_REF] Davoli | Transcriptome analysis of skeletal muscle tissue to identify genes involved in pre-slaughter stress response in pigs[END_REF]. Table 3 Components of the molecular response initiated by pre slaughter stress in two muscles of cows as revealed by transcriptomic signatures. The differentially expressed genes (DEGs), the main regulators and the main targets of the 3 datasets of the DEGs as identified by Pathway Studio are listed. Query of genetic information was performed with the QTL module included in ProteINSIDE in order to retrieve information on the location of the genes encoding proteins of interest within published Quantitative trait loci (QTLs) for meat and carcass. This module interrogates the publicly available QTL database "Animal QTLdb".

Type of response Gene name

Newly translated transcription factors and their related biological pathways

While the short-term response to stress may be primarily driven by changes in protein phosphorylation (e.g., reversible phosphorylation [START_REF] Mato | The first evidence of global meat phosphoproteome changes in response to pre-slaughter stress[END_REF]), as well as enzyme activity or protein abundance, our study provided convincing evidence that the response to stress includes a transcriptional component, as previously reported in two studies [START_REF] Davoli | Transcriptome analysis of skeletal muscle tissue to identify genes involved in pre-slaughter stress response in pigs[END_REF][START_REF] Zhao | Functional genomic analysis of variation on beef tenderness induced by acute stress in angus cattle[END_REF]. Indeed, functional annotation of the lists of DEGs revealed the enrichment of GO terms related to the regulation of gene expression and transcription. It is well accepted that the primary response to stress involves the activation of pre-existing TFs by phosphorylation [START_REF] Sabban | Stress-triggered activation of gene expression in catecholaminergic systems: dynamics of transcriptional events[END_REF]. Our data indicate that newly translated TFs may also relay the stress response, as 11 of the DEGs encode TFs. Eight of these TFs were identified in both muscles, of which some were detected as nodes in the molecular networks associated with the response to stress. The majority of the differentially expressed TFs were upregulated, except for two muscle regulatory factors (MYOG and MYOD1) and a transcriptional repressor (HES1). MYOG and MYOD1 are basic helix-loop-helix family TFs essential for myogenesis, including during the regenerative process [START_REF] Zammit | Function of the myogenic regulatory factors Myf5, MyoD, Myogenin and MRF4 in skeletal muscle, satellite cells and regenerative myogenesis[END_REF]. HES1 is a downstream target of Notch [START_REF] Borggrefe | The Notch signaling pathway: transcriptional regulation at Notch target genes[END_REF]. It is also a master regulator of glucocorticoid receptor-dependent gene expression. It is silenced by the primary stress hormones glucocorticoids [START_REF] Revollo | HES1 is a master regulator of glucocorticoid receptor-dependent gene expression[END_REF]. The observed downregulation of HES1 was not surprising since stressed cows showed high plasma and urinary cortisol levels [START_REF] Bourguet | Characterising the emotional reactivity of cows to understand and predict their stress reactions to the slaughter procedure[END_REF]. Of the upregulated TFs, 4 were basic leucine zipper (bZip) TFs: FOS, ATF3, CEBPB, and CEBPD. ATF3, a member of the mammalian cAMP-responsive element-binding protein (CREB) family, is induced by various stresses. ATF3 is a sensor for a wide range of conditions and modulates the immune response, atherogenesis, cell cycle, apoptosis, and glucose homeostasis [START_REF] Jadhav | Activating transcription factor 3 in immune response and metabolic regulation[END_REF]. ATF3 has been considered an adaptive response gene with a dual mode of action to activate (as a homodimer) or repress (as a heterodimer) target gene expression. It was proposed that ATF3 functions as a "hub" of the cellular adaptiveresponse network that helps cells adapt to disturbances of homeostasis [START_REF] Hai | ATF3, a hub of the cellular adaptive-response network, in the pathogenesis of diseases: is modulation of inflammation a unifying component?[END_REF]. ATF3 was also found to be differentially expressed following acute stress induced by surgery in Angus beef [START_REF] Zhao | Functional genomic analysis of variation on beef tenderness induced by acute stress in angus cattle[END_REF]. The bZip proteins CEBPB and CEBPD are members of the C/EBP family, which participates in a number of biological responses, including energy metabolism, cell proliferation and differentiation, and immune responses [START_REF] Ramji | CCAAT/enhancer-binding proteins: structure, function and regulation[END_REF]. Their binding sites are found in the regulatory regions of a large number of acute phase proteins. A dual role was proposed for the C/EBP proteins as mediators of both inflammatory responses and glucocorticoid effects [START_REF] Nerlov | The C/EBP family of transcription factors: a paradigm for interaction between gene expression and proliferation control[END_REF][START_REF] Roos | The emerging role of C/EBPs in glucocorticoid signaling: lessons from the lung[END_REF]. CEBPD expression is induced by inflammatory effectors and hypoxia and promotes proinflammatory signalling and hypoxia adaptation [START_REF] Balamurugan | The many faces of C/EBPδ and their relevance for inflammation and cancer[END_REF]. CEBPB was also recently identified as a novel regulator of satellite cell homeostasis that promotes differentiation at the expense of self-renewal [START_REF] Alsudais | CCAAT/enhancer binding protein β is required for satellite cell self-renewal[END_REF].

Overrepresented TF binding sites in the promoters of DEGs and related biological pathways

Several cis-transcriptional modules were located in the promoters of the DEGs. Modules common to both muscles were detected mainly in the promoters of common DEGs, while muscle-specific cis-transcriptional modules were detected in the promoters of muscle-specific DEGs. However, some specific cis-transcriptional modules were detected in the promoters of common DEGs. FOS was targeted by 4 common modules in both muscles plus 1 specific module in the ST. ATF3 was targeted by 1 common module in both muscles and by 3 specific modules (1 in the LT and 2 in the ST). Examination of cis-transcriptional modules of DEGs from both muscles revealed that binding sites for the transcription factor SP1 and for members of the ETS family are often included in those modules. SP1 is ubiquitously expressed and, in addition to functioning as a 'housekeeping' TF, may be a key mediator of gene expression induced by insulin and other hormones [START_REF] Solomon | A critical role of Sp1 transcription factor in regulating gene expression in response to insulin and other hormones[END_REF]. ETS1 is a highly conserved TF that controls the expression of cytokines, chemokines and angiogenesis factors [START_REF] Russell | Transcription factor Ets-1 in cytokine and chemokine gene regulation[END_REF]. ETS binding sites were found in the promoter of common differentially expressed TFs as well as in the promoter of 11 of the 15 LT-specific DEGs.

Other biological pathways related to the response to stress

Our study provided additional evidence that the response to stress interacts with the immune response, inflammatory response, and chemotaxis, as well as the production of interleukins (IL-16 in the LT; IL-1 B and IL-6 as main regulators and targets of the DEGs; and IL-10 and IL-13 in the list of common targets of the DEGs). This is consistent with previous studies examining the response to stress in livestock animals: A transcriptional shift in acquired and innate immunity pathways was reported in the peripheral blood of psychosocially stressed pigs [START_REF] Oster | Transcriptional responses of PBMC in psychosocially stressed animals indicate an alerting of the immune system in female but not in castrated male pigs[END_REF]. Amplified inflammatory activity was also detected in blood neutrophil expression in young bulls following truck transportation for 9 h (Buckham [START_REF] Sporer | Transportation stress in young bulls alters expression of neutrophil genes important for the regulation of apoptosis, tissue remodeling, margination, and anti-bacterial function[END_REF] and in the LT muscle of Angus beef exposed to acute stress induced by surgery [START_REF] Zhao | Functional genomic analysis of variation on beef tenderness induced by acute stress in angus cattle[END_REF]. Moreover, a conserved transcriptional response to chronic social stress involving increased expression of proinflammatory genes (including IL-6 and IL-8) has been reported in blood leukocytes [START_REF] Powell | Social stress up-regulates inflammatory gene expression in the leukocyte transcriptome via β-adrenergic induction of myelopoiesis[END_REF] in mice and humans. In our study, changes in chemokine and cytokine expression in muscle were most likely part of the adaptive mechanisms contributing to the stress response (Fig. 3). IL-16 is a lymphocyte chemoattractant factor also classified as an "alarmin" [START_REF] Rider | Alarmins: feel the stress[END_REF]. IL-6 and IL-8 are also regarded as myokines released from muscle in response to contractions [START_REF] Brandt | The role of exercise-induced myokines in muscle homeostasis and the defense against chronic diseases[END_REF]. Muscle-derived IL-6 may mediate some of the anti-inflammatory and insulin-sensitizing effects of physical exercise (Covarrubias [START_REF] Anthony | IL-6 strikes a balance in metabolic inflammation[END_REF].

Another striking result of our study is the upregulation of transcripts related to the carbohydrate metabolic pathway, e.g., transcripts encoding PDK4 (an inactivator of pyruvate dehydrogenase complex; targeted by 1 cis-transcriptional module in the LT and 3 modules in the ST), PFKFB3 (a glycolysis regulator; targeted by 1 cis-transcriptional module in the LT and 2 cis-transcriptional modules in the ST) and SLC25A25 (a mitochondrial ATP transporter; targeted by 2 cis-transcriptional modules in the LT). This illustrates a switch in energy metabolism in the muscles of animals exposed to exercise and psychological stress towards anaerobic metabolism to support ATP production for muscle contraction. PDK4 plays a pivotal role in controlling metabolic flexibility [START_REF] Zhang | The pivotal role of pyruvate dehydrogenase kinases in metabolic flexibility[END_REF], and its expression is increased in response to moderate intensity exercise.

Analysis of molecular networks also highlighted the contribution of the response to oxygen levels/hypoxia in the response to stress, albeit via different transcripts and different contractile and metabolic muscle types. Consistently, the TF hypoxia inducible factor (HIF1A) was identified as a main common regulator of the DEGs. This could be a signature of oxygen imbalance or of the physical activity imposed on the cows. Thus, it may not be surprising that the expression of PFKB3, a downstream target of HIF, was upregulated. Hypoxia was also demonstrated to cross-talk with the Notch signalling pathways, which regulate satellite cell quiescence and self-renewal [START_REF] Liu | Hypoxia promotes satellite cell self-renewal and enhances the efficiency of myoblast transplantation[END_REF]. Since quiescent satellite cells have a low metabolic rate, fewer mitochondria and anaerobic metabolism, this is likely part of the adaptive signature of muscle to stress. Thus, the combined signatures of hypoxia, the Notch signalling pathway [START_REF] Fukada | Molecular signature of quiescent satellite cells in adult skeletal muscle[END_REF], MYOD1 downregulation [START_REF] Kopan | The intracellular domain of mouse Notch: a constitutively activated repressor of myogenesis directed at the basic helix-loop-helix region of MyoD[END_REF], and IRFD1 (an inducer of regenerative myogenesis) downregulation further indicate that quiescent satellite cells are stress targets and most likely physical activity targets.

Putative effects on meat quality

Finally, the transcriptomic muscle response to preslaughter stress may have an impact on meat quality through energy metabolism and hypoxia. Indeed, anaerobic glycolysis is highly relevant to beef quality since it is involved in postmortem protein degradation and hence beef tenderization during meat ageing (reviewed by [START_REF] Maltin | Determinants of meat quality: tenderness[END_REF]. This process is regulated by the decline in muscle pH due to the conversion of glycogen into lactate following the lack of oxygen after slaughtering. Stress was shown to markedly affect meat tenderness by increasing postmortem ultimate pH [START_REF] Purchas | An assessment of the role of pH differences in determining the relative tenderness of meat from bulls and steers[END_REF] due to the depletion of glycogen stores by stress prior to slaughtering, which leads to dark-cutting meats. Reliable indicators of the occurrence of high pH and preslaughter stress were identified in the sarcoplasmic proteome of muscle [START_REF] Fuente-Garcia | Search for proteomic biomarkers related to bovine pre-slaughter stress using liquid isoelectric focusing (OFFGEL) and mass spectrometry[END_REF][START_REF] Sentandreu | Protein biomarkers of bovine defective meats at a glance: gel-free hybrid quadrupole-orbitrap analysis for rapid screening[END_REF]. They were mainly involved in metabolism, chaperone-and stress-related processes, muscle contractility/fibre organization, and transport activities. In our study, several genes encoded by the DEGs and the common regulators or targets of the DEGs were located in bovine QTLs of the meat and carcass group associated with meat quality parameters known to be impacted by stress: muscle pH (DLL4, ERBB2), shear force (CDIPT, CEBPD, DNAJB4, GPAM, RAB3IL1, MAPK1, and TLR4), tenderness score (ADRB2 CDIPT RAB3IL1, and IFNG), compression (ADRB2), and juiciness (ATP1B1, DFFB, and RAB3IL1). Some genes were also mapped in QTLs associated with marbling score and with lipid class contents. However, the relationships between transcript levels and these meat quality parameters remain to be studied.

CONCLUSIONS

Exposure to emotional stress (novelty, social isolation, presence of active humans, noise) and physical effort (walking) prior to slaughter induced a transcriptional response in two muscles in cows. Our data provide evidence of a coordinated response in two muscles of stressed animals due to the identification of common target genes, associated functions, cis-transcriptional modules, regulators and downstream targets. The response included an interplay between metabolic changes (glycolytic), hypoxia, inflammatory process, and satellite cell renewal/quiescence, likely due to elevated cortisol. However, the relative contribution of mechanisms related to stress and to physical activity induced by walking the labyrinth remains to be elucidated.

From an animal production perspective, the identification of gene networks activated by stress will improve the understanding of the molecular mechanisms of meat conversion and beef quality defects caused by preslaughter stressful conditions suffered by cattle. The target stress-responsive gene network could be modulated by management factors (on farm nutrition, antioxidant supplementation, etc.) to reduce the adverse impact of stress.

FoldFigure 1

 1 Figure1Venn diagram visualizing the intersection of the lists of the gene names of the differentially expressed genes (DEGs) in response to pre slaughter stress in the M. longissimus thoracis (LT) and in the M. semitendinosus (ST). A subset of 24 common DEGs was assigned to a set of core stress responsive genes. The two subsets of DEGs only in the LT (n = 38) or in the ST (n = 8) were considered as components of the muscle-specific response to stress. Transcriptional regulators are underlined: Transcription factor (unbroken line), transcriptional modulator (dotted line).Full-size  DOI: 10.7717/peerj.13150/fig-1

Figure 2

 2 Figure2Common GO terms across muscles for the differentially expressed genes (DEGs) in response to preslaughter stress. Lists of DEGs were submitted to functional annotation compared to the microarray background (data available in Additional File 3). The intersection of the lists of GO terms was computed at http://bioinformatics.psb.ugent.be/webtools/Venn/. Gene names capitalized in bold are common DEG between muscles. LT: M. longissimus thoracis; ST: M. semitendinosus.Full-size  DOI: 10.7717/peerj.13150/fig-2

Table 2

 2 Over-represented transcriptional modules in the promoter of the stress-responsive genes in the muscles of cows. The transcriptional modules were searched with the module inspector function of Genomatix, their occurrence was examined in the promoters of genes of the experimental datasets and the number of target genes was determined in each dataset.

	Muscle Module	p	Occurrence	Number of	Gene ID
			value	of module	target genes
	LT	ETSF_ETSF_01	0.076 16	15	IL16 SERPINE1 HES1 HSPBAP1 CDIPT XYLT2 TUBB3 THBS1 MED23
						GPAM PIGM CEBPD HEYL HES6 MYOD1
		SP1F_CAAT_02	0.040 10	9	NME6 SDC4 PDPR HES6 PFKFB3 CDIPT PMP22 THBS1 IFRD1
		CAAT_AP1F_01	0.035 8	8	SLC25A25 SERPINE1 NME6 IMP3 HSPBAP1 SLC2A3 THBS1 ATF3
		SP1F_EBOX_SP1F_01	0.024 8	7	DFFB GLUL PDK4 IMP3 PMP22 XYLT2 CEBPD
		CAAT_SP1F_01	0.088 5	5	SERPINE1 ATP1B1 GLUL GEM HES6
		GATA_GATA_GATA_01 0.037 5	3	NME6 SLC16A6 MED23
		YY1F_SRFF_02	0.016 3	3	SLC2A3 ATF3 FOS
		SORY_SORY_EGRF_01	0.061 3	3	MUSK ATP1B1 RAB3IL1
		NFKB_NFKB_01	0.064 3	3	SLC25A25 GLUL GEM
		HNF1_GATA_01	0.098 3	3	MED23 PLD1 ATP1B1
		KLFS_NR2F_KLFS_01	0.024 3	2	SERPINE1 TUBB3
		STAF_SP1F_01	0.026 2	2	GLUL HEYL
		RXRF_EBOX_01	0.043 2	2	PDPR RAB3IL1
		AP1F_SMAD_01	0.055 2	2	IL16 THBS1
		ETSF_AP1F_04	0.067 2	2	ACOT11 HSPBAP1
		CEBP_MYBL_03	0.076 2	2	ACOT11 HSPBAP1
		AARF_CEBP_01	0.091 2	2	ABRA NME6
		BRNF_RXRF_02	0.066 4	1	DLL4
		NFKB_ETSF_01	0.007 2	1	DLL4
		SRFF_AP1F_01	0.047 1	1	FOS
		ETSF_SP1F_SMAD_01	0.062 1	1	HEYL
		YY1F_SRFF_01	0.076 1	1	FOS
		PAX8_NKXH_01	0.076 1	1	PMP22
		ETSF_SRFF_01	0.091 1	1	FOS
	ST	SP1F_SP1F_06	0.002 30	14	PGF GADD45A SLC16A6 ADAMTS9 CYP1A1 SLC2A8 SDC4 PMP22
						TUBB3 IFRD1 HYAL2 ATF3 HES6 HEYL
		NFKB_SP1F_03	0.002 12	8	SLC2A8 SDC4 MYLK4 PGF LRP4 PMP22 HEYL CEBPD
		SP1F_ETSF_04	0.087 8	8	ABRA SDC4 PGF LCAT PMP22 CYP1A1 SMAD7 HES6
		SMAD_E2FF_01	0.088 12	7	SLC2A8 SDC4 IFRD1 PDK4 CEBPD HES6 FOS
		SP1F_YY1F_01	0.044 10	7	PGF ABRA GEM SDC4 SLC2A8 ATF3 HES6
		SP1F_CAAT_02	0.039 6	5	SDC4 PFKFB3 IFRD1 PMP22 HES6
		SP1F_EBOX_SP1F_01	0.085 4	4	MYLK4 PMP22 PDK4 CEBPD
		RUSH_EGRF_01	0.049 3	3	SDC4 GADD45A SPOCK2
		IRFF_NFAT_01	0.084 3	3	MYLK4 ADAMTS9 IFRD1
		GATA_GATA_GATA_01 0.013 4	2	SLC16A6 ADAMTS9
		MYOD_MYOD_03	0.066 3	2	SPOCK2 HES6
		AP1F_ETSF_04	0.013 2	2	IFRD1 HYAL2
		YY1F_SRFF_02	0.028 2	2	ATF3 FOS
		ZFHX_ZFHX_NKXH_01 0.037 2	2	GADD45A ADAMTS9

Table 2

 2

	(continued )
	Muscle Module	p	Occurrence	Number of	Gene ID
		value	of module	target genes
	SMAD_HIFF_01	0.032 2	1	PFKFB3
	SP1F_MZF1_01	0.035 2	1	PMP22
	ETSF_SP1F_SMAD_01	0.016 1	1	HEYL
	SRFF_AP1F_01	0.024 1	1	FOS
	YY1F_SRFF_01	0.039 1	1	FOS
	PAX8_NKXH_01	0.039 1	1	PMP22
	ETSF_SRFF_01	0.047 1	1	FOS
	MEF2_MYOD_01	0.054 1	1	SLC16A6
	KLFS_CREB_KLFS_01	0.070 1	1	SLC2A8
	CAAT_SREB_01	0.077 1	1	IFRD1
	GATA_HNF1_02	0.077 1	1	PDK4

Notes:

LT: Longissimus thoracis muscle; ST: Semitendinosus muscle. Modules in bold were in common between muscles. TFs genes are underlined.

Table 3

 3 Proposed as a protein biomarker for high ultimate pH (pHu) meat in[START_REF] Sentandreu | Protein biomarkers of bovine defective meats at a glance: gel-free hybrid quadrupole-orbitrap analysis for rapid screening[END_REF].

	(continued )
	Type of response Gene	Transcription	Location in a bovine QTL
		name	regulator
		INS
		MAPK1		Shear force
		MAPK14
		TGFB1
		TNF		Marbling score
	Common main	BCL2
	targets	BGLAP
		CDKN1A
		COL3A1
		ERBB2		Muscle pH, Marbling score
		FN1
		ICAM1		Marbling score
		IL6		Marbling score
		MMP2
		PPARG	TF
		SELE		Juiciness
		SLC2A4
		TLR4		Shear force
		VEGFA
	Notes:
	Only QTL related to meat quality are shown in the table.

*

Cassar-Malek et al. (2022), PeerJ, DOI 10.7717/peerj.13150

ACKNOWLEDGEMENTS

The authors thank Geneviève Gentes and the HELIXIO Company (formerly Imaxio, Clermont-Limagne Biopôle) for performing the transcriptomic analyses and Dominique Bauchart, Denis Durand, Claudia Terlouw, Brigitte Picard and Joelle Henry-Berger for scientific discussions.

Data Availability

The following information was supplied regarding data availability:

The raw data are available in the Supplemental File. The transcriptomic datasets are available at GEO: GSE119912.

Grant Disclosures

The following grant information was disclosed by the authors: French National Agency for Research. 6th Framework Programme of the European Union: FOOD-CT-2006-36241.

Competing Interests

The authors declare that they have no competing interests.

Author Contributions

Isabelle Cassar-Malek conceived and designed the experiments, performed the experiments, analyzed the data, prepared figures and/or tables, authored or reviewed drafts of the paper, and approved the final draft.

Lise Pomiès analyzed the data, prepared figures and/or tables, authored or reviewed drafts of the paper, and approved the final draft. Anne de la Foye analyzed the data, authored or reviewed drafts of the paper, and approved the final draft. Jérémy Tournayre analyzed the data, authored or reviewed drafts of the paper, and approved the final draft. Céline Boby performed the experiments, analyzed the data, prepared figures and/or tables, authored or reviewed drafts of the paper, and approved the final draft. Jean-François Hocquette conceived and designed the experiments, performed the experiments, analyzed the data, authored or reviewed drafts of the paper, and approved the final draft.

Animal Ethics

The following information was supplied relating to ethical approvals (i.e., approving body and any reference numbers):

The experiment was conducted in 2007. Before 2013 submission to the Ethics Committee was not mandatory in France. However, the Animal Facility of INRAE is approved for animal experimentation and that the scientists and technicians were authorized for animal experimentation. Experimental procedures and animal holding facilities respected French animal protection legislation, including licensing of experimenters. French Veterinary Services controlled and approved them (B63 345 17).

Supplemental Information

Supplemental information for this article can be found online at http://dx.doi.org/10.7717/ peerj.13150#supplemental-information.