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Two-hybrid screening

[[Image:Two hybrid assay.svg|thumb|300px| Overview of two-hybrid assay, checking for interactions between two proteins, called here Bait and Prey.

A. The Gal4 transcription factor gene produces a two-domain protein (BD and AD) essential for transcription of the reporter gene (LacZ).

B,C. Two fusion proteins are prepared: Gal4BD+Bait and Gal4AD+Prey. Neither of them are usually sufficient to initiate transcription (of the reporter gene) alone.

D. When both fusion proteins are produced and the Bait part of the first fusion protein interacts with the Prey part of the second, transcription of the reporter gene occurs. ]]

Two-hybrid screening (originally known as yeast two-hybrid system or Y2H) is a molecular biology technique used to discover protein–protein interactions (PPIs) and protein–DNA interactions by testing for physical interactions (such as binding) between two proteins or a single protein and a DNA molecule, respectively.

The premise behind the test is the activation of downstream reporter gene(s) by the binding of a transcription factor onto an upstream activating sequence (UAS). For two-hybrid screening, the transcription factor is split into two separate fragments, called the DNA-binding domain (DBD or often also abbreviated as BD) and activating domain (AD). The BD is the domain responsible for binding to the UAS and the AD is the domain responsible for the activation of transcription. The Y2H is thus a protein-fragment complementation assay.

History
Pioneered by Stanley Fields and Ok-Kyu Song in 1989, the technique was originally designed to detect protein–protein interactions using the Gal4 transcriptional activator of the yeast Saccharomyces cerevisiae. The Gal4 protein activated transcription of a gene involved in galactose utilization, which formed the basis of selection. Since then, the same principle has been adapted to describe many alternative methods, including some that detect protein–DNA interactions or DNA-DNA interactions, as well as methods that use different host organisms such as Escherichia coli or mammalian cells instead of yeast.

Basic premise
The key to the two-hybrid screen is that in most eukaryotic transcription factors, the activating and binding domains are modular and can function in proximity to each other without direct binding. This means that even though the transcription factor is split into two fragments, it can still activate transcription when the two fragments are indirectly connected.

The most common screening approach is the yeast two-hybrid assay. In this approach the researcher knows where each prey is located on the used medium (agar plates). Millions of potential interactions in several organisms have been screened in the latest decade using high-throughput screening systems (often using robots) and over thousands of interactions have been detected and categorized in databases as BioGRID. This system often utilizes a genetically engineered strain of yeast in which the biosynthesis of certain nutrients (usually amino acids or nucleic acids) is lacking. When grown on media that lacks these nutrients, the yeast fail to survive. This mutant yeast strain can be made to incorporate foreign DNA in the form of plasmids. In yeast two-hybrid screening, separate bait and prey plasmids are simultaneously introduced into the mutant yeast strain or a mating strategy is used to get both plasmids in one host cell.

The second high-throughput approach is the library screening approach. In this set up the bait and prey harboring cells are mated in a random order. After mating and selecting surviving cells on selective medium the scientist will sequence the isolated plasmids to see which prey (DNA sequence) is interacting with the used bait. This approach has a lower rate of reproducibility and tends to yield higher amounts of false positives compared to the matrix approach.

Plasmids are engineered to produce a protein product in which the DNA-binding domain (BD) fragment is fused onto a protein while another plasmid is engineered to produce a protein product in which the activation domain (AD) fragment is fused onto another protein. The protein fused to the BD may be referred to as the bait protein, and is typically a known protein the investigator is using to identify new binding partners. The protein fused to the AD may be referred to as the prey protein and can be either a single known protein or a library of known or unknown proteins. In this context, a library may consist of a collection of protein-encoding sequences that represent all the proteins expressed in a particular organism or tissue, or may be generated by synthesising random DNA sequences. Regardless of the source, they are subsequently incorporated into the protein-encoding sequence of a plasmid, which is then transfected into the cells chosen for the screening method. This technique, when using a library, assumes that each cell is transfected with no more than a single plasmid and that, therefore, each cell ultimately expresses no more than a single member from the protein library.

If the bait and prey proteins interact (i.e., bind), then the AD and BD of the transcription factor are indirectly connected, bringing the AD in proximity to the transcription start site and transcription of reporter gene(s) can occur. If the two proteins do not interact, there is no transcription of the reporter gene. In this way, a successful interaction between the fused protein is linked to a change in the cell phenotype.

The challenge of separating cells that express proteins that happen to interact with their counterpart fusion proteins from those that do not, is addressed in the following section.

Fixed domains
In any study, some of the protein domains, those under investigation, will be varied according to the goals of the study whereas other domains, those that are not themselves being investigated, will be kept constant. For example, in a two-hybrid study to select DNA-binding domains, the DNA-binding domain, BD, will be varied while the two interacting proteins, the bait and prey, must be kept constant to maintain a strong binding between the BD and AD. There are a number of domains from which to choose the BD, bait and prey and AD, if these are to remain constant. In protein–protein interaction investigations, the BD may be chosen from any of many strong DNA-binding domains such as Zif268. A frequent choice of bait and prey domains are residues 263–352 of yeast Gal11P with a N342V mutation and residues 58–97 of yeast Gal4, respectively. These domains can be used in both yeast- and bacterial-based selection techniques and are known to bind together strongly.

The AD chosen must be able to activate transcription of the reporter gene, using the cell's own transcription machinery. Thus, the variety of ADs available for use in yeast-based techniques may not be suited to use in their bacterial-based analogues. The herpes simplex virus-derived AD, VP16 and yeast Gal4 AD have been used with success in yeast whilst a portion of the α-subunit of E. coli RNA polymerase has been utilised in E. coli-based methods.

Whilst powerfully activating domains may allow greater sensitivity towards weaker interactions, conversely, a weaker AD may provide greater stringency.

Construction of expression plasmids
A number of engineered genetic sequences must be incorporated into the host cell to perform two-hybrid analysis or one of its derivative techniques. The considerations and methods used in the construction and delivery of these sequences differ according to the needs of the assay and the organism chosen as the experimental background.

There are two broad categories of hybrid library: random libraries and cDNA-based libraries. A cDNA library is constituted by the cDNA produced through reverse transcription of mRNA collected from specific cells of types of cell. This library can be ligated into a construct so that it is attached to the BD or AD being used in the assay. A random library uses lengths of DNA of random sequence in place of these cDNA sections. A number of methods exist for the production of these random sequences, including cassette mutagenesis. Regardless of the source of the DNA library, it is ligated into the appropriate place in the relevant plasmid/phagemid using the appropriate restriction endonucleases.

E. coli-specific considerations
By placing the hybrid proteins under the control of IPTG-inducible lac promoters, they are expressed only on media supplemented with IPTG. Further, by including different antibiotic resistance genes in each genetic construct, the growth of non-transformed cells is easily prevented through culture on media containing the corresponding antibiotics. This is particularly important for counter selection methods in which a lack of interaction is needed for cell survival.

The reporter gene may be inserted into the E. coli genome by first inserting it into an episome, a type of plasmid with the ability to incorporate itself into the bacterial cell genome with a copy number of approximately one per cell.

The hybrid expression phagemids can be electroporated into E. coli XL-1 Blue cells which after amplification and infection with VCS-M13 helper phage, will yield a stock of library phage. These phage will each contain one single-stranded member of the phagemid library.

Recovery of protein information
Once the selection has been performed, the primary structure of the proteins which display the appropriate characteristics must be determined. This is achieved by retrieval of the protein-encoding sequences (as originally inserted) from the cells showing the appropriate phenotype.

E. coli
The phagemid used to transform E. coli cells may be "rescued" from the selected cells by infecting them with VCS-M13 helper phage. The resulting phage particles that are produced contain the single-stranded phagemids and are used to infect XL-1 Blue cells. The double-stranded phagemids are subsequently collected from these XL-1 Blue cells, essentially reversing the process used to produce the original library phage. Finally, the DNA sequences are determined through dideoxy sequencing.

Controlling sensitivity
The Escherichia coli-derived Tet-R repressor can be used in line with a conventional reporter gene and can be controlled by tetracycline or doxicycline (Tet-R inhibitors). Thus the expression of Tet-R is controlled by the standard two-hybrid system but the Tet-R in turn controls (represses) the expression of a previously mentioned reporter such as HIS3, through its Tet-R promoter. Tetracycline or its derivatives can then be used to regulate the sensitivity of a system utilising Tet-R.

Sensitivity may also be controlled by varying the dependency of the cells on their reporter genes. For example, this may be affected by altering the concentration of histidine in the growth medium for his3-dependent cells and altering the concentration of streptomycin for aadA dependent cells. Selection-gene-dependency may also be controlled by applying an inhibitor of the selection gene at a suitable concentration. 3-Amino-1,2,4-triazole (3-AT) for example, is a competitive inhibitor of the HIS3-gene product and may be used to titrate the minimum level of HIS3 expression required for growth on histidine-deficient media.

Sensitivity may also be modulated by varying the number of operator sequences in the reporter DNA.

Non-fusion proteins
A third, non-fusion protein may be co-expressed with two fusion proteins. Depending on the investigation, the third protein may modify one of the fusion proteins or mediate or interfere with their interaction.

Co-expression of the third protein may be necessary for modification or activation of one or both of the fusion proteins. For example, S. cerevisiae possesses no endogenous tyrosine kinase. If an investigation involves a protein that requires tyrosine phosphorylation, the kinase must be supplied in the form of a tyrosine kinase gene.

The non-fusion protein may mediate the interaction by binding both fusion proteins simultaneously, as in the case of ligand-dependent receptor dimerization.

For a protein with an interacting partner, its functional homology to other proteins may be assessed by supplying the third protein in non-fusion form, which then may or may not compete with the fusion-protein for its binding partner. Binding between the third protein and the other fusion protein will interrupt the formation of the reporter expression activation complex and thus reduce reporter expression, leading to the distinguishing change in phenotype.

Split-ubiquitin yeast two-hybrid
One limitation of classic yeast two-hybrid screens is that they are limited to soluble proteins. It is therefore impossible to use them to study the protein–protein interactions between insoluble integral membrane proteins. The split-ubiquitin system provides a method for overcoming this limitation. In the split-ubiquitin system, two integral membrane proteins to be studied are fused to two different ubiquitin moieties: a C-terminal ubiquitin moiety ("Cub", residues 35–76) and an N-terminal ubiquitin moiety ("Nub", residues 1–34). These fused proteins are called the bait and prey, respectively. In addition to being fused to an integral membrane protein, the Cub moiety is also fused to a transcription factor (TF) that can be cleaved off by ubiquitin specific proteases. Upon bait–prey interaction, Nub and Cub-moieties assemble, reconstituting the split-ubiquitin. The reconstituted split-ubiquitin molecule is recognized by ubiquitin specific proteases, which cleave off the transcription factor, allowing it to induce the transcription of reporter genes.

Fluorescent two-hybrid assay
Zolghadr and co-workers presented a fluorescent two-hybrid system that uses two hybrid proteins that are fused to different fluorescent proteins as well as LacI, the lac repressor. The structure of the fusion proteins looks like this: FP2-LacI-bait and FP1-prey where the bait and prey proteins interact and bring the fluorescent proteins (FP1 = GFP, FP2=mCherry) in close proximity at the binding site of the LacI protein in the host cell genome. The system can also be used to screen for inhibitors of protein–protein interactions.

Enzymatic two-hybrid systems: KISS
While the original Y2H system used a reconstituted transcription factor, other systems create enzymatic activities to detect PPIs. For instance, the KInase Substrate Sensor ("KISS"), is a mammalian two-hybrid approach has been designed to map intracellular PPIs. Here, a bait protein is fused to a kinase-containing portion of TYK2 and a prey is coupled to a gp130 cytokine receptor fragment. When bait and prey interact, TYK2 phosphorylates STAT3 docking sites on the prey chimera, which ultimately leads to activation of a reporter gene.

One-hybrid
The one-hybrid variation of this technique is designed to investigate protein–DNA interactions and uses a single fusion protein in which the AD is linked directly to the binding domain. The binding domain in this case however is not necessarily of fixed sequence as in two-hybrid protein–protein analysis but may be constituted by a library. This library can be selected against the desired target sequence, which is inserted in the promoter region of the reporter gene construct. In a positive-selection system, a binding domain that successfully binds the UAS and allows transcription is thus selected.

Note that selection of DNA-binding domains is not necessarily performed using a one-hybrid system, but may also be performed using a two-hybrid system in which the binding domain is varied and the bait and prey proteins are kept constant.

Three-hybrid

RNA-protein interactions have been investigated through a three-hybrid variation of the two-hybrid technique. In this case, a hybrid RNA molecule serves to adjoin together the two protein fusion domains—which are not intended to interact with each other but rather the intermediary RNA molecule (through their RNA-binding domains). Techniques involving non-fusion proteins that perform a similar function, as described in the 'non-fusion proteins' section above, may also be referred to as three-hybrid methods.

One-two-hybrid
Simultaneous use of the one- and two-hybrid methods (that is, simultaneous protein–protein and protein–DNA interaction) is known as a one-two-hybrid approach and expected to increase the stringency of the screen.

Host organism
Although theoretically, any living cell might be used as the background to a two-hybrid analysis, there are practical considerations that dictate which is chosen. The chosen cell line should be relatively cheap and easy to culture and sufficiently robust to withstand application of the investigative methods and reagents. The latter is especially important for doing high-throughput studies. Therefore the yeast S. cerevisiae has been the main host organism for two-hybrid studies. However it is not always the ideal system to study interacting proteins from other organisms. Yeast cells often do not have the same post translational modifications, have a different codon use or lack certain proteins that are important for the correct expression of the proteins. To cope with these problems several novel two-hybrid systems have been developed. Depending on the system used agar plates or specific growth medium is used to grow the cells and allow selection for interaction. The most common used method is the agar plating one where cells are plated on selective medium to see of interaction takes place. Cells that have no interaction proteins should not survive on this selective medium.

S. cerevisiae (yeast)
The yeast S. cerevisiae was the model organism used during the two-hybrid technique's inception. It is commonly known as the Y2H system. It has several characteristics that make it a robust organism to host the interaction, including the ability to form tertiary protein structures, neutral internal pH, enhanced ability to form disulfide bonds and reduced-state glutathione among other cytosolic buffer factors, to maintain a hospitable internal environment. The yeast model can be manipulated through non-molecular techniques and its complete genome sequence is known. Yeast systems are tolerant of diverse culture conditions and harsh chemicals that could not be applied to mammalian tissue cultures.

A number of yeast strains have been created specifically for Y2H screens, e.g. Y187 and AH109, both produced by Clontech. Yeast strains R2HMet and BK100 have also been used.

Candida albicans
C. albicans is a yeast with a particular feature: it translates the CUG codon into serine rather than leucine. Due to this different codon usage it is difficult to use the model system S. cerevisiae as a Y2H to check for protein-protein interactions using C. albicans genes. To provide a more native environment a C. albicans two-hybrid (C2H) system was developed. With this system protein-protein interactions can be studied in C. albicans itself. A recent addition was the creation of a high-throughput system.

E. coli
Bacterial two hybrid methods (B2H or BTH) are usually carried out in E. coli and have some advantages over yeast-based systems. For instance, the higher transformation efficiency and faster rate of growth lends E. coli to the use of larger libraries (in excess of 108). The absence of requirements for a nuclear localisation signal to be included in the protein sequence and the ability to study proteins that would be toxic to yeast may also be major factors to consider when choosing an experimental background organism.

The methylation activity of certain E. coli DNA methyltransferase proteins may interfere with some DNA-binding protein selections. If this is anticipated, the use of an E. coli strain that is defective for a particular methyltransferase may be an obvious solution. The B2H may not be ideal when studying eukaryotic protein-protein interactions (e.g. human proteins) as proteins may not fold as in eukaryotic cells or may lack other processing.

Mammalian cells
In recent years a mammalian two hybrid (M2H) system has been designed to study mammalian protein-protein interactions in a cellular environment that closely mimics the native protein environment. Transiently transfected mammalian cells are used in this system to find protein-protein interactions. Using a mammalian cell line to study mammalian protein-protein interactions gives the advantage of working in a more native context. The post-translational modifications, phosphorylation, acylation and glycosylation are similar. The intracellular localization of the proteins is also more correct compared to using a yeast two hybrid system. It is also possible with the mammalian two-hybrid system to study signal inputs. Another big advantage is that results can be obtained within 48 hours after transfection.

Arabidopsis thaliana
In 2005 a two hybrid system in plants was developed. Using protoplasts of A. thaliana protein-protein interactions can be studied in plants. This way the interactions can be studied in their native context. In this system the GAL4 AD and BD are under the control of the strong 35S promoter. Interaction is measured using a GUS reporter. In order to enable a high-throughput screening the vectors were made gateway compatible. The system is known as the protoplast two hybrid (P2H) system.

Aplysia californica
The sea hare A californica is a model organism in neurobiology to study among others the molecular mechanisms of long-term memory. To study interactions, important in neurology, in a more native environment a two-hybrid system has been developed in A californica neurons. A GAL4 AD and BD are used in this system.

Bombyx mori
An insect two-hybrid (I2H) system was developed in a silkworm cell line from the larva or caterpillar of the domesticated silk moth, Bombyx mori (BmN4 cells). This system uses the GAL4 BD and the activation domain of mouse NF-κB P65. Both are under the control of the OpIE2 promoter.

Determination of sequences crucial for interaction
By changing specific amino acids by mutating the corresponding DNA base-pairs in the plasmids used, the importance of those amino acid residues in maintaining the interaction can be determined.

After using bacterial cell-based method to select DNA-binding proteins, it is necessary to check the specificity of these domains as there is a limit to the extent to which the bacterial cell genome can act as a sink for domains with an affinity for other sequences (or indeed, a general affinity for DNA).

Drug and poison discovery
Protein–protein signalling interactions pose suitable therapeutic targets due to their specificity and pervasiveness. The random drug discovery approach uses compound banks that comprise random chemical structures, and requires a high-throughput method to test these structures in their intended target.

The cell chosen for the investigation can be specifically engineered to mirror the molecular aspect that the investigator intends to study and then used to identify new human or animal therapeutics or anti-pest agents.

Determination of protein function
By determination of the interaction partners of unknown proteins, the possible functions of these new proteins may be inferred. This can be done using a single known protein against a library of unknown proteins or conversely, by selecting from a library of known proteins using a single protein of unknown function.

Zinc finger protein selection
To select zinc finger proteins (ZFPs) for protein engineering, methods adapted from the two-hybrid screening technique have been used with success. A ZFP is itself a DNA-binding protein used in the construction of custom DNA-binding domains that bind to a desired DNA sequence.

By using a selection gene with the desired target sequence included in the UAS, and randomising the relevant amino acid sequences to produce a ZFP library, cells that host a DNA-ZFP interaction with the required characteristics can be selected. Each ZFP typically recognises only 3–4 base pairs, so to prevent recognition of sites outside the UAS, the randomised ZFP is engineered into a 'scaffold' consisting of another two ZFPs of constant sequence. The UAS is thus designed to include the target sequence of the constant scaffold in addition to the sequence for which a ZFP is selected.

A number of other DNA-binding domains may also be investigated using this system.

Strengths

 * Two-hybrid screens are low-tech; they can be carried out in any lab without sophisticated equipment.
 * Two-hybrid screens can provide an important first hint for the identification of interaction partners.
 * The assay is scalable, which makes it possible to screen for interactions among many proteins. Furthermore, it can be automated, and by using robots many proteins can be screened against thousands of potentially interacting proteins in a relatively short time. Two types of large screens are used: the library approach and the matrix approach.
 * Yeast two-hybrid data can be of similar quality to data generated by the alternative approach of coaffinity purification followed by mass spectrometry (AP/MS).

Weaknesses
The reason for this high error rate lies in the characteristics of the screen:
 * The main criticism applied to the yeast two-hybrid screen of protein–protein interactions are the possibility of a high number of false positive (and false negative) identifications. The exact rate of false positive results is not known, but earlier estimates were as high as 70%. This also, partly, explains the often found very small overlap in results when using a (high throughput) two-hybrid screening, especially when using different experimental systems.
 * Certain assay variants overexpress the fusion proteins which may cause unnatural protein concentrations that lead to unspecific (false) positives.
 * The hybrid proteins are fusion proteins; that is, the fused parts may inhibit certain interactions, especially if an interaction takes place at the N-terminus of a test protein (where the DNA-binding or activation domain is typically attached).
 * An interaction may not happen in yeast, the typical host organism for Y2H. For instance, if a bacterial protein is tested in yeast, it may lack a chaperone for proper folding that is only present in its bacterial host. Moreover, a mammalian protein is sometimes not correctly modified in yeast (e.g., missing phosphorylation), which can also lead to false results.
 * The Y2H takes place in the nucleus. If test proteins are not localized to the nucleus (because they have other localization signals) two interacting proteins may be found to be non-interacting.
 * Some proteins might specifically interact when they are co-expressed in the yeast, although in reality they are never present in the same cell at the same time. However, in most cases it cannot be ruled out that such proteins are indeed expressed in certain cells or under certain circumstances.

Each of these points alone can give rise to false results. Due to the combined effects of all error sources yeast two-hybrid have to be interpreted with caution. The probability of generating false positives means that all interactions should be confirmed by a high confidence assay, for example co-immunoprecipitation of the endogenous proteins, which is difficult for large scale protein–protein interaction data. Alternatively, Y2H data can be verified using multiple Y2H variants or bioinformatics techniques. The latter test whether interacting proteins are expressed at the same time, share some common features (such as gene ontology annotations or certain network topologies), have homologous interactions in other species.

title: Two-hybrid screening


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Wikipedia talk:Articles for deletion/Peter A Barnard

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 * Osiris, that's not the point. Even if Mr. Barnard does not mind, Wikipedia is not the place for school-kid fun. If you want to start your own wiki to play around, you can create one at Wikicities. jglc | t | c 19:48, 9 Jun 2005 (UTC)

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Talk:Jože Plečnik

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RT-21 Temp 2S

The RT-21 Temp 2S (Темп-2С) was a mobile intercontinental ballistic missile developed by the Soviet Union during the Cold War. It was assigned the NATO reporting name SS-16 Sinner and carried the industry designation 15Zh42 (15Ж42).

The RT-21 was the first mobile ICBM developed in the world. Its innovative concept and design were created by Alexander Nadiradze. The RT-21M Pioner and succeeding missile complexes relied on the RT-21 base concept and were used by Nadiradze for many of his later projects. The program became mired in a series of treaty complications, including questions regarding its use of theatre missile launchers. It is unlikely that the RT-21 ultimately reached deployment, and by the mid-1980s, the program had been scrapped. Its maximum period of storage on a launcher was 5 years, and preparation time for launch was 40 minutes.

title: RT-21 Temp 2S


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WA Reds

The Western Reds were a rugby league football club based in Perth, Western Australia. Founded in 1992 as the Western Reds, they entered into the Australian Rugby League competition in 1995 before defecting to the rival Super League competition in 1997, where they rebranded themselves as the Perth Reds. However, by the end of the year the Reds had become a casualty of the Super League War peace deal and were shut down. The name Reds was named after the native Red Kangaroos. The Reds entered a state of limbo for the next decade but were revived as a lower-level club in 2006 by the WARL and ARL, under the name WA Reds.

Western Reds (1992–1996)
The Reds had recruited well in 1993–94, and signed Peter Mulholland as their first coach. St. George fullback Michael Potter (twice winner of the Dally M Award), 1992 Rookie of year and CLEO bachelor of the year Matthew Rodwell, and Australian and NSW rep player Brad Mackay were three of the major signings.

Their first game, played at the WACA, was watched by a record 24,392, with the Reds defeating St George 28–16. When News Limited began its "blitzkrieg" in April 1995, the Reds aligned themselves with Super League along with nine other Australian Rugby League clubs. That season the Reds were the best performing of the three expansion teams introduced, winning 11 of their 22 games, including 8 at home, which drew an average crowd of around 13,000, larger than that of many Sydney teams.

The Reds even recruited local identity and decorated WAFL and West Coast Eagles (AFL) player Adrian Barich. Barich, having grown up in Canberra reverted from Australian rules football to rugby league, finishing his career with seasons with the Reds, however he never played in first grade.

After declaring that the club may not have the financial resources to compete in the 1996 ARL Optus Cup, a major sponsorship with the Rupert Murdoch-owned Sunday Times gave the club some much needed money. By mid 1996 club support had dwindled to just over 6,000.

Perth Reds (1997)
In 1997 the Reds became one of eight ARL teams to join the rival Super League during the dispute known as the Super League war. They changed their name to the Perth Reds for the 1997 Super League season, and adopted a jersey of red, white and black. Although the club had made some promising signings, such as Rodney Howe and Robbie Kearns, the crushing $10 million debt that hung over the club (from having to pay the airfares for all visiting teams) at the end of the season led to Super League axing the Perth Reds on 1 October 1997.

Hiatus (1998–2005)
Rugby league has continued to be played in Western Australia since the Reds left the top-flight competition with the Swan Brewery Cup continuing and NRL matches being staged at various times since 1998. On 8 May 1999, Melbourne Storm played Western Suburbs Magpies at Lathlain Park in Perth, with Melbourne running out winners 62–6. In 2005, Cronulla took their home game against the New Zealand Warriors to Perth Oval, and played in front of around 13,000 spectators.

WA Reds (2006–2011)
The WARL resurrected the Reds in 2006 with the intent of joining the National Rugby League in the future. In 2008 the Reds joined the Jim Beam Cup and played out of Perth Oval.

In the 2009 Bundaberg Red Cup the Reds were winless until the back-end of the season when they won 3 of their 4 last games, giving them something to build on for the next season. Darwin Rugby League product Aaron Barnes was named the Reds' player of the year. Unfortunately for financial reasons the Reds snr team was withdrawn from the Bundy Cup for 2010.

In 2010 the WA Reds entered an Under 18's team in the S. G. Ball Cup competition. The first trial game on 30 January 2010 was against the runners-up of the Western Australia Rugby League competition the Central Bulldogs – they won 38–20. The WARL have set up two junior academies to help develop players for a return to the NRL with the SG Ball side providing a staging ground for player development. The WA Reds juniors won their first SG Ball game beating the Balmain Tigers at ME Stadium 28–8.

Whilst the team found it tough going against often bigger and more experienced sides they did claim a couple of scalps and at the end of the season Curtis Rona was signed by the Sydney Roosters, earning a call up to the under 20s side during 2010.

A new bid logo was launched in 2010 and membership for fans was made available. NRL CEO David Gallop recognised the WA Reds 2013 bid on a number of occasions, speaking positively about the potential for a Perth team in the NRL, but as of now, no such offer has become official.

West Coast Pirates (2012–present)
In 2012 the WARL launched the West Coast Pirates as the Perth bid team for an NRL licence. As of 2012 West Coast competes in the SG Ball Cup instead of the WA Reds.

Players

 * 1995 U/21 Reds
 * Jade Koteka
 * David Hignett
 * Loa Tupou
 * Paul Muller
 * Darren Mullholland
 * Duncan Arkley
 * Will Bramwell
 * Ward Denman
 * Matt Geyer
 * Andrew Lippiat
 * Chris Scarisbrick
 * Leon Ruri
 * Aaron Ritchie
 * Nick Twiddle
 * David Huber
 * Shaun Owen
 * Brock McDonald
 * Myles Ritchie
 * Wayne Blazey
 * Brett Hyland
 * Brian McCarthey
 * Graeme Shield
 * Jarred Millar
 * Chris Lott
 * Jason Edmunds (Captain)
 * Tony Hemana
 * Byron Hutton
 * Ryan Gundry
 * Adam Beard
 * Dwayne Evans
 * Daniel Cross
 * Michael Jackson
 * Dane Dorahy
 * Ryan Dissegna
 * Sean Edmunds

 * 1st Grade
 * Mark Anderson
 * Simon Kricheldorff
 * Mark Geyer
 * Matt Geyer
 * Rodney Howe
 * Robbie Kearns
 * Brad Mackay
 * Barrie-Jon Mather
 * Simon Robbie
 * Scott Wilson
 * David Dowell
 * Shayne McMenemy
 * Danny Davies

WA Reds (2006–2011)

 * Liam Mulhall
 * Savi Hafoka
 * Pikari Te Wara
 * Clae Morgan
 * Heath Egglestone
 * Ryan Dickson
 * Matt Doeg
 * James Blake
 * Issac Thomas
 * John Phipps
 * James McGowan
 * Delane Edwards
 * Taurean Sheehan
 * Joel Freeman
 * Gareth Morton
 * Aaron Barnes
 * Nathan Searle
 * Carlin Miller
 * Michael Elphick
 * Ben McCrone
 * Frank Matthewman

 * Recent and current NRL WARL-produced players
 * Bryson Goodwin (South Sydney Rabbitohs)
 * Bronx Goodwin (St. George Illawarra Dragons)
 * Cory Paterson (Newcastle Knights)
 * Jon Green (St. George Illawarra Dragons)
 * Daniel Holdsworth (Salford City Reds)
 * Matt Petersen (Wakefield Trinity Wildcats)
 * Lee Te Maari (Parramatta Eels)
 * Curtis Rona (Canterbury Bulldogs)
 * Waqa Blake (Parramatta Eels)

Internationals

 * Rodney Howe (1997)
 * Julian O'Neill (1997)
 * Barrie-Jon Mather (1997)
 * Shayne McMenemy (2007)
 * Halvor Harris (2015)
 * Bradley Williams (2015)

Coaches

 * John Dorahy (U/21 1995)
 * Peter Mulholland (1995–96)
 * Dean Lance (1997)

Club records
Biggest Win
 * 28 points, 34–6 against Canterbury Bulldogs at Perth Oval on 23 March 1997.

Biggest Loss
 * 42 points, 0–42 against Sydney Bulldogs at Parramatta Stadium on 9 April 1995.

Most Consecutive Wins
 * 2 matches, 1–8 July 1995
 * 2 matches, 21–28 July 1995
 * 2 matches, 12–18 August 1995
 * 2 matches, 14–20 July 1996
 * 2 matches, 23–27 March 1997

Most Consecutive Loses
 * 9 matches, 13 April – 23 June 1996

Most First Grade Matches
 * 59 – Matthew "Matt" Fuller (1995–97)

Most First Grade Points
 * 210 – Chris Ryan (21 tries, 63 goals)

Most First Grade Tries
 * 21 – Chris Ryan

Most Points in a Season
 * 136 – Chris Ryan (9 tries, 50 goals)

Most Tries in a Season
 * 10 – Matthew "Matt" Fuller (1995)

Most Points in a Match
 * 26 – Julian O'Neill (4 tries, 5 goals. v Canterbury, 1997)

Most Tries in a Match
 * 4 – Julian O'Neill (v Canterbury, 1997)

Most Goals in a Match
 * 6 – Julian O'Neill (v Auckland, 1996)
 * 6 – Damien Chapman (v Penrith, 1997)

title: WA Reds


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Curtiss SOC Seagull

The Curtiss SOC Seagull was an American single-engined scout observation seaplane, designed by Alexander Solla of the Curtiss-Wright Corporation for the United States Navy. The aircraft served on battleships and cruisers in a seaplane configuration, being launched by catapult and recovered from a sea landing. The wings folded back against the fuselage for storage aboard ship. When based ashore or on carriers the single float was replaced by fixed wheeled landing gear.

Curtiss delivered 258 SOC aircraft, in versions SOC-1 through SOC-4, beginning in 1935. The SOC-3 design was the basis of the Naval Aircraft Factory SON-1 variant, of which the NAF delivered 64 aircraft from 1940.

The aircraft served as an important observation craft during WW2 for the U.S. Navy, although the Vought OS2U Kingfisher served in greater numbers.

Design and development
The SOC was ordered for production by the United States Navy in 1933 and first entered service in 1935. The first order was for 135 SOC-1 models, which was followed by 40 SOC-2 models for landing operations and 83 SOC-3s. A variant of the SOC-3 was built by the Naval Aircraft Factory and was known as the SON-1.

Operational history
The first ship the SOC was assigned to was the light cruiser USS Marblehead (CL-12) in November 1935; by the end of the decade, the SOC had replaced its predecessor throughout the fleet. Production came to an end in 1938. By 1941, most battleships had transitioned to the Vought OS2U Kingfisher and cruisers were expected to replace their aging SOCs with the third generation SO3C Seamew. The SO3C, however, suffered from a weak engine and plans to adopt it as a replacement were scrapped. The SOC, despite belonging to an earlier generation, went on to execute its missions of gunfire observation and limited range scouting missions.

Through the first six months of naval service, the SOC was known as the XO3C-1, The designation was changed to SOC when it was decided to merge its scouting and observation roles. The SOC was not called the Seagull until 1941, when the U.S. Navy began the wholesale adoption of popular names for aircraft in addition to their alpha-numeric designations. The name 'Seagull' had earlier been given to two civil Curtiss aircraft, a Curtiss Model 18 and a Model 25, both converted Curtiss MF flying boats.

When operating as a seaplane, returning SOCs would land on the relatively smooth ocean surface created on the sheltered side of the vessel as it made a wide turn, after which the aircraft would be winched back onto the deck.

When the SOC was replaced by the OS2U Kingfisher, most remaining airframes were converted into trainers; they remained in use until 1945. With the failure of the Curtiss SO3C Seamew, many SOCs in second line service were returned to frontline units starting in late 1943. They saw service aboard warships in the combat zone for the rest of World War II. This is one of the few instances in aviation history in which an older aircraft type, that was retired or sent to second line service, replaced the new aircraft type that was intended to replace it.

In certain roles such as an observation aircraft for battleships, they served until 1949, and were eventually superseded by longer range radar and helicopters.

Variants

 * XO3C-1 (Curtiss Model 71): Prototype aircraft, powered by 550 hp (410 kW) Pratt & Whitney R-1340-12 engine. One built, redesignated XSOC-1 on 23 March 1935.
 * SOC-1 (Curtiss Model 71A): Initial production version, with 550 hp (410 kW) Pratt & Whitney R-1340-18 engine enclosed in NACA cowling. Interchangeable float and wheeled undercarriage. 135 built.
 * SOC-2 (Curtiss Model 71B): Minor changes, with R-1340-22 engine. 40 built. Wheeled undercarriage only.
 * XSO2C-1 (Curtiss Model 71C): Improved version. One prototype only, no production.
 * SOC-3 (Curtiss Model 71E): Similar to SOC-2, but with interchangeable undercarriage. 83 built by Curtiss as SOC-3 with further 64 built by the Naval Aircraft Factory as the SON-1.
 * SOC-3A: All SOC-4s were transferred to the U.S. Navy in 1942 (BuNo 48243, 48244, 48245, respectively), which modified them SOC-3A standard, meaning the fitting of a deck arrester gear.
 * SOC-4 (Curtiss Model 71F): The U.S. Coast Guard acquired the final three SOC-3 Seagulls produced by Curtiss in 1938 and these were designated as SOC-4s. They were assigned the USCG call numbers V171, V172, and V173.
 * SO2C: One built for evaluation based on the SOC-3, but with a 5-foot fuselage stretch and powered by a R-1340-35.
 * SON-1: SOC-3 aircraft produced by the Naval Aircraft Factory, 64 built.

Operators

 * 🇺🇸 United States
 * United States Navy
 * United States Marine Corps
 * United States Coast Guard

title: Curtiss SOC Seagull


text:
National Export Credits Guarantee Board (Sweden)

Swedish National Export Credits Guarantee Board (Exportkreditnämnden, EKN) is a Swedish government agency that answers to the Ministry for Foreign Affairs. The agency is located in Stockholm.

Its aim is to promote Swedish exports by issuing guarantees, functioning as insurances, by which the Government of Sweden assumes certain risks. The customers include export companies and banks.


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Alanna Nash

Alanna Kay Nash (born August 16, 1950) is an American journalist and biographer.

Born in Louisville, Kentucky, Nash holds a master's degree from the Columbia University Graduate School of Journalism and is the author of several acclaimed books. She is a 1972 graduate of Stephens College. A feature writer for The New York Times, Stereo Review, Entertainment Weekly, and USA Weekend, she was named the Society of Professional Journalists' National Member of the Year in 1994. In 1977, Nash's job afforded her the opportunity to become one of the journalists to view the remains of Elvis Presley. In her dust jacket biography for her book Baby, Let's Play House, she was described as "the first journalist to see Elvis Presley in his casket".

Dolly Parton
Nash's much-reprinted 1978 biography of Dolly Parton included material gathered from a long interview with its subject, that took place over the course of several days. The biography, Dolly, was published just as Parton reached mainstream appeal outside of the country music world, via her now-famous pop-music crossover. Nash has closely followed Parton's career since, having also written numerous magazine articles about her and reviewing a number of Parton's albums.

Other projects
In 1988 Nash received acclaim for her book on Jessica Savitch, Golden Girl: The Story of Jessica Savitch, which was the basis for the 1996 motion picture Up Close & Personal, as well as a made-for-television movie named Almost Golden. In the 1990s Nash began researching the life of Elvis Presley in order to write a book. Although there were already several hundred Presley books on the market, her 1995 book, Elvis Aaron Presley: Revelations from the Memphis Mafia, provided what Entertainment Weekly called "stunning allegations". As a result of Nash being able to get the collaboration of employees Marty Lacker, Lamar Fike, and Presley's first cousin, Billy Smith who lived and worked with Presley throughout his life, she provided a look at Presley not previously published.

Colonel Tom Parker
Her research into Presley led to a second book on Colonel Tom Parker. While covering Presley's death, Alanna Nash had seen what most of the throng of reporters there at the time considered as somewhat bizarre decorum by Parker when he came to the funeral dressed in a Hawaiian shirt and baseball cap. For her 1995 book, Nash had interviewed Colonel Parker but her examination into his life kept unfolding with so many twists and turns that it led to six years of exhaustive research including travel to his birthplace Breda in the Netherlands for documents and interviews. Her book, The Colonel: The Extraordinary Story of Colonel Tom Parker and Elvis Presley, was published on July 15, 2003, to acclaim. Billboard called it a "classic of music industry reporting". Other positive reviews came from The Washington Post, The New York Review of Books, Variety, and Publishers Weekly, among others. In the UK, Mojo music magazine said her book was "the most incisive and comprehensive look at the life of the elusive Colonel available" and the reviewer for The Observer lauded the book as "perhaps the most thoroughly researched music book ever written" — before adding that "sadly most of the story has been told before. Nash simply adds layer after layer of padding."

For her reporting on Colonel Parker, Nash was voted one of the "Heavy 100 of Country Music" by Esquire magazine and earned the 2004 CMA Media Achievement Award and the 2004 Belmont Book Award. She has reviewed for Stereo Review magazine, Reader's Digest, and Amazon.com as well as others.

title: Alanna Nash


text:
Nicotiana alata

Nicotiana alata is a species of tobacco. It is called jasmine tobacco, sweet tobacco, winged tobacco, tanbaku, and Persian tobacco.

Nictoiana alata is mainly grown as an ornamental plant; numerous cultivars and hybrids are derived from it. In Iran, narghila tobacco is sometimes produced from N. alata; it is not chopped like cigarette tobacco, but broken up by hand.

 * Flowering season: Summer to fall
 * Light requirements: Sun to partial shade
 * Flower color: Lime green, maroon red, white, yellow, pink, and crimson
 * Height: 30 to 60 cm [12 to 24 inches]
 * Spacing: 30 to 40 cm [12 to 14 inches]
 * Comments: Low drought tolerance

Has a lovely fragrance in the evening to night. Most varieties bloom late-afternoon to evening. Seedlings do best when gradually introduced to outdoor environment over the course of a week. Plants tend to self-sow. (Color may be the original or occasionally several colors on one plant). Deadhead spent flower stalks for best blooming results.

title: Nicotiana alata


text:
National Inspectorate of Strategic Products (Sweden)

Inspectorate of Strategic Products (Inspektionen för strategiska produkter) is a Swedish government agency that answers to the Ministry for Foreign Affairs. The agency is located in Stockholm.

The agency controls the export of military equipment and dual-use products, ie products that may have both a civilian and a military use.


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Brita (company)

Brita GmbH is a German manufacturer of water filters headquartered in Taunusstein near Wiesbaden, Hesse. The company's manufacturing facilities are located in China, Germany, Switzerland, Italy, and the United Kingdom. Brita products are distributed in 69 countries.

Products

Brita produces water jugs (BPA-free, made of styrene methyl methacrylate copolymer), kettles and tap attachments with integrated disposable filters. The filters can be recycled.

Their primary filtering mechanism consists of activated carbon and ion-exchange resin. The activated carbon is produced from coconut shells. According to Brita, the filters have two effects:
 * The activated carbon removes substances that may impair taste, such as chlorine and chlorine compounds.
 * The ion-exchange resin reduces the carbonate hardness (limescale) as well as copper and lead.

In the United Kingdom, Hinari and Breville sell electric kettles incorporating Brita technology.

History
Brita was founded in 1966 by Heinz Hankammer. The company was first named AquaDeMat before he later named the company after his daughter. The company secured the first patent for domestic water filter use in the 1970s.

In 1988, The Clorox Company, based in Oakland, California, entered in a licensing-and-distribution agreement with the German company for North and South America. In 1995, it added the Canadian rights by acquiring Canada's Brita International Holdings. In 2000, Clorox acquired the sole rights to the brand in the Americas and Brita agreed to a non-compete clause until 2005. In 2008, Brita returned to the North American market under the brand Mavea, only to withdraw again in 2016. However, products are still sold in the US under the Brita brand.

title: Brita (company)


text:
Remote job entry

Remote job entry, or Remote Batch, is the procedure for sending requests for non-interactive data processing tasks (jobs) to mainframe computers from remote workstations, and by extension the process of receiving the output from such jobs at a remote workstation.

The RJE workstation is called a remote because it usually is located some distance from the host computer. The workstation connects to the host through a modem, digital link, packet-switching network or local area network (LAN). RJE is similar to uux and SSH, except that the workstation sends a complete job stream rather than a single command and that the user does not receive any output until the completion of the job.. The terms Remote Batch, Remote Job System and Remote Job Processing are also used for RJE facilities.

Examples
Remote Job Entry (RJE) is also the name of an OS/360 component that provided RJE services. An RJE workstation operator may have complete console control of the job flow between the workstation and mainframe, depending on local configuration and policy.

Houston Automatic Spooling Priority (HASP) initially supported job entry from terminals using Synchronous transmit-receive (STR); eventually HASP II supported only Binary Synchronous Communications (BSC), and added the Multi-leaving protocol for BSC programmable work stations; this protocol is incompatible with that used by OS/360 RJE and is the basis for protocols used for job submission from programmable work stations for, e.g., Attached Support Processor (ASP), JES2, JES3, OS/VS1 Remote Entry Services (RES), VM RSCS, as well as the later protocols for Network Job Entry (NJE) in, e.g., JES2, JES3, VM RSCS.

Conversational Remote Job Entry (CRJE) is a component of OS/360 and OS/VS1 that provides job submission, job retrieval and editing for a user at an interactive terminal.

Remote Entry Services] (RES) is a component of OS/VS1 that provides RJE services. An RES workstation operator may have complete console control of the job flow between the workstation and mainframe, depending on local configuration and policy.

Remote Spooling Communications Subsystem (RSCS) is, depending on the release, a component of or extra cost product in VM that provides RJE services. The RSCS in the free VM/370 only supported BSC; IBM added support for Systems Network Architecture (SNA), NJE and TCP/IP over several chargeable versions.

Network Job Entry (NJE) is Store and forward networking for transmitting, e.g., card files, jobs, printed output, among peers. The initial versions of NJE for JES2, JES3, VSE POWER and VM RSCS used BSC multileaving, but IBM quickly added support for Channel-to-channel adapters. IBM later added support for SNA and, ultimately, TCP/IP.

NETRJS is the protocol developed by the Campus Computing Network at UCLA to deliver batch jobs to the Remote Job Service (RJS) on their IBM 360 Model 91. This protocol was originally assigned to ARPANET Initial Connection Protocol sockets 71, 73, and 75, and later reassigned to Internet ports 71–74. RJS is a subsystem of OS/360 MVS written by UCLA to support remote batch from card-reader/printer terminals.

RJE workstations
Early RJE workstations were "dumb" (non-programmable) devices using byte-synchronous communications protocols such as IBM BISYNC or STR, or equivalents from other vendors. Later, programmable devices or small computers were used, and IBM developed a protocol called HASP multileaving for use with HASP, and later, e.g., ASP, JES2, JES3, RSCS. The IBM System/360 Model 20 and 1130, the Mohawk Data MDS 2400 and the UNIVAC 1004, were popular. Later still RJE workstations switched to bit-oriented full duplex protocols such as IBM Synchronous Data Link Control, HDLC, or X.25. The Internet Engineering Task Force has defined RFCs for internet remote job entry protocols, but they are now considered obsolete or legacy.

The 200 USER Terminal is a remote batch terminal and protocol developed by the Control Data Corporation for their CDC 6000 series and CDC 3000 series mainframe computers in the 1960s. A 200 USER Terminal consisted of a low speed punched card reader, a line printer, and a CRT operators console. It typically communicated with a remote mainframe via synchronous modem. The software subsystem on the mainframe side was called Export-Import 200, and later, the Remote Batch Facility (RBF). Other remote batch terminals using the UT200 protocol included the CDC 731, 732, and 734. Software emulators for the UT200 protocol were also written for a number of minicomputer systems.

Network Job Entry
RJE is well suited to organizations that had a single large central computer center. However, in large organizations with multiple data centers, there was an interest in a peer-to-peer transfer of, e.g., submitted jobs, printer output. Following customer requests, IBM developed a suite of facilities, derived from BITNET and VNET, known as Network Job Entry. As part of that software, IBM provided commands to transmit datasets among nodes (complexes of computers with a collective name). NJE allows a batch job to control where it would run and where its output would be processed; similarly, NJE allows an interactive user to send printed or punched output to a different node.

IBM has integrated NJE facilities into its mainframe software, and it is no longer available as separate products. However, the NJE support in JES3 requires the Batch Data Transmission (BDT) program product and the NJE support in z/VM requires the Remote Spooling Communications Subsystem (RSCS) program product. NJE supports Binary Synchronous Communications (BSC), Channel-to-channel adapter (CTCA), Systems Network Architecture (SNA) and TCP/IP connections among its nodes.

title: Remote job entry


text:
Invest Sweden

Invest Sweden (Myndigheten för utländska investeringar i Sverige), a.k.a. Invest in Sweden Agency (Delegationen för Utländska Investeringar i Sverige) 1995–2010, was a Swedish government agency organised under the Minister for Foreign Affairs, promoting business and investment opportunities in Sweden to foreign investors. The objective was to stimulate an increased flow of foreign direct investment to Sweden.

In 2013, the agency was merged with the organisation Swedish Trade Council, to form a new organisation named Business Sweden.

title: Invest Sweden


text:
Ghislaine Dommanget

Ghislaine Marie Françoise Dommanget (13 October 1900 – 30 April 1991) was a French actress, and the Princess consort of Monaco from 1946 to 1949.

Early life
Dommanget was born in Reims, France, as the daughter of Colonel Robert Joseph Dommanget (1867–1957) and his wife, Marie Louise Meunier (1867–1957).

Personal life
In 1923, she was married to Paul Diey (1863–1931). After his death in 1931, she had a relationship with actor André Brulé.

She broke up with Brulé and married the reigning Prince of Monaco, Louis II, on 24 July 1946 — the first bride of a member of the House of Grimaldi without a dowry.

Following Louis II's death on 9 May 1949, and the accession of her step-grandson, Rainier III, she became Dowager Princess of Monaco, a title she held until her death.

Honours

 * National honours
 * 🇲🇨 Monaco: Dame Grand Cross of the Order of Saint-Charles (24 October 1946).
 * Foreign honours
 * 🇫🇷 France:
 * Knight of the Order of the Legion of Honour (25 June 1947).
 * Commander of the Order of Public Health (14 July 1948).
 * Holy See: Benemerenti Medal (8 January 1948).

title: Ghislaine Dommanget


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Fabian Nicieza

Fabian Nicieza (December 31, 1961) is an Argentine-American comic book writer and editor who is best known for his work on Marvel titles such as X-Men, X-Force, New Warriors, Nomad, Cable, Deadpool and Thunderbolts, for all of which he helped create numerous characters, among them Deadpool, Domino, Shatterstar, and Silhouette.

Early life
Nicieza was born in Buenos Aires, Argentina, the son of Omar and Irma Riguetti Nicieza. He was four years old when his family moved to the United States. Growing up in New Jersey, Nicieza learned to read and write from comic books. He lived first in Sayreville, New Jersey and moved to Old Bridge Township, where he attended Madison Central High School, from which he graduated in 1979. He studied at Rutgers University, interning at the ABC television network before graduating in 1983 with a degree in advertising and public relations. His brother is Mariano Nicieza, also a comic book writer and editor.

Career
Until 1985, Nicieza worked for the Berkley Publishing Group, starting in the production department and becoming a managing editor.

Marvel Comics
In 1985, Nicieza joined the staff at Marvel Comics, initially as a manufacturing assistant, later moving to the promotions department as an advertising manager. During this period he began to take his first freelance work for Marvel, writing short articles for Marvel's promotional magazine Marvel Age.

Nicieza's first published comics story came with Psi-Force No. 9 (July 1987), a title in Marvel's short-lived New Universe imprint. This led to his becoming that title's regular writer from #16 (Feb. 1988) until #32 (June 1989), the final issue. This led to fill-in work on titles such as Classic X-Men, for which he provided backup stories, and in the Marvel Annuals' 1989 summer crossover "Atlantis Attacks".

After Tom DeFalco, then Marvel's editor-in-chief, created the superhero team the New Warriors, using existing characters, in Thor No. 412 (Dec. 1989), he selected Nicieza to write the spin-off series. Nicieza recalled "I took the assignment for two reasons. First, I saw a lot of potential in these characters that had already been deemed useless. And secondly, I really wanted to write a monthly book." Collaborating with pencilers Mark Bagley and later Darick Robertson, primarily, Nicieza went on to write the title for most of its first 53 issues (July 1990 – November 1994). Years later, Nicieza said that he considers the first 25 issues of New Warriors to be the best work of his career.

Also in 1990 Nicieza began short runs on comics such as Alpha Flight (#87–101), Avengers (#317–325) and Avengers Spotlight, as well as the miniseries Nomad, which in turn led him to write the ongoing series Nomad vol. 2 in 1992. That year, Nicieza became editor of Marvel's children's imprint, Star Comics. Shortly afterward, he left the Marvel staff and began freelance writing for the company.

Nicieza's projects in this period included the first four issues of National Football League-approved superhero NFL SuperPro (Oct. 1991 – Feb. 1992), and, with penciler Kevin Maguire, the four-issue  miniseries Adventures of Captain America (also known by its cover-logo treatment, The Adventures of Captain America: Sentinel of Liberty) (Sept. 1991 – Jan. 1992), an origin-story retelling set in the 1940s.

The X-Men
In 1991, Nicieza joined with artist Rob Liefeld in co-plotting and writing the final three issues of the New Mutants. In those issues Liefeld and Nicieza created the characters Deadpool and Shatterstar as well as the super team, X-Force. Liefeld and Nicieza then produced an ongoing X-Force title. Nicieza initially worked on the title as scripter; after the departure of Liefeld in No. 12 he became its full writer, which he remained until 1995. By the end of 1992, Nicieza became regular scripter for X-Men vol. 2, beginning with No. 12 (Sept. 1992), working primarily with penciler Andy Kubert throughout his run. For the next three years, Nicieza was among the writers and editors of one of Marvel's most popular superhero franchises during a time of such popular, multi-series crossover story arcs as "X-Cutioner's Song", "Phalanx Covenant" and "Age of Apocalypse".

During this period Nicieza wrote the first Cable miniseries as well as the first few issues of the character's subsequent ongoing series. He also wrote the first solo Deadpool series, Deadpool: the Circle Chase in 1993. These series expanded the characters' personalities and established key background information for both characters, all things which were later used by other writers on those characters' subsequent ongoing books.

However, in 1995, in a dispute with then editor-in-chief Bob Harras over the future direction of his plotlines on X-Force, Nicieza was fired from the X-titles, leaving X-Force with No. 43 and X-Men with No. 45. He later remarked, "I never wanted to leave [X-Force], and never felt my firing was justified. ... I don't recall being given a reason [for being fired], and I also don't recall asking for one. ... Considering it was a Top 10 selling title at the time, I felt it was a wholly unjustified decision."

Acclaim Comics
After 1995, Nicieza wrote short runs of Captain Marvel  (vol. 2, 1995), Spider-Man: The Final Adventure (1995) and stories for Mighty Morphin Power Rangers before leaving the company in 1996. That year Nicieza did his first work for rival publisher DC Comics, co-writing Justice League: Midsummer Nightmare with Mark Waid which relaunched the Justice League as the JLA. He also worked for Twist and Shout Comics writing and pencilling back-up stories in X-Flies Special #1 and Dirtbag #7.

Later in 1996 Nicieza joined Acclaim Comics as senior vice-president and editor-in-chief. He was charged with revamping the companies intellectual properties which had previously formed Valiant Comics' Valiant Universe. Nicieza as editor oversaw the new version, dubbed "VH2", which re-imagined characters such as Solar, X-O Manowar, and Ninjak.

Nicieza himself wrote the Turok title as well as a new series, Troublemakers. Turok met with success as a video game adaptation, and Nicieza was promoted to president and publisher of Acclaim Comics in 1997. He also wrote a Turok novella during this period. However, after staff cuts and most of the lines' cancellation, Nicieza left Acclaim in 1999.

Freelance work
Returning to freelance work, Marvel and the X-Men, Nicieza co-wrote the Magneto Wars crossover through Uncanny X-Men #366–367 and X-Men vol. 2, #86–87, with artist Alan Davis in 1999. This led to the successive Magneto limited series Magneto Rex (1999) and Magneto: Dark Seduction (2000), as well as an ongoing Gambit (1999) series which he wrote for the first 24 issues of its 25-issue run.

Also in 1999, Nicieza began writing Thunderbolts with #34. He continued to write the book (initially with old partner Mark Bagley on art, later with Patrick Zircher and Chris Batista) up until No. 75 when the title was revamped. The revamp was unsuccessful, and in 2004 the original version of the team was resurrected, initially in an Avengers/Thunderbolts miniseries, then later in the New Thunderbolts series with Nicieza again as writer.

Nicieza also worked on several limited series at Marvel and DC around the turn of the century. At Marvel he wrote Citizen V (2001), Citizen V and the V Battalion: Everlasting (2002), X-Men Forever (2001), and X-Force vol. 2, as well as the short-lived ongoing series Hawkeye (2003); while at DC, he wrote the six-issue miniseries Supermen of America (1999) and the Elseworlds project JLA: Created Equal (2000), as well as some issues of the children's comic Justice League Adventures.

In 2003 Nicieza co-created, with artist Stefano Raffaele, the horror miniseries The Blackburne Covenant, published by Dark Horse Comics. That same year he returned to two of his old characters with the Marvel series Cable and Deadpool, of which he wrote all 50 issues.

In 2006, Nicieza returned to DC with a three-issue arc in Action Comics #841–843 (July–Sept. 2006), co-written with Kurt Busiek. Nicieza also wrote JSA Classified #28 (September 2007). He is also one of the co-writers for The 99, an "Islamic culture-based comic book" with Kuwaiti Naif Al-Mutawa, Other late-2000s DC work includes Nightwing and Robin, both titles being cancelled in connection with "Batman R.I.P." and Nicieza then wrote an Azrael : Death's Dark Knight mini-series, part of the Battle for the Cowl storyline which dealt with the "Batman R.I.P." aftermath. After Death's Dark Knight concluded, Nicieza wrote the new Azrael ongoing series from issue #1–13 (December 2009 – December 2010).

Nicieza began writing the DC series Red Robin from issues #13-26, the final issue (Aug. 2010 - Aug. 2011). DC announced Nicieza would be writing Legion Lost, a spinoff of Legion of Superheroes as part of DC's line wide relaunch initiative in September 2011. Nicieza wrote the first six issues before leaving the title. In 2016, for the comics company Shatner Singularity, he adapted a Stan Lee poem into the graphic novel Stan Lee's 'God Woke'. That work won the 2017 Independent Publisher Book Awards' Outstanding Books of the Year Independent Voice Award.

In 2023, it was announced that Nicieza would be returning to the X-Men line of books to write the fifth volume of Cable, as a four-issue mini-series beginning in January, 2024.

Non-comics work
In non-comics works, Nicieza co-scripted the direct-to-DVD animated feature Hot Wheels World Race, and the animated DVD feature The Black Belt Club, based on the Scholastic book series. In 2021, Nicieza created and executive produced the animated streaming series Superhero Kindergarten, based on the comic book series of the same name by Stan Lee. In 2021, Nicieza's first non-comic book was released: Suburban Dicks, a satirical crime novel set in the New Jersey suburbs. A second book with the same characters, The Self-Made Widow, was published in 2022.

title: Fabian Nicieza


text:
Ailsa Mellon Bruce

Ailsa Mellon Bruce (June 28, 1901 – August 25, 1969) was a prominent American socialite and philanthropist who established the Avalon Foundation.

Early life
Ailsa was born in Pittsburgh, Pennsylvania, on June 28, 1901. She was the daughter of the banker and diplomat Andrew W. Mellon and Nora Mary (née McMullen) Mellon (1879–1937). Her parents divorced in 1912 and from 1921 to 1932, Ailsa served as her father's official hostess during his tenure as United States Secretary of the Treasury, and again when he was U.S. Ambassador to the United Kingdom in 1932–1933. Her only sibling was brother Paul Mellon (1907–1999), who was also a philanthropist and was known as a prominent owner/breeder of thoroughbred racehorses.

Ailsa attended Miss Porter's School in Farmington, Connecticut and spent her summers as a teenager in Europe.

Philanthropy and legacy
Bruce established the Avalon Foundation in 1940, which made grants to colleges and universities, medical schools and hospitals, youth programs and community services, churches, environmental projects, and an array of cultural and arts organizations. In 1947, the Avalon Foundation was instrumental in the establishment of the Hampton National Historic Site in Maryland.

In 1957, when Fortune prepared its first list of the wealthiest Americans, it estimated that Ailsa Mellon Bruce, her brother, Paul, and her cousins, Sarah Mellon and Richard King Mellon, were all among the richest eight people in the United States, with fortunes of between 400 and 700 million dollars each. In 1968, Ailsa and Paul donated $20 million to build an annex to the National Gallery of Art in Washington, D.C.

Ailsa Bruce was a silent partner, with Nona McAdoo Park and Sophie Meldrim Shonnard, founding Chez Ninon in 1928, on Madison Avenue.

At her death in 1969, Ailsa Bruce bequeathed 153 paintings, primarily by 19th-century French artists, to the National Gallery of Art, as well as establishing a fund for future acquisitions. Among the many works acquired by the Gallery through the Ailsa Mellon Bruce Fund was the portrait of Ginevra de' Benci, the only painting by Leonardo da Vinci in the United States. In 1969, the assets of Paul Mellon’s Old Dominion Foundation were merged into his sister's Avalon Foundation, which was renamed the Andrew W. Mellon Foundation in honor of their father.

Personal life
She dated Prince Otto Bismarck, the grandson of Otto von Bismarck, the "Iron Chancellor", and was close to marrying him, but decided to marry David Bruce, an American, instead.

On May 23, 1926, she married David Kirkpatrick Este Bruce (1898–1977), a scion of a prominent Virginia family including his father William Cabell Bruce, a U.S. Senator from Maryland, and brother James Cabell Bruce, the U.S. Ambassador to Argentina. Their engagement, marriage (which was attended by President and Mrs. Coolidge) and honeymoon were followed closely by the news media. In 1933, after seven years of marriage, Ailsa gave birth to her only child:

 * Audrey Sheila Bruce (1934–1967), who married Stephen Richard Currier (1930–1967), who went missing during a flight in the Caribbean in January 1967 and were never recovered. They were the founders of the Taconic Foundation, a charitable giving organization, which was instrumental in the formation of the Council for United Civil Rights Leadership.

She obtained a divorce from her husband in Palm Beach, Florida in April 1945 on the grounds of "desertion and mental cruelty", receiving sole custody of their 11-year-old daughter. Following their divorce, her ex-husband would later become the United States Ambassador to the United Kingdom from 1961 to 1969, the same position her father held.

After her divorce, Mrs. Bruce was in a long rumored relationship with G. Lauder Greenway of the Lauder Greenway Family. In addition to their personal links, Greenway was a longtime trustee of Bruce's Avalon Foundation.

She died on August 25, 1969, at Roosevelt Hospital in New York City. She had homes at 960 Fifth Avenue, a beach house in Atlantic Beach, NY, and a 121-acre estate in Syosset, New York on Long Island. Her obituary in The New York Times called her the "Richest Woman in U.S."

Descendants
When Audrey and her husband, Stephen Currier, died in a presumed plane crash in 1967, leaving three young children – Andrea Currier, Lavinia Currier, and Michael Stephen Currier (1961–1998), she decided to bequeath her collection of 18th-century English furniture and ceramics to the Carnegie Institute of Pittsburgh, Pennsylvania.

title: Ailsa Mellon Bruce


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History of Saturday Night Live (2005–2010)

Saturday Night Live is an American sketch comedy series created and produced by Lorne Michaels for most of the show's run. The show has aired on NBC since 1975.

2005-2006 saw the debuts of Bill Hader, Andy Samberg, and Kristen Wiig. Samberg came aboard the show with his collaborators from The Lonely Island, and the trio would help usher SNL into the digital era with their viral short films. The 33rd season was interrupted by the 2007 writers strike. Production was paused in November and did not resume until February 2008 after the strike ended.

The 2008 U.S. presidential election gave the show opportunities to parody politicians such as Sarah Palin, John McCain, and Barack Obama, which led to high ratings. Weekend Update was largely co-anchored by Seth Meyers and Amy Poehler, until 2009-2010 when Meyers became the solo anchor.

Hader, Samberg, and Wiig join (2005–2006)
Saturday Night Live promised changes for the 2005–06 season, one of which was broadcasting in high-definition. Lorne Michaels added four new featured players: Andy Samberg, Bill Hader, Kristen Wiig (partway through the season), and Jason Sudeikis, who was added for the last three episodes of the previous season. Hader quickly became popular for his impersonations, such as of Vincent Price and Al Pacino, and characters such as New York City correspondent Stefon. Wiig, who first appeared when Jason Lee hosted, gained popularity with impersonations of Drew Barrymore, Felicity Huffman and Megan Mullally, also creating memorable characters such as the Two A-Holes (with Sudeikis) and Target Lady.

Leaving after this season were Rachel Dratch and Tina Fey, who had committed to working on Fey's new sitcom 30 Rock, as well as Horatio Sanz, Finesse Mitchell, and Chris Parnell, who were let go due to budget cuts. Fey had missed several episodes early in the season while on maternity leave, as had Maya Rudolph. Horatio Sanz took Fey's place on Weekend Update while she was away.

The Lonely Island
Andy Samberg was hired alongside his longtime friends and collaborators Jorma Taccone and Akiva Schaffer; the trio were in the comedy troupe The Lonely Island. Taccone and Shaffer joined the writing staff. The three would create a new SNL mainstay feature in the SNL Digital Shorts, the most popular being "Lazy Sunday".

Cast
Repertory players
 * Fred Armisen
 * Rachel Dratch
 * Tina Fey
 * Will Forte
 * Darrell Hammond
 * Seth Meyers
 * Finesse Mitchell
 * Chris Parnell
 * Amy Poehler
 * Maya Rudolph
 * Horatio Sanz
 * Kenan Thompson

Featured players
 * Bill Hader
 * Andy Samberg
 * Jason Sudeikis
 * Kristen Wiig (first episode: November 12, 2005)

bold denotes Weekend Update anchor

2006–2007 season
SNL had a smaller cast in the 2006–2007 season due to "massive budget cuts" at NBC. Lorne Michaels said that cutting staff was chosen over reducing from 20 the number of original episodes produced. A separate announcement confirmed the departure of Tina Fey, who left to focus on her new show 30 Rock. Rachel Dratch, one of the stars in the 30 Rock pilot (though subsequently replaced by Jane Krakowski), also did not return.

In September 2006, it was announced that Seth Meyers would replace Tina Fey as co-anchor on Weekend Update.

Kristen Wiig, Bill Hader, Andy Samberg, and Jason Sudeikis were all promoted to repertory players at the beginning of their second season (the third for Sudeikis including his role as a featured player).

The final cast list included just 11 members, the lowest number since the 1997–1998 season, and the first time since the 1997–98 season that no featured players were hired.

Cast
Repertory players

 * Fred Armisen
 * Will Forte
 * Bill Hader
 * Darrell Hammond
 * Seth Meyers
 * Amy Poehler

 * Maya Rudolph
 * Andy Samberg
 * Jason Sudeikis
 * Kenan Thompson
 * Kristen Wiig

bold denotes Weekend Update anchor

Writers' strike (2007–2008)
The 2007–2008 season of Saturday Night Live began on September 29, 2007 with LeBron James hosting the episode and musical guest Kanye West. Production was suspended due to the 2007 Writers Guild of America strike, after which only four episodes were produced. The last episode before the hiatus was Maya Rudolph's final as a cast member.

The show's first episode after the writers' strike was aired on February 23, 2008, with former head writer and cast member Tina Fey as host. Casey Wilson debuted as a featured player in the episode. During the show's hiatus, Lorne Michaels held auditions for someone to play Barack Obama in political sketches for the upcoming 2008 election. Jordan Peele and Donald Glover were among those who auditioned for the role. Ultimately, cast member Fred Armisen was given the part, and he debuted as Obama in the first post-strike episode. Armisen's portrayal attracted some criticism as the comic is not himself black, but of Asian and Latino descent.

Cast
Repertory players
 * Fred Armisen
 * Will Forte
 * Bill Hader
 * Darrell Hammond
 * Seth Meyers
 * Amy Poehler
 * Maya Rudolph (final episode: November 3, 2007)
 * Andy Samberg
 * Jason Sudeikis
 * Kenan Thompson
 * Kristen Wiig

Featured players
 * Casey Wilson (first episode: February 23, 2008)

bold denotes Weekend Update anchor

Election coverage (2008–2009)
The 2008–2009 season of Saturday Night Live began September 13, 2008 with host Michael Phelps and musical guest Lil' Wayne. New cast members included Abby Elliott, Michaela Watkins, and Bobby Moynihan. SNL saw some of its highest ratings during its coverage of the 2008 presidential election. In the season premiere, Tina Fey appeared as Sarah Palin and Amy Poehler appeared as Hillary Clinton in A Nonpartisan Message from Governor Sarah Palin & Senator Hillary Clinton in the cold open. Fey would portray Palin five more times during the season, with the real Palin making an appearance on the fifth episode. The quote "I can see Russia from my house!", which was said during a sketch, is commonly misattributed to the real Palin. Fey won a Primetime Emmy Award for Outstanding Guest Actress in a Comedy Series for her portrayal.

Amy Poehler, who had been away from the Weekend Update desk because of maternity leave, returned to Update in December 2008 for her farewell episode.

Cast
Repertory players
 * Fred Armisen
 * Will Forte
 * Bill Hader
 * Darrell Hammond
 * Seth Meyers
 * Amy Poehler (final episode: December 13, 2008)
 * Andy Samberg
 * Jason Sudeikis
 * Kenan Thompson
 * Kristen Wiig

Featured players
 * Abby Elliott (first episode: November 15, 2008)
 * Bobby Moynihan
 * Michaela Watkins (first episode: November 15, 2008)
 * Casey Wilson

bold denotes Weekend Update anchor

2009–2010 season
The 2009–2010 season of Saturday Night Live began September 26, 2009 with host Megan Fox and musical guest U2. New cast members included Nasim Pedrad and Jenny Slate. In early 2010, an online campaign was created on Facebook to get actress Betty White to host an episode of SNL. In March 2010, White confirmed that she would be hosting the May 8 episode. White's episode garnered around 12 million viewers, the highest since the 2008 election.

This would be the final season for Will Forte, who had been on the show for eight seasons. Jenny Slate's contract was not renewed for the following season.

Cast
Repertory players
 * Fred Armisen
 * Will Forte
 * Bill Hader
 * Seth Meyers
 * Andy Samberg
 * Jason Sudeikis
 * Kenan Thompson
 * Kristen Wiig

Featured players
 * Abby Elliott
 * Bobby Moynihan
 * Nasim Pedrad
 * Jenny Slate

bold denotes Weekend Update anchor


text: User talk:80.177.108.116

title: User talk:80.177.108.116


text:
Harlington railway station

Harlington railway station is located in Bedfordshire. It is named after the village of Harlington, on the outskirts of which it is located, but serves a wide rural area including the larger villages of Toddington and Barton-le-Clay.

History
It was built by the Midland Railway in 1868 on its extension to St. Pancras. The original intention had been to call it "Harlington for Toddington". The station buildings still exist and were carefully restored in the early 1980s. The station is situated on the Midland Main Line and managed by Thameslink.

Stationmasters
In 1909 the station master, William Drake, was killed at the station whilst directing shunting operations at the station. A verdict of accidental death was recorded.

 * T. Tomblin 1870 - 1875
 * Frederick Christian 1875 - 1898 (afterwards station master at Shefford)
 * G.G. Best 1898 - 1903
 * William Drake 1903 - 1909
 * Ernest Joseph Clulow 1909 - ca. 1911 (formerly station master at Godmanchester)
 * J.J. Davies ca. 1914 - 1924
 * Robert Arthur Gill 1924 - 1931 (formerly station master at Hemel Hempstead)
 * Frederick Charles Watson 1933 - 1940 (afterwards station master at Kenilworth)
 * A. Latimer from 1940 (formerly station master at Kimbolton, also station master at Leagrave)

Services
All services at Harlington are operated by Thameslink using EMUs.

The typical off-peak service in trains per hour is:
 * 4 tph to Bedford
 * 2 tph to Brighton
 * 2 tph to Three Bridges via Redhill

During the peak hours, the station is served by additional services to and from Rainham, Sutton and East Grinstead.

The station is also served by a half-hourly night service between Bedford and Three Bridges on Sunday to Friday nights.

Facilities
Harlington station has the following facilities:
 * Shelters on each platform
 * 1 telephone
 * 1 Ticket Machine
 * Cycle storage for 44 bikes
 * Car park with 127 spaces

The station has a PlusBus scheme where train and bus tickets can be bought together for a cheaper price. It is in the same area as Flitwick station.

As well as Harlington village itself, the station also serves the villages of Barton-le-Clay, Toddington and Westoning.

Ticket Office opening hours
The ticket office is open for just over 7½ hours per day Mondays to Friday and 6 hours per day on Saturday.

In January 2009, the previous franchisee First Capital Connect proposed that the ticket office at Harlington railway station would open for just four hours per day. The proposals were for the office to open at 0645 (previously 0600) and close at 1030 (currently 1850) on weekdays. There would be no weekend opening under these proposals. The single automated ticket machine, which was stolen in summer 2008, was replaced the same week that the proposals were announced. Subsequently, whilst a reduction in hours was agreed, although not to the degree set out in the initial proposal (see above).

title: Harlington railway station


text:
Rosario Salazar

Rosario Inés Consuelo Yolanda Salazar (previously McFarland; also known as "Rosie" and "Ro-Ro") is a fictional character on the American television sitcom Will & Grace, portrayed by actress Shelley Morrison. Originally due to appear just once, the character was so popular with audiences that the show's writers made her a semi-regular character.

Early life
A native of El Salvador, Rosario was a school teacher, who received a bachelor's degree in clinical psychology from the University of Texas and who almost got a master's degree (three credits short); she also attended business school. At some point, she moved to The Bronx, presumably while studying business. However, Karen states that she bought Rosario from Rosario's parents, and another comment made by Jack states that Karen smuggles staff in from Latin America, namely Mexico, and probably El Salvador as well. While there, she also performed with a then-unknown Jennifer Lopez, including a production of "Tea for Two" at the Shalom Retirement Center. At some point, she was also paid to tape pornographic videos for Tommy Lee. One of Rosario's brothers is a political prisoner, and remarks made by Karen suggests she has children, possibly still in El Salvador.

Series overview
Although she had been mentioned by Karen since the pilot episode, Rosario's first appearance was in the season-one finale, "Object of My Rejection." Due to Karen's patronizing tone, we're made to think she doesn't speak English well. Rosario actually speaks English fluently albeit with a thick accent, but is an undocumented immigrant.

To keep her in the country, Karen has her marry Jack McFarland, her gay friend, to get her green card. The marriage was later terminated at the end of season two, when Rosario asked for a divorce to pursue other romantic interests. However, she maintained a close relationship with Jack after (often echoing hers with Karen) and claimed to still think of him as family. Rosario was one of few people with whom Stan remained in contact after he faked his death (at the end of season five). When Karen learned that Rosario had kept this news from her (in season eight), she fired her. Rosario worked as a bar attendant until Karen took her back. During a live episode, it is revealed by Karen that Rosario earns $350,000 per year.

In the show's series finale, Rosario was shown to have lived with Karen and Jack in the years after the show ended, growing older with them and possibly still working for them despite being in a wheelchair (though she states that she is only in the wheelchair due to a sprained ankle). Rosario is usually seen wearing a Members Only jacket over her maid's uniform. She has a particular fondness for Subway sandwiches. Morrison was a guest star in the first two seasons, but from season three until the eighth season, she was credited as a main cast member in the episodes she appeared.

Series revival
In the series revival in 2017, Rosario is an unseen character, referred to but never appearing. Morrison had declined to reprise her role, having retired from acting. The character was written out of the show in the episode "Rosario's Quinceañera", in which Rosario dies of a heart attack off screen.

Relationship with Karen Walker
In 1985, Rosario met Karen Walker at a club, while working as a cigarette lady, and went to work for her as a maid, which was where she was working when the series began in 1998. Rosario and Karen are very close, but their relationship is love-hate. They are notorious for swapping insults back and forth. Rosario is just as feisty as Karen, and is one of the few people that can go toe-to-toe with her.

In earlier episodes, they often yell and argue with each other at the same time. However, many of those fights end with an emotional embrace or light-hearted exchange between the two, which makes it clear that they really do care about each other. Examples of this include the season 3 episode "My Uncle, The Car," where Karen becomes sad at almost losing Rosario in a bet to her rival Beverley Leslie, and the season 7 episode "Partners," where Karen ignores the plight of a very sick Rosario because she is afraid of losing her. Rosario also mentions in season 4's "The Third Wheel Gets The Grace" that, for their 15-year anniversary, all she wanted to do was spend time with Karen.

When Rosario dies in the series revival, Karen arranges the funeral, giving Rosario the quinceañera she never had. She bolts out of the funeral before it starts, however, drowning her sorrows at a bar, unable to handle the loss of her friend. She later comes back to the church to sit next to the casket and say goodbye, admitting with rare emotion that Rosario was her best friend, her "sparring partner", and her "everything".

title: Rosario Salazar


text:
File:PollyPromo.jpg

Fair use in Polly (song)
Though this image is subject to copyright, its use is covered by the U.S. fair use laws because:
 * 1) It's a low resolution copy of a CD/album cover.
 * 2) It doesn't limit the copyright owner's rights to sell the CD/album in any way, in fact, it may encourage sales.
 * 3) Because of the low resolution, copies could not be used to make illegal copies of the album artwork on another CD.
 * 4) The image is itself a subject of discussion in the article or used in the infobox thereof.
 * 5) The image on the cover is significant because it was made by a famous artist.

Source

 * 1) Derived from a digital capture (scan/photo) of the album/CD cover (creator of this digital version is irrelevant as the copyright in all equivalent images is still held by the same party). Copyright held by the record company or the artist. Claimed as fair use regardless.

title: File:PollyPromo.jpg