Currently available shRNA vectors and libraries have limited utility for in vivo studies of antiviral T cell responses. This hampers rapid identification of biological targets in vivo that could be manipulated therapeutically in the context of vaccines and adoptive immunotherapy to treat viral infections successfully. The overarching goals of Core B are to clone novel and superior shRNAmirs that cover the mouse genome more comprehensively than any other currently available tools, to do so in a vector that is useful for extended in vivo studies of adoptively transferred lymphocytes in the context of viral infections, and to make them available immediately to laboratories conducting Projects 1-3 of this U19 proposal, and ultimately to the research community at large. Our specific objectives are to clone two novel comprehensive gene-family shRNA libraries in a retroviral vector suitable for in vivo analysis of the murine immune system (Aim 1);to produce ready-to-transfect retroviral-shRNAmir DNA from cloned libraries for virus production (Aim 2);and, to clone smaller focused retroviral-shRNAmir sets and build custom delivery constructs (Aim 3), to facilitate specific follow-up studies outlined in Research Projects 1-3.
There are very limited tools available to analyze rapidly the requirements of each individual gene in the entire cell that lymphocytes use to mediate clearance of viral pathogens. We will produce the first comprehensive short hairpin RNA libraries that can disable individually hundreds of genes in parallel, in vivo, to elucidate gene function in T cells during viral infections.
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|Martinez, Gustavo J; Hu, Joyce K; Pereira, Renata M et al. (2016) Cutting Edge: NFAT Transcription Factors Promote the Generation of Follicular Helper T Cells in Response to Acute Viral Infection. J Immunol 196:2015-9|
|Pedros, Christophe; Zhang, Yaoyang; Hu, Joyce K et al. (2016) A TRAF-like motif of the inducible costimulator ICOS controls development of germinal center TFH cells via the kinase TBK1. Nat Immunol 17:825-33|
|Crotty, Shane (2015) A brief history of T cell help to B cells. Nat Rev Immunol 15:185-9|
|Nance, J Philip; BÃ©langer, Simon; Johnston, Robert J et al. (2015) Bcl6 middle domain repressor function is required for T follicular helper cell differentiation and utilizes the corepressor MTA3. Proc Natl Acad Sci U S A 112:13324-9|
|Martinez, Gustavo J; Pereira, Renata M; Ã„ijÃ¶, Tarmo et al. (2015) The transcription factor NFAT promotes exhaustion of activated CD8âº T cells. Immunity 42:265-78|
|Hatzi, Katerina; Nance, J Philip; Kroenke, Mark A et al. (2015) BCL6 orchestrates Tfh cell differentiation via multiple distinct mechanisms. J Exp Med 212:539-53|
|Choi, Youn Soo; Gullicksrud, Jodi A; Xing, Shaojun et al. (2015) LEF-1 and TCF-1 orchestrate T(FH) differentiation by regulating differentiation circuits upstream of the transcriptional repressor Bcl6. Nat Immunol 16:980-90|
|Nance, J Philip; BÃ©langer, Simon; Johnston, Robert J et al. (2015) Cutting edge: T follicular helper cell differentiation is defective in the absence of Bcl6 BTB repressor domain function. J Immunol 194:5599-603|
|Crotty, Shane; Pipkin, Matthew E (2015) In vivo RNAi screens: concepts and applications. Trends Immunol 36:315-22|
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