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.

Public Health Relevance

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.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Program--Cooperative Agreements (U19)
Project #
1U19AI109976-01
Application #
8711628
Study Section
Special Emphasis Panel (ZAI1-ZL-I (J1))
Project Start
Project End
Budget Start
2014-03-01
Budget End
2015-02-28
Support Year
1
Fiscal Year
2014
Total Cost
$403,797
Indirect Cost
$65,145
Name
La Jolla Institute
Department
Type
DUNS #
603880287
City
La Jolla
State
CA
Country
United States
Zip Code
92037
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Nance, J Philip; Bélanger, 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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