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Immunity. 2014 Aug 21;41(2):325-38. doi: 10.1016/j.immuni.2014.08.002.

In vivo RNA interference screens identify regulators of antiviral CD4(+) and CD8(+) T cell differentiation.

Author information

1
Division of Signaling and Gene Expression, La Jolla Institute for Allergy and Immunology, La Jolla, CA 92037, USA; Division of Vaccine Discovery, La Jolla Institute for Allergy and Immunology, La Jolla, CA 92037, USA; Department of Cancer Biology, The Scripps Research Institute, Jupiter, FL 33458, USA.
2
Division of Vaccine Discovery, La Jolla Institute for Allergy and Immunology, La Jolla, CA 92037, USA.
3
Department of Cancer Biology, The Scripps Research Institute, Jupiter, FL 33458, USA.
4
Division of Cell Biology, La Jolla Institute for Allergy and Immunology, La Jolla, CA 92037, USA.
5
Division of Signaling and Gene Expression, La Jolla Institute for Allergy and Immunology, La Jolla, CA 92037, USA.
6
Division of Vaccine Discovery, La Jolla Institute for Allergy and Immunology, La Jolla, CA 92037, USA. Electronic address: shane@lji.org.
7
Department of Cancer Biology, The Scripps Research Institute, Jupiter, FL 33458, USA. Electronic address: mpipkin@scripps.edu.

Abstract

Classical genetic approaches to examine the requirements of genes for T cell differentiation during infection are time consuming. Here we developed a pooled approach to screen 30-100+ genes individually in separate antigen-specific T cells during infection using short hairpin RNAs in a microRNA context (shRNAmir). Independent screens using T cell receptor (TCR)-transgenic CD4(+) and CD8(+) T cells responding to lymphocytic choriomeningitis virus (LCMV) identified multiple genes that regulated development of follicular helper (Tfh) and T helper 1 (Th1) cells, and short-lived effector and memory precursor cytotoxic T lymphocytes (CTLs). Both screens revealed roles for the positive transcription elongation factor (P-TEFb) component Cyclin T1 (Ccnt1). Inhibiting expression of Cyclin T1, or its catalytic partner Cdk9, impaired development of Th1 cells and protective short-lived effector CTL and enhanced Tfh cell and memory precursor CTL formation in vivo. This pooled shRNA screening approach should have utility in numerous immunological studies.

PMID:
25148027
PMCID:
PMC4160313
DOI:
10.1016/j.immuni.2014.08.002
[Indexed for MEDLINE]
Free PMC Article

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