title: In vivo RNA interference screens identify regulators of antiviral CD4(+) and CD8(+) T cell differentiation.
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.
authors.affiliation: 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.
authors.forename: Runqiang
authors.initials: R
authors.lastname: Chen
authors.affiliation: Division of Vaccine Discovery, La Jolla Institute for Allergy and Immunology, La Jolla, CA 92037, USA.
authors.forename: Simon
authors.initials: S
authors.lastname: Bélanger
authors.affiliation: Department of Cancer Biology, The Scripps Research Institute, Jupiter, FL 33458, USA.
authors.forename: Megan A
authors.initials: MA
authors.lastname: Frederick
authors.affiliation: Division of Vaccine Discovery, La Jolla Institute for Allergy and Immunology, La Jolla, CA 92037, USA.
authors.forename: Bin
authors.initials: B
authors.lastname: Li
authors.affiliation: Division of Vaccine Discovery, La Jolla Institute for Allergy and Immunology, La Jolla, CA 92037, USA.
authors.forename: Robert J
authors.initials: RJ
authors.lastname: Johnston
authors.affiliation: Division of Cell Biology, La Jolla Institute for Allergy and Immunology, La Jolla, CA 92037, USA.
authors.forename: Nengming
authors.initials: N
authors.lastname: Xiao
authors.affiliation: Division of Cell Biology, La Jolla Institute for Allergy and Immunology, La Jolla, CA 92037, USA.
authors.forename: Yun-Cai
authors.initials: YC
authors.lastname: Liu
authors.affiliation: Division of Cell Biology, La Jolla Institute for Allergy and Immunology, La Jolla, CA 92037, USA.
authors.forename: Sonia
authors.initials: S
authors.lastname: Sharma
authors.affiliation: Division of Vaccine Discovery, La Jolla Institute for Allergy and Immunology, La Jolla, CA 92037, USA.
authors.forename: Bjoern
authors.initials: B
authors.lastname: Peters
authors.affiliation: Division of Signaling and Gene Expression, La Jolla Institute for Allergy and Immunology, La Jolla, CA 92037, USA.
authors.forename: Anjana
authors.initials: A
authors.lastname: Rao
authors.affiliation: Division of Vaccine Discovery, La Jolla Institute for Allergy and Immunology, La Jolla, CA 92037, USA. Electronic address: shane@lji.org.
authors.forename: Shane
authors.initials: S
authors.lastname: Crotty
authors.affiliation: Department of Cancer Biology, The Scripps Research Institute, Jupiter, FL 33458, USA. Electronic address: mpipkin@scripps.edu.
authors.forename: Matthew E
authors.initials: ME
authors.lastname: Pipkin
mesh_terms: D000818:Animals; D015496:CD4-Positive T-Lymphocytes / Q000276:immunology*; D018414:CD8-Positive T-Lymphocytes / Q000276:immunology*; D002454:Cell Differentiation / Q000235:genetics / Q000276:immunology*; D056906:Cyclin T / Q000096:biosynthesis / Q000235:genetics; D042863:Cyclin-Dependent Kinase 9 / Q000096:biosynthesis / Q000235:genetics; D007156:Immunologic Memory / Q000276:immunology; D008213:Lymphocyte Activation / Q000276:immunology; D008216:Lymphocytic Choriomeningitis / Q000276:immunology; D008217:Lymphocytic choriomeningitis virus / Q000276:immunology*; D051379:Mice; D008810:Mice, Inbred C57BL; D035683:MicroRNAs / Q000235:genetics; D000074462:Positive Regulatory Domain I-Binding Factor 1; D034622:RNA Interference / Q000276:immunology*; D034741:RNA, Small Interfering; D011948:Receptors, Antigen, T-Cell / Q000235:genetics; D020825:T-Box Domain Proteins / Q000235:genetics; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology; D018417:Th1 Cells / Q000276:immunology; D014157:Transcription Factors / Q000235:genetics; D014161:Transduction, Genetic / Q000379:methods; D000098815:T-bet Transcription Factor
chemical_list: C503685:Ccnt1 protein, mouse; D056906:Cyclin T; D035683:MicroRNAs; C513258:Prdm1 protein, mouse; D034741:RNA, Small Interfering; D011948:Receptors, Antigen, T-Cell; D020825:T-Box Domain Proteins; D000098815:T-bet Transcription Factor; D014157:Transcription Factors; D000074462:Positive Regulatory Domain I-Binding Factor 1; C467972:Cdk9 protein, mouse; D042863:Cyclin-Dependent Kinase 9