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

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


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.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • CD4-Positive T-Lymphocytes / immunology*
  • CD8-Positive T-Lymphocytes / immunology*
  • Cell Differentiation / genetics
  • Cell Differentiation / immunology*
  • Cyclin T / biosynthesis
  • Cyclin T / genetics
  • Cyclin-Dependent Kinase 9 / biosynthesis
  • Cyclin-Dependent Kinase 9 / genetics
  • Immunologic Memory / immunology
  • Lymphocyte Activation / immunology
  • Lymphocytic Choriomeningitis / immunology
  • Lymphocytic choriomeningitis virus / immunology*
  • Mice
  • Mice, Inbred C57BL
  • MicroRNAs / genetics
  • Positive Regulatory Domain I-Binding Factor 1
  • RNA Interference / immunology*
  • RNA, Small Interfering
  • Receptors, Antigen, T-Cell / genetics
  • T-Box Domain Proteins / genetics
  • T-Lymphocytes, Cytotoxic / immunology
  • Th1 Cells / immunology
  • Transcription Factors / genetics
  • Transduction, Genetic / methods


  • Ccnt1 protein, mouse
  • Cyclin T
  • MicroRNAs
  • Prdm1 protein, mouse
  • RNA, Small Interfering
  • Receptors, Antigen, T-Cell
  • T-Box Domain Proteins
  • T-box transcription factor TBX21
  • Transcription Factors
  • Positive Regulatory Domain I-Binding Factor 1
  • Cdk9 protein, mouse
  • Cyclin-Dependent Kinase 9