Mature T cell responses are controlled by microRNA-142
Yaping Sun, … , Thomas Saunders, Pavan Reddy
Yaping Sun, … , Thomas Saunders, Pavan Reddy
Published June 22, 2015
Citation Information: J Clin Invest. 2015;125(7):2825-2840. https://doi.org/10.1172/JCI78753.
View: Text | PDF
Research Article Immunology

Mature T cell responses are controlled by microRNA-142

Abstract

T cell proliferation is critical for immune responses; however, the molecular mechanisms that mediate the proliferative response are poorly understood. MicroRNAs (miRs) regulate various molecular processes, including development and function of the immune system. Here, utilizing multiple complementary genetic and molecular approaches, we investigated the contribution of a hematopoietic-specific miR, miR-142, in regulating T cell responses. T cell development was not affected in animals with a targeted deletion of Mir142; however, T cell proliferation was markedly reduced following stimulation both in vitro and in multiple murine models of graft-versus-host disease (GVHD). miR-142–deficient T cells demonstrated substantial cell-cycling defects, and microarray and bioinformatics analyses revealed upregulation of genes involved in cell cycling. Moreover, 2 predicted miR-142 target genes, the atypical E2F transcription factors E2f7 and E2f8, were most highly upregulated in miR-142–deficient cells. Clustered regularly interspaced short palindromic repeat interference–mediated (CRISPRi-mediated) silencing of E2F7 and E2F8 in miR-142–deficient T cells ameliorated cell-cycling defects and reduced GVHD, and overexpression of these factors in WT T cells inhibited the proliferative response. Together, these results identify a link between hematopoietic-specific miR-142 and atypical E2F transcription factors in the regulation of mature T cell cycling and suggest that targeting this interaction may be relevant for mitigating GVHD.

Authors

Yaping Sun, Katherine Oravecz-Wilson, Nathan Mathewson, Ying Wang, Richard McEachin, Chen Liu, Tomomi Toubai, Julia Wu, Corinne Rossi, Thomas Braun, Thomas Saunders, Pavan Reddy

×

Figure 1

Generation of miR-142 KO mice and its impact on T cell functional responses.

Options: View larger image (or click on image) Download as PowerPoint
Generation of miR-142 KO mice and its impact on T cell functional respon...
(A) Scheme of the Mir142 locus and targeting vector used to generate Mir142 null alleles. 5′ and 3′ arm probes (5′ probe and 3′ probe) and 5′ and 3′ primers (5F/5R and 3F/3R) for genotyping are shown. (B) A representative genotyping result illustrating WT (Mir142+/+), heterozygous (Mir142+/–), and null mice (Mir142–/–). (C) Zygosity determination by TaqMan qPCR using Tert as positive control and probes specific for 3′ and 5′ arms designed for Mir142 gene homologous recombination to confirm the deletion of the Mir142 gene in homozygous KO mice and single-copy loss in heterozygous mice. Data represent a combination of 6 independent experiments. P values were obtained using unpaired t test. **P < 0.01. (D) The loss of miR-142 expression at the RNA level in purified T cells from miR-142 KO mice confirmed by TaqMan qPCR using specific probes against miR-142–3p. Data represent a combination of 3 independent experiments. (E and F) Significantly reduced proliferation of miR-142 KO T cells determined by tritium incorporation and decreased apoptosis determined by annexin V staining when stimulated in vitro with anti-CD3 and anti-CD28 Abs for 2 to 6 days (E) or stimulated in vitro with allogeneic (allo) splenocytes for 2 to 6 days (F). Syn, syngeneic. Data shown represent results from 3 or 4 independent experiments (mean ± SEM). P values were obtained using the Holm-Sidak method. *P < 0.05; **P < 0.01.