Polycomb Group Protein Yin Yang 1 Controls DNA Methylation in Murine T Lymphocyte Development

Abstract

Transcription factors and epigenetic regulators play key functions in T lymphocyte lineage commitment and development. Mammalian PcG proteins are major negative regulators of gene expression that typically bind as large complexes (such as Polycomb Repressive Complex 1 (PRC1) and PRC2) to gene regulatory regions selectively targeted by an as yet unknown mechanism. While nearly all mammalian PcG proteins are lack of specific DNA binding capacities, transcription factor Yin Yang 1 (YY1) can bind at the DNA sites specifically and recruit other PcG proteins to specific DNA sequences. A small 25 amino acid YY1 domain (the REPO domain) is necessary and sufficient for the recruitment of PcG proteins, PcG-mediated stable transcriptional repression and histone modification. While YY1ΔREPO is competent for DNA binding, transcriptional activation, transient transcriptional repression and interaction with transcriptional coregulators such as HDACs, YY1ΔREPO is defective in all YY1 PcG functions. Our prior study demonstrated that deletion of YY1 in the hematopoietic system leads to T cell development blockage at the double-negative 3 (DN3) stage. Interestingly, ectopic expression of YY1∆REPO in YY1 null background by retroviral bone marrow transplantation could not rescue YY1 deficiency mediated T cell survival defect. This suggests that the YY1 REPO domain/PcG function plays a vital role in T cell development. Herein, we generated the YY1 REPO domain conditional knockout mouse model ( Yy1 f/∆REPO Vav-Cre) by CRISPR/Cas9 to further dissect the YY1 REPO domain/PcG function in T cell development. Yy1 f/∆REPO Vav-Cre ( Yy1 -/∆REPO) mice have reduced thymus weight and cellularity with medulla aplasia. Compared with wild-type (WT) and YY1 heterozygous ( Yy1 +/-) mice, Yy1 -/∆REPO mice have an increased percentage of DN3 cells with decreased DN4 percentage supporting a developmental blockage at the DN3 stage. Yy1 -/∆REPO DN T cells fail to go through TCRβ rearrangement with increased apoptosis and reduction of TCRβ + cells in the thymus compared with WT and Yy1 +/-cells. Interestingly, RNA-seq analysis in sorted primary DN3 cells revealed the genetic network governing DNA methylation, histone methylation, chromosome maintenance, and translation initiation complex formation are deregulated in Yy1 -/∆REPO mice compared to WT and Yy1 +/-mice. DN T cell development requires precise and dynamic regulation of histone and DNA methylation and demethylation. Interestingly, the DNA demethylase enzymes ten-eleven translocation (TET) family proteins Tet1 and Tet2 are downregulated in Yy1 -/∆REPO DN3 thymocytes compared with WT and Yy1 +/- DN3 cells. Our data show that the YY1 REPO domain/PcG function is required for DN3-DN4 transition via regulating TCRβ rearrangement and the YY1 REPO domain/PcG function plays a critical role in proper DNA methylation status in early T cell development. As PcG proteins modify histone via trimethylation of histone H3 on lysine 27, we will further assess the impact of YY1 REPO domain/PcG function on histone methylation in T cell development. By utilizing a powerful mouse model for identifying YY1 functions as PcG domain-dependent or PcG domain-independent, our study has generated fundamental new insights into the epigenetic landscape in T cell development.