Thpok promotes CD4+-lineage differentiation by recruiting the NuRD chromatin remodeling complex

Abstract

Abstract T cell precursors develop into helper CD4 or cytotoxic CD8 T cells in thymus. Thpok, a BTB/POZ family transcription factor, enforces MHC II restricted thymocytes commitment to the CD4 lineage, while Runx3 protein drives MHC I restricted thymocytes to the CD8 lineage. However, the molecular mechanisms underlying Thpok functions during CD4 T cell development are not understood. Replacing the BTB domain from Thpok with that from Bcl6, another BTB/POZ zinc finger transcription factor that recruits NCOR-family co-repressors through its BTB domain, abrogated CD4 T cell development in thymus; this suggests a unique feature associated with Thpok BTB domain. Here, using mass-spectrometry, we identified nucleosome remodeling and deacetylase (NuRD) complex as a novel Thpok cofactor. We demonstrated that the Thpok BTB domain is essential for NuRD recruitment. Moreover, we identified BTB domain amino acid residues critical for NuRD complex interaction, and showed that mutation of these Thpok residues disrupts CD4 T cell development in vivo. Reciprocally, a fusion protein reconstituting NuRD binding to a BTB-less version of Thpok (which cannot bind NuRD) restored CD4 T cells in vivo. In addition, we found that CD4-lineage thymocytes expressing that fusion protein have a transcriptome similar to that of thymocytes expressing wild type Thpok. Finally, we found that NuRD binding is required for Thpok to antagonize Runx3 expression in thymocytes. Thus, our results demonstrate that NuRD recruitment is both necessary and sufficient for the functions of the Thpok BTB domain in CD4 T cell development and it is also required to antagonize Runx3-dependent differentiation towards CD8 lineage.