PROSER1 mediates TET2 O-GlcNAcylation to regulate DNA demethylation on UTX-dependent enhancers and CpG islands.

Xiaokang Wang,Tanner Martinez,Yurii Sedkov,Kristian Helin,Jesper Christensen,Patrick Schreiner,Wojciech Rosikiewicz,Hans-Martin Herz,Shondra M Pruett-Miller,Baranda S Hansen,Beisi Xu

doi:10.26508/lsa.202101228

Abstract

DNA methylation at enhancers and CpG islands usually leads to gene repression, which is counteracted by DNA demethylation through the TET protein family. However, how TET enzymes are recruited and regulated at these genomic loci is not fully understood. Here, we identify TET2, the glycosyltransferase OGT and a previously undescribed proline and serine rich protein, PROSER1 as interactors of UTX, a component of the enhancer-associated MLL3/4 complexes. We find that PROSER1 mediates the interaction between OGT and TET2, thus promoting TET2 O-GlcNAcylation and protein stability. In addition, PROSER1, UTX, TET1/2, and OGT colocalize on many genomic elements genome-wide. Loss of PROSER1 results in lower enrichment of UTX, TET1/2, and OGT at enhancers and CpG islands, with a concomitant increase in DNA methylation and transcriptional down-regulation of associated target genes and increased DNA hypermethylation encroachment at H3K4me1-predisposed CpG islands. Furthermore, we provide evidence that PROSER1 acts as a more general regulator of OGT activity by controlling O-GlcNAcylation of multiple other chromatin signaling pathways. Taken together, this study describes for the first time a regulator of TET2 O-GlcNAcylation and its implications in mediating DNA demethylation at UTX-dependent enhancers and CpG islands and supports an important role for PROSER1 in regulating the function of various chromatin-associated proteins via OGT-mediated O-GlcNAcylation.

Highlights

To assess the relationship between UTX, PROSER1, and TET2 in more detail we FLAG affinity-purified UTX followed by glycerol gradient fractionation and observed that PROSER1 and TET2 co-migrated with UTX in fractions containing RBBP5, a core subunit of the MLL3/4 complexes (Fig 1C)
These results indicate that PROSER1 and TET2 associate with the MLL3/4 complexes
Genome-wide we find that PROSER1 regulates the chromatin association of TET1/2 to mediate DNA demethylation at UTX/H3K4me1-dependent enhancers and CpG islands (CGIs) (Fig 7A)

Summary

Introduction

Based on our findings that PROSER1 regulates TET1 and TET2 protein stability and their occupancy on UTX/H3K4me1-dependent enhancers (Figs 3D and 4C and D), we investigated how the genome-wide DNA methylation pattern was affected in PROSER1 KO cells. Differentially methylated regions extending over 500 bp (DMR500) or 1,000 bp (DMR1000) which were associated with increased DNA methylation and overlapping with at least one PROSER1, UTX, or H3K4me1 peak showed a decrease of UTX, TET1, TET2, OGT, H3K4me1, H3K4me2, and H3K4me3 enrichment and were associated with gene repression of their nearest target gene in PROSER1 KO versus WT cells (Figs 5D, S4D, and S5B and C).

Results

Conclusion