GADD45A Recruits Demethylation Proteins to CpG Island Promoters

Previous studies have shown that the antisense long noncoding RNA (lncRNA) TCF21 antisense RNA inducing promoter demethylation (TARID) activates transcription of the tumor suppressor gene TCF21 by recruiting growth arrest and DNA damage inducible A (GADD45A) and stimulating TET1-mediated DNA demethylation. Building on these findings, Arab and colleagues found that the lncRNA TARID formed a DNA–RNA hybrid R-loop at the CpG island promoter of TCF21. The R-loop–forming sequence overlapped with a region of DNA demethylation in the TCF21 promoter, and degradation of this R-loop by RNase H1 decreased TCF21 expression, suggesting that TARID-dependent R-loop formation directs promoter demethylation and is necessary for TCF21 transcriptional activation. GADD45A, a stress response protein implicated in repair-mediated DNA demethylation, directly interacted with the TARID–TCF21 R-loop and with DNA–RNA hybrid structures in vitro and recruited the TET1 demethylase in an R-loop–dependent manner. Transcription of TARID and TCF21 were temporally regulated during the cell cycle, as TARID transcription occurred in early- to mid-S phase and was followed by accumulation of R-loops at the TCF21 promoter, GADD45A binding to R-loops, and TCF21 demethylation and transcriptional activation. Consistent with a role for R-loops as sites of promoter demethylation, global levels of oxidized 5′methylcytosine derivatives were enriched at R-loops. Chromatin immunoprecipitation sequencing of mouse embryonic stem cells revealed that TET1 was associated with thousands of genomic R-loops, in particular those found in CpG island regions around transcription start sites and a subset of regions associated with antisense lncRNAs. TET1 binding to R-loops was also detected at the promoters of orthologous genes in human primary skin fibroblasts, and knockdown of GADD45A and GADD45B specifically reduced TET1 recruitment to these R-loop–containing promoters. These findings identify GADD45A as an epigenetic reader of promoter-associated R-loops that targets demethylation factors to CpG island promoters to regulate transcription.

Arab K, Karaulanov E, Musheev M, Trnka P, Schäfer A, Grummt I, et al. GADD45A binds R-loops and recruits TET1 to CpG island promoters. Nat Genet 2019;51:217–23.

• ©2019 American Association for Cancer Research.