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J Biol Chem. 2019 Sep 3. pii: jbc.RA119.010160. doi: 10.1074/jbc.RA119.010160. [Epub ahead of print]

The finger loop of the SRA domain in the E3 ligase UHRF1 is a regulator of ubiquitin targeting and is required for maintaining DNA methylation.

Author information

1
Van Andel Research Institute, United States.
2
Center for Epigenetics, Van Andel Research Institute, United States of America.

Abstract

The SET and RING associated (SRA) protein domain is conserved across bacteria and eukaryota and coordinates extrahelical or 'flipped' DNA bases. A functional SRA domain is required for ubiquitin-like with PHD and RING finger domains 1 (UHRF1) E3 ubiquitin ligase activity toward histone H3, a mechanism for recruiting the DNA methylation maintenance enzyme DNA methyltransferase 1 (DNMT1). The SRA domain supports UHRF1 oncogenic activity in colon cancer cells, highlighting that UHRF1 SRA antagonism could be a cancer therapeutic strategy. Here, we used molecular dynamics simulations, DNA binding assays, in vitro ubiquitination reactions, and DNA methylation analysis, to identify the SRA finger loop as a regulator of UHRF1 ubiquitin targeting and DNA methylation maintenance. A chimeric UHRF1 (finger swap) with diminished E3 ligase activity toward nucleosomal histones, despite tighter binding to unmodified or asymmetric or symmetrically methylated DNA, uncouples DNA affinity from regulation of E3 ligase activity. Our model suggests that SRA domains sample DNA bases through flipping in the presence or absence of a cytosine modification and that specific interactions of the SRA finger loop with DNA are required for downstream host protein function. Our findings provide insight into allosteric regulation of UHRF1 E3 ligase activity, suggesting that UHRF1's SRA finger loop regulates its conformation and function.

KEYWORDS:

DNA binding protein; DNA methylation; E3 ubiquitin ligase; SRA domain; allosteric regulation; epigenetics; molecular dynamics; string method in collective variables; ubiquitin-like with PHD and RING finger domains 1 (UHRF1)

PMID:
31481468
DOI:
10.1074/jbc.RA119.010160
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