Enhancement of NEIL1 protein-initiated oxidized DNA base excision repair by heterogeneous nuclear ribonucleoprotein U (hnRNP-U) via direct interaction

Muralidhar L. Hegde, Srijita Banerjee, Pavana M. Hegde, Larry J. Bellot, Tapas K. Hazra, Istvan Boldogh, Sankar Mitra

Research output: Contribution to journalArticle

34 Scopus citations

Abstract

Repair of oxidized base lesions in the human genome, initiated by DNA glycosylases, occurs via the base excision repair pathway using conserved repair and some non-repair proteins. However, the functions of the latter noncanonical proteins in base excision repair are unclear. Here we elucidated the role of heterogeneous nuclear ribonucleoprotein-U (hnRNP-U), identified in the immunoprecipitate of human NEIL1, a major DNA glycosylase responsible for oxidized base repair. hnRNP-U directly interacts with NEIL1 in vitro via the NEIL1 common interacting C-terminal domain, which is dispensable for its enzymatic activity. Their in-cell association increases after oxidative stress. hnRNP-U stimulates the NEIL1 in vitro base excision activity for 5-hydroxyuracil in duplex, bubble, forked, or single-stranded DNA substrate, primarily by enhancing product release. Using eluates from FLAG-NEIL1 immunoprecipitates from human cells, we observed 3-fold enhancement in complete repair activity after oxidant treatment. The lack of such enhancement in hnRNP-U-depleted cells suggests its involvement in repairing enhanced base damage after oxidative stress. The NEIL1 disordered C-terminal region binds to hnRNP-U at equimolar ratio with high affinity (Kd = ∼54 nM). The interacting regions in hnRNP-U, mapped to both termini, suggest their proximity in the native protein; these are also disordered, based on PONDR (Predictor of Naturally Disordered Regions) prediction and circular dichroism spectra. Finally, depletion of hnRNP-U and NEIL1 epistatically sensitized human cells at low oxidative genome damage, suggesting that the hnRNP-U protection of cells after oxidative stress is largely due to enhancement of NEIL1-mediated repair.

Original languageEnglish (US)
Pages (from-to)34202-34211
Number of pages10
JournalJournal of Biological Chemistry
Volume287
Issue number41
DOIs
StatePublished - Oct 5 2012

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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