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SPRTN is a mammalian DNA-binding metalloprotease that resolves DNA-protein crosslinks.

ABSTRACT: Ruijs-Aalfs syndrome is a segmental progeroid syndrome resulting from mutations in the SPRTN gene. Cells derived from patients with SPRTN mutations elicit genomic instability and people afflicted with this syndrome developed hepatocellular carcinoma. Here we describe the molecular mechanism by which SPRTN contributes to genome stability and normal cellular homeostasis. We show that SPRTN is a DNA-dependent mammalian protease required for resolving cytotoxic DNA-protein crosslinks (DPCs)- a function that had only been attributed to the metalloprotease Wss1 in budding yeast. We provide genetic evidence that SPRTN and Wss1 function distinctly in vivo to resolve DPCs. Upon DNA and ubiquitin binding, SPRTN can elicit proteolytic activity; cleaving DPC substrates and itself. SPRTN null cells or cells derived from patients with Ruijs-Aalfs syndrome are impaired in the resolution of covalent DPCs in vivo. Collectively, SPRTN is a mammalian protease required for resolving DNA-protein crosslinks in vivo whose function is compromised in Ruijs-Aalfs syndrome patients.

SUBMITTER: Lopez-Mosqueda J

PROVIDER: S-EPMC5127644 | biostudies-literature | 2016 Nov

REPOSITORIES: biostudies-literature

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SPRTN is a mammalian DNA-binding metalloprotease that resolves DNA-protein crosslinks.

Lopez-Mosqueda Jaime J Maddi Karthik K Prgomet Stefan S Kalayil Sissy S Marinovic-Terzic Ivana I Terzic Janos J Dikic Ivan I

eLife 20161117

Ruijs-Aalfs syndrome is a segmental progeroid syndrome resulting from mutations in the <i>SPRTN</i> gene. Cells derived from patients with SPRTN mutations elicit genomic instability and people afflicted with this syndrome developed hepatocellular carcinoma. Here we describe the molecular mechanism by which SPRTN contributes to genome stability and normal cellular homeostasis. We show that SPRTN is a DNA-dependent mammalian protease required for resolving cytotoxic DNA-protein crosslinks (DPCs)- ...[more]

PMID: 27852435

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Project description:Free radical attack on C1' of deoxyribose forms the oxidized abasic (AP) site 2-deoxyribonolactone (dL). In vitro, dL traps the major base excision DNA repair enzyme DNA polymerase beta (Pol?) in covalent DNA-protein crosslinks (DPC) via the enzyme's N-terminal lyase activity acting on 5'-deoxyribose-5-phosphate residues. We previously demonstrated formation of Pol?-DPC in cells challenged with oxidants generating significant levels of dL. Proteasome inhibition under 1,10-copper-ortho-phenanthroline (CuOP) treatment significantly increased Pol?-DPC accumulation and trapped ubiquitin in the DPC, with Pol? accounting for 60-70 % of the total ubiquitin signal. However, the identity of the remaining oxidative ubiquityl-DPC remained unknown. In this report, we surveyed whether additional AP lyases are trapped in oxidative DPC in mammalian cells in culture. Poly(ADP-ribose) polymerase 1 (PARP1), Ku proteins, DNA polymerase ? (Pol?), and the bifunctional 8-oxoguanine DNA glycosylase 1 (OGG1), were all trapped in oxidative DPC in mammalian cells. We also observed significant trapping of Pol?, PARP1, and OGG1 in cells treated with the alkylating agent methylmethane sulfonate (MMS), in addition to dL-inducing agents. Ku proteins, in contrast, followed a pattern of trapping similar to that for Pol?: MMS failed to produce Ku-DPC, while treatment with CuOP or (less effectively) H2O2 gave rise to significant Ku-DPC. Unexpectedly, NEIL1 and NEIL3 were trapped following H2O2 treatment, but not detectably in cells exposed to CuOP. The half-life of all the AP lyase-DPC ranged from 15-60 min, consistent with their active repair. Accordingly, CuOP treatment under proteasome inhibition significantly increased the observed levels of DPC in cultured mammalian cells containing PARP1, Ku protein, Pol?, and OGG1 proteins. As seen for Pol?, blocking the proteasome led to the accumulation of DPC containing ubiquitin. Thus, the ubiquitin-dependent proteolytic mechanisms that control Pol?-DPC removal may also apply to a broad array of oxidative AP lyase-DPC, preventing their toxic accumulation in cells.

Dataset Information

SPRTN is a mammalian DNA-binding metalloprotease that resolves DNA-protein crosslinks.

Publications

SPRTN is a mammalian DNA-binding metalloprotease that resolves DNA-protein crosslinks.

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