Structural Insight into DNA-Dependent Activation of Human Metalloprotease Spartan

Li, Faxiang; Raczynska, J.E.; Chen, James Z.; Yu, Hongtao

doi:10.1016/j.celrep.2019.02.082

Cited by 45 publications

(55 citation statements)

References 52 publications

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“…Our data suggest that the p97-TEX264 complex enables TOP1cc proteolysis by SPRTN. This conclusion is supported by a recent strutural analysis of SPRTN's active site 61 . The catalytic core of SPRTN is located within a narrow groove that is only accessible to small, flexible peptides, such as the disordered tails of histones.…”

Section: Discussionsupporting

confidence: 57%

“…A recent structural study demonstrated that the protease active site of SPRTN is located within a narrow groove that is only accessible to flexible peptide structures and not globular proteins HA S1 X X Denature: SDS/95 °C TEX264 TEX264-associated proteins TEX264 S1 + ARTICLE such as TOP1 61 . Therefore, large DPCs, such as TOP1ccs, likely need to undergo remodelling before their cleavage.…”

Section: Resultsmentioning

confidence: 99%

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TEX264 coordinates p97- and SPRTN-mediated resolution of topoisomerase 1-DNA adducts

et al. 2020

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Eukaryotic topoisomerase 1 (TOP1) regulates DNA topology to ensure efficient DNA replication and transcription. TOP1 is also a major driver of endogenous genome instability, particularly when its catalytic intermediate-a covalent TOP1-DNA adduct known as a TOP1 cleavage complex (TOP1cc)-is stabilised. TOP1ccs are highly cytotoxic and a failure to resolve them underlies the pathology of neurological disorders but is also exploited in cancer therapy where TOP1ccs are the target of widely used frontline anti-cancer drugs. A critical enzyme for TOP1cc resolution is the tyrosyl-DNA phosphodiesterase (TDP1), which hydrolyses the bond that links a tyrosine in the active site of TOP1 to a 3' phosphate group on a single-stranded (ss)DNA break. However, TDP1 can only process small peptide fragments from ssDNA ends, raising the question of how the~90 kDa TOP1 protein is processed upstream of TDP1. Here we find that TEX264 fulfils this role by forming a complex with the p97 ATPase and the SPRTN metalloprotease. We show that TEX264 recognises both unmodified and SUMO1-modifed TOP1 and initiates TOP1cc repair by recruiting p97 and SPRTN. TEX264 localises to the nuclear periphery, associates with DNA replication forks, and counteracts TOP1ccs during DNA replication. Altogether, our study elucidates the existence of a specialised repair complex required for upstream proteolysis of TOP1ccs and their subsequent resolution.

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Section: Discussionsupporting

confidence: 57%

Section: Resultsmentioning

confidence: 99%

TEX264 coordinates p97- and SPRTN-mediated resolution of topoisomerase 1-DNA adducts

et al. 2020

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“…ArdC is structurally similar to DNA-binding dependent metalloproteases involved in the maintenance of genetic stability such as Spartan or IrrE. Human Spartan protein cleaves DNA-protein crosslinks [15] meanwhile IrrE plays a central regulatory role in DNA protection and repair pathways in response to radiation [16]. ArdC structure differs from other known plasmidcoded antirestriction proteins.…”

Section: Discussionmentioning

confidence: 99%

“…The closest structural homologues were the DNA binding metalloproteases Spartan (PDB: 6MDW; Z-score: 6.4) and IrrE (PDB: 3DTE; Zscore: 5.5). Spartan is a protein involved in the cleavage of proteins irreversibly crosslinked to DNA to preserve this way genome stability [15]. IrrE protects D. radiodurans from UV radiation DNA damage by proteolysis of a transcriptional regulator, DdrO, involved in SOS response [16].…”

Section: Ardc Is Needed In Recipient Cellsmentioning

confidence: 99%

ArdC protein overpasses the recipient hsdRMS restriction system broadening conjugation host range

González-Montes

Campo

Cruz

et al. 2019

Preprint

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Plasmids, when transferred by conjugation, must overpass restriction-modification systems of the recipient cell. We demonstrate that protein ArdC, encoded by broad host range plasmid R388, was required for conjugation from Escherichia coli to Pseudomonas putida, but not from E. coli to E. coli. Surprisingly, expression of ardC was required in the recipient cells, but not in the donor cells. Besides, ardC was not required for conjugation if the hsdRMS system was deleted in P. putida recipient cells. Thus, ArdC has antirestriction activity against HsdRMS system, and consequently broadens R388 plasmid host range. The crystal structure of ArdC was solved both in the absence and in the presence of Mn 2+ . ArdC is composed of a non-specific ssDNA binding N-terminal domain and a C-terminal metalloprotease domain, although the metalloprotease activity is not needed for antirestriction function. We also observed by RNA-seq that ArdC-dependent conjugation triggers an SOS response in the P. putida recipient cells. Our findings give new insights, and open new questions, into the antirestriction strategies developed by plasmids to counteract bacterial restriction strategies. ResultsardC is required for R388 conjugation from E. coli to P. putida R388 plasmid is composed by three functional sectors ( Fig EV1). One for general maintenance (modules of replication, stable inheritance and establishment) located in the leading region, a sector for Ab R and integration and a third one for conjugation (modules of DNA transfer replication and mating pore formation) [11]. We expected the stable inheritance and establishment region to be required in interspecies conjugation. pSU2007, a Kn R R388 derivative, is transferred with different efficiencies from E. coli BW27783-Nx R to other bacteria ( Fig EV2). All conjugation experiments were done at the recipient cells optimal growing temperature except for P. putida KT2440 to which we observed that the conjugation was more effective at 37 °C. The transfer of pIC10 (R388∆kfrA-orf14), an R388 derivative without the stability and maintenance region, is similar to pSU2007 in all species but in P. putida KT2440 where the conjugation frequencies dropped around 1000 times. We have performed the conjugation experiments at both the optimal E. coli growing temperature (37 °C) or P. putida growing temperature (30 °C). In the stability and maintenance gene region deleted in pIC10 there are 13 genes that code for proteins homologous to some with predicted function of: fertility inhibition in a different system (kfrA, nuc1, nuc2 and osa) [12], hypothetical proteins of unknown function (orf7, orf8, orf9, orf12 and orf14), transcriptional regulators (ardk and klcB), ssDNA binding

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“…Furthermore, the protease activity of SPRTN is tightly regulated with a switch that depends on its DNA binding, ubiquitination, and autocleavage [14][15][16][17][18]. Both single-and double-stranded DNA can activate the protease activity of SPRTN, with single-stranded DNA being more effective [14][15][16]107]. SPRTN can be monoubiquitinated, but only unmodified SPRTN binds to chromatin [15].…”

Section: Proteolysis-dependent Repair Mechanisms Targeting Cross-linkmentioning

confidence: 99%

DNA–protein cross-link repair: what do we know now?

2020

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When a protein is covalently and irreversibly bound to DNA (i.e., a DNA-protein cross-link [DPC]), it may obstruct any DNA-based transaction, such as transcription and replication. DPC formation is very common in cells, as it can arise from endogenous factors, such as aldehyde produced during cell metabolism, or exogenous sources like ionizing radiation, ultraviolet light, and chemotherapeutic agents. DPCs are composed of DNA, protein, and their cross-linked bonds, each of which can be targeted by different repair pathways. Many studies have demonstrated that nucleotide excision repair and homologous recombination can act on DNA molecules and execute nuclease-dependent DPC repair. Enzymes that have evolved to deal specifically with DPC, such as tyrosyl-DNA phosphodiesterases 1 and 2, can directly reverse cross-linked bonds and release DPC from DNA. The newly identified proteolysis pathway, which employs the proteases Wss1 and SprT-like domain at the N-terminus (SPRTN), can directly hydrolyze the proteins in DPCs, thus offering a new venue for DPC repair in cells. A deep understanding of the mechanisms of each pathway and the interplay among them may provide new guidance for targeting DPC repair as a therapeutic strategy for cancer. Here, we summarize the progress in DPC repair field and describe how cells may employ these different repair pathways for efficient repair of DPCs.

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Structural Insight into DNA-Dependent Activation of Human Metalloprotease Spartan

Cited by 45 publications

References 52 publications

TEX264 coordinates p97- and SPRTN-mediated resolution of topoisomerase 1-DNA adducts

TEX264 coordinates p97- and SPRTN-mediated resolution of topoisomerase 1-DNA adducts

ArdC protein overpasses the recipient hsdRMS restriction system broadening conjugation host range

DNA–protein cross-link repair: what do we know now?

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