Spartan/C1orf124, a Reader of PCNA Ubiquitylation and a Regulator of UV-Induced DNA Damage Response

Abstract

SUMMARY PCNA is a key component of DNA replication and repair machineries. DNA damage-induced PCNA ubiquitylation serves as a molecular mark to orchestrate postreplication repair. Here, we have identified and characterized Spartan, a protein that specifically recognizes ubiquitylated PCNA and plays an important role in cellular resistance to UV radiation. In vitro, Spartan engages ubiquitylated PCNA via both a PIP box and a UBZ domain. In cells, Spartan is recruited to sites of UV damage in a manner dependent upon the PIP box, the UBZ domain, and PCNA ubiquitylation. Furthermore, Spartan colocalizes and interacts with Rad18, the E3 ubiquitin ligase that modifies PCNA. Surprisingly, while Spartan is recruited by ubiquitylated PCNA, knockdown of Spartan compromised chromatin association of Rad18, monoubiquitylation of PCNA, and localization of Pol η to UV damage. Thus, as a “reader” of ubiquitylated PCNA, Spartan promotes an unexpected feed-forward loop to enhance PCNA ubiquitylation and translesion DNA synthesis.

Proliferating Cell Nuclear Antigen (PCNA)-binding Protein C1orf124 Is a Regulator of Translesion Synthesis

“…Our findings agree with the observations in a recent study characterizing the function of C1orf124 in the UV light-induced damage response (39). However, the precise mechanism by which C1orf124 confers cellular resistance to UV damage is still unclear.…”

The ADP-ribosyltransferase PARP10/ARTD10 Interacts with Proliferating Cell Nuclear Antigen (PCNA) and Is Required for DNA Damage Tolerance

“…In conclusion, PARP10 seem to be part of a positive feedback loop that recognizes and amplifies the PCNA ubiquitination signal. This is strikingly similar to the situation previously described for Spartan/C1orf124, another reader of PCNA ubiquitination involved in translesion synthesis (52)(53)(54). It will be important to investigate if Spartan and PARP10 work together.…”

The DNA-binding box of human SPARTAN contributes to the targeting of Polη to DNA damage sites

“…We mapped the DNA-binding box in the SPARTAN sequence and generated a point mutant SPARTAN that was partially defective in DNA binding. Previously, we and others described that the ubiquitylation of PCNA regulates the subnuclear targeting of SPARTAN [24][25][26]28]. We demonstrate that the DNA binding of SPARTAN is not essential for its intranuclear distribution.…”

“…We and others have described previously that Spartan has an important function at the stalled replication fork in the targeting of Polη to the site of DNA damage induced by UV irradiation [24][25][26][27][28]. Since it has also been suggested that Spartan is the human homologue of yeast WSS1, which is a DNAdependent protease [33], we decided to test whether SPARTAN can bind DNA and whether this biochemical activity affects its function after UV damage.…”

“…The targeting of SPARTAN to the stalled replication fork depends mainly on its PCNA-interacting (PIP) and ubiquitin-binding zinc finger (UBZ) domains, which can mediate its interaction with Ub-PCNA. Additionally, SPARTAN plays an important role in the correct targeting of Polη to the stalled replication fork; in its absence, UV-induced Polη foci formation is highly defective [24][25][26][27][28]. Other publications suggest that SPARTAN is responsible for the targeting of p97 (VCP, vasolinecontaining protein), which is an ATP-dependent segregase and facilitates the dissociation of Polη from the site of action [29,30].…”