Ultraviolet A (UVA) radiation induces DNA damage both directly, by forming cyclobutane pyrimidine dimers (CPDs), and indirectly, by generating oxidative stress. Cells rely on nucleotide excision repair (NER) and translesion synthesis (TLS) to tolerate these lesions. Xeroderma pigmentosum variant (XP-V) cells, deficient in DNA polymerase eta (pol eta), exhibit a heightened risk of skin cancer due to impaired TLS. While XP-V patients are considered NER-proficient, our findings challenge this assumption by demonstrating that UVA-induced oxidative stress impaired NER activity, leading to increased C > T transitions at CPD sites. Whole-exome sequencing of UVA-irradiated XP-V cells revealed a substantial rise in mutations, with a distinct C > T signature characteristic of defective CPD repair. Notably, pretreatment with the antioxidant N-acetylcysteine (NAC) mitigated this effect, reducing C > T transitions through enhanced NER function and decreasing C > A transversions via its antioxidant properties. These results redefine the mutagenic landscape of XP-V cells, revealing that oxidatively generated damage to NER proteins-rather than TLS deficiency alone-contributes to their elevated mutation burden. Our findings suggest that antioxidant strategies may partially protect XP-V patients from UVA-driven mutagenesis enhancing the cells' DNA repair capacity, ultimately reducing skin cancer and contributing to better overall health outcomes.

DNA polymerase (pol) η is vital for accurately replicating DNA opposite ultraviolet light (UV)-induced cyclobutane pyrimidine dimers and cisplatin-induced intrastrand purine crosslinks. While human POLH deficiency is linked to the disease xeroderma pigmentosum variant, the functional consequences of germline and somatic POLH variants remain largely unexplored. We characterized nine nonsynonymous POLH germline variants, five of which have also been found in various tumors. Enzyme activity was first assessed using recombinant pol η (residues 1-432) proteins. Variants F17S, C227Y, and R356X displayed substantially reduced or nearly abolished polymerase activity opposite cis-syn cyclobutane thymine dimer (CTD) compared to the wild-type. Cellular effects were then evaluated in POLH-deficient human embryonic kidney (HEK) 293 cells. Unlike cells transfected with wild-type POLH, cells transfected with F17S, R81C, C227Y, or R356X variants failed to rescue UV- and cisplatin-sensitivity. Interestingly, the R81C variant protein was undetectable in transfected cells. Further steady-state kinetic analysis revealed that the F17S, C227Y, and R356X variants had 3- to 5000-fold reductions in kcat/Km values for correct dATP insertion opposite CTD, while the R81C variant exhibited kinetics comparable to the wild-type enzyme. CRISPR/Cas9-mediated knock-in of the R81C variant in HEK 293T cells was associated with significantly impaired pol η protein expression and increased cisplatin sensitivity. Notably, R81C and R356X mutations have been reported in skin cancer samples. These findings suggest that R81C, F17S, C227Y, and R356X POLH variants-underexpressed or hypoactive-may be insufficient to protect cells from UV radiation and cisplatin, highlighting their potential implications for individual susceptibility to UV and cisplatin damage.

Thirteen children with xeroderma pigmentosum variant C were evaluated using the Dermoscopic Photoaging Assessment Scale (DPAS), the Glogau scale, and the Sun Protection Behavior Scale (SPBS). Most patients exhibited signs of epidermal photoaging, with pigmentary and vascular changes and poor sun protection behavior (mean SPBS score: 18.92 ± 5.69). The mean DPAS score of 13.15 correlated with the Glogau scale and ocular severity score. Sun protection behavior improved significantly at 6 months after repeated reinforcement of sun protection measures (35 ± 5.68).

DNA damage tolerance pathways that allow for the completion of replication following fork arrest are critical in maintaining genome stability during cell division. The main DNA damage tolerance pathways include strand switching, replication fork reversal and translesion synthesis (TLS). The TLS pathway is mediated by specialised DNA polymerases that can accommodate altered DNA structures during DNA synthesis, and are important in allowing replication to proceed after fork arrest, preventing fork collapse that can generate more deleterious double-strand breaks in the genome. TLS may occur directly at the fork, or at gaps remaining behind the fork, in the process of post-replication repair. Inactivating mutations in the human POLH gene encoding the Y-family DNA polymerase Pol η causes the skin cancer-prone genetic disease xeroderma pigmentosum variant (XPV). Pol η also contributes to chemoresistance during cancer treatment by bypassing DNA lesions induced by anti-cancer drugs including cisplatin. We review the current understanding of the canonical role of Pol η in translesion synthesis following replication arrest, as well as a number of emerging non-canonical roles of the protein in other aspects of DNA metabolism.