DNA damage responses protect xeroderma pigmentosum variant from UVC-induced clastogenesis

Abstract

Lack of DNA polymerase eta and the attendant defect in bypass replication of pyrimidine dimers induced in DNA by ultraviolet light (UV) underlie the enhanced mutagenesis and carcinogenesis observed in xeroderma pigmentosum variant (XP-V). We investigated whether diploid XP-V fibroblasts growing in culture are also more susceptible to UV-induced clastogenesis than normal human fibroblasts (NHF). This study utilized diploid fibroblasts immortalized by the ectopic expression of human telomerase. The cell lines displayed checkpoint responses to DNA damage comparable with those measured in the parental strains. Shortly after exposure to low doses of UVC (< or =4 J/m2), XP-V cells accumulated daughter strand gaps in excess of normal controls (>25-fold). Daughter strand gaps generated in UV-irradiated S phase cells are potential precursors of chromatid-type chromosomal aberrations. Nonetheless, chromatid-type chromosomal aberrations were only 1.5 to 2 times more abundant in XP-V than in NHF exposed to the same UVC dose. XP-V cells, however, displayed S phase delays at lower doses of UVC and for longer periods of time than NHF. These results support the hypothesis that aberrant DNA structures activate S phase checkpoint responses that increase the time available for postreplication repair. Alternatively, cells that cannot be properly repaired remain permanently arrested and never reach mitosis. These responses protect human cells from chromosomal aberrations, especially when other pathways, such as accurate lesion bypass, are lost.

Caffeine and human DNA metabolism: the magic and the mystery

“…The velocity sedimentation methodology used to determine the steady-state distribution of sizes of nascent DNA 30-45 min after irradiation of log phase cultures with either 1 J/m 2 UVC or 1.5 Gy IR has been described previously [23,29].…”

“…A system of post-replication repair that includes efficient translesion synthesis of cyclobutane pyrimidine dimers by DNA pol η also provides a measure of protection. Accurate translesion synthesis by DNA pol η reduces mutagenesis by a factor of 4 [20], and decreases clastogenesis and cytotoxicity by a factor of two-fold or less [20,23]. Caffeine is also known to inhibit the repair of gaps in daughter-strand DNA in UVC-damaged cells [24].…”

p53 suppression overwhelms DNA polymerase η deficiency in determining the cellular UV DNA damage response

“…XP-V cells immortalized with hTERT show high frequencies of DNA breaks but few frank chromosomal anomalies [43]. These observations suggest that checkpoint function protects the broken DNA from generating major chromosomal changes.…”

“…In XP-V cells that lack the translesional DNA Pol η, UV damage to DNA causes an asymmetric replication fork arrest due to a replication block on the leading strand of DNA [41][42][43][44][45][46]. These structures trigger the activation of intra-S-phase checkpoints [44].…”

“…Each of the three means of inactivating p53 are distinct, although all appear to destabilize the S phase in UV damaged cells and contribute to higher levels of DNA breakage (Fig 4) and increased importance of the ATR signal transduction pathway. XP-V cells transformed with SV40 uniquely show increased UV sensitivity when ATR is inhibited by caffeine [7,35,43]; this may be due to the elevated levels of p53 in equilibrium with large T that mimic high levels of damage and induce an apoptotic response [28]. XP-V cells transformed by HPV16(E6/E7) are highly UV sensitive without further sensitization [23], suggesting that there are additional targets for the E6 ubiquitin ligase that play supporting roles in the UV response.…”