Enhanced S phase delay and inhibition of replication of an undamaged shuttle vector in UVC-irradiated xeroderma pigmentosum variant

Xeroderma pigmentosum variant (XP-V) cells are defective in bypass replicat ion of UVC-induced thymine dimers in DNA because they lack a novel DNA poly merase (polymerase eta). In this study the effects of WC on S phase cells w ere compared in fibroblasts derived from normal donors (IDH4) and XP-V pati ents (CTag) and immortalized by expression of the SV40 large T antigen. The se transformed fibroblasts did not activate the G(1) checkpoint or inhibit replicon initiation when damaged by UVC or gamma -rays, The transformed W-V cells (CTag) retained the increased sensitivity to UVC-induced inhibition of DNA strand growth previously observed with their diploid counterpart. Ce ll cycle progression analyses showed that CTag cells displayed a stronger S phase delay than transformed fibroblasts from normal individuals (IDH4) af ter treatment with only 2 J/m(2) UVC, Low doses of UVC also caused a lag in CTag cell proliferation. The extent of replication of an episomal DNA (pSV 011), not previously exposed to radiation, was measured after the host cell s were irradiated with 1-3 J/m(2) UVC, Replication of pSV011 was barely aff ected in irradiated IDH4 cells. Plasmid replication was inhibited by 50% in irradiated CTag cells and this inhibition could not be accounted for by in creased killing of host cells by UVC, These results suggest that even in tr ansformed cells WC induces DNA damage responses that are reflected in trans ient cell cycle arrest, delay in proliferation and inhibition of episomal D NA replication. These responses are enhanced in CTag cells, presumably beca use of their bypass replication defect, The accumulation of replication com plexes blocked at thymine dimers and extended single-stranded regions in ch romosomal DNA might sequester replication factors that are needed for plasm id and chromosomal replication. Alternatively, aberrant replication structu res might activate a signal transduction pathway that down-regulates DNA sy nthesis.