RESUMEN
Repair of DNA interstrand crosslinks (ICLs) predominantly involves the Fanconi anemia (FA) pathway and homologous recombination (HR). The HR repair system eliminates DNA double strand breaks (DSBs) that emerge during ICLs removal. The current study presents a novel cell line, CLV8B, representing a new complementation group of Chinese hamster cell mutants hypersensitive to DNA crosslinking factors. CLV8B exhibits increased sensitivity to various DNAdamaging agents, including compounds leading to DSBs formation (bleomycin and 6thioguanine), and is extremely sensitive to poly (ADP-ribose) polymerase inhibitor (>400fold), which is typical for HRdefective cells. In addition, this cell line exhibits a reduced number of spontaneous and induced sister chromatid exchanges, which suggests likely impairment of HR in CLV8B cells. However, in contrast to other known HR mutants, CLV8B cells do not show defects in Rad51 foci induction, but only slight alterations in the focus formation kinetics. CLV8B is additionally characterized by a considerable chromosomal instability, as indicated by a high number of spontaneous and MMCinduced chromosomal aberrations, and a twice as large proportion of cells with abnormal centrosomes than that in the wild type cell line. The molecular defect present in CLV8B does not affect the efficiency and stabilization of replication forks. However, stalling of the forks in response to replication stress is observed relatively rarely, which suggests an impairment of a signaling mechanism. Exposure of CLV8B to crosslinking agents results in Sphase arrest (as in the wild type cells), but also in larger proportion of G2/Mphase cells and apoptotic cells. CLV8B exhibits similarities to HR and/or FAdefective Chinese hamster mutants sensitive to DNA crosslinking agents. However, the unique phenotype of this new mutant implies that it carries a defect of a yet unidentified gene involved in the repair of ICLs.
