It has been shown that the Chinese hamster cell mutant V-C8 is sensitive to different DNA damaging agents, such as mitomycin C (MMC), alkylating agents, UV light, and X-rays. We found that V-C8 is also sensitive to the following radiomimetic agents: bleomycin (approximately 2-fold, based on D10 values), H2O2 (approximately 2-fold), streptonigrin (approximately 11-fold), and etoposide (approximately 8-fold). Two independent spontaneous MMC-resistant revertants isolated from V-C8 cells show a level of cell killing by X-rays, EMS, and UV light which is similar to that of wild-type cells, suggesting that the observed pattern of cross-sensitivity of V-C8 cells to a wide spectrum of DNA damaging agents results from a single mutation. V-C8 cells also display radioresistant DNA synthesis following gamma-irradiation which, however, remained almost unchanged in the V-C8 revertants. The measurement of the level and rate of repair of DNA single- and double-strand breaks (SSBs and DSBs, respectively) by the DNA elution technique showed that the V-C8 mutant has a slower repair of DSBs induced by gamma-rays. The described unique phenotype of V-C8 cells suggested that V-C8 represents a novel type of mutant amongst X-ray-sensitive hamster cell mutants. To confirm this, complementation analysis with other X-ray-sensitive mutants was performed. V-C8 cells were fused with EM9, XR-1, xrs5, sxi-1, V-3, V-E5, irs3, and BLM2 mutant cells, representing different complementation groups. All the obtained hybrids regained X-ray resistance (or bleomycin resistance in the case of V-C8/BLM2 hybrids) similar to that of wild-type cells, indicating that V-C8 represents a new complementation group. The results presented indicate that V-C8 is defective in a gene involved in a pathway operating in the responses to different DNA damaging agents in mammalian cells.