ROLE OF THE HUMAN Y-BOX-BINDING PROTEIN YB-1 IN CELLULAR-SENSITIVITY TO THE DNA-DAMAGING AGENTS CISPLATIN, MITOMYCIN-C, AND ULTRAVIOLET-LIGHT

The Y box-binding protein (YB-1) binds to DNA sequences, present in th e control regions of many genes, that contain an inverted CCAAT box. T he binding activity of a nuclear factor, designated MDR-NF1, to an inv erted CCAAT box in the promoter of the multidrug resistance 1 (MDR1) g ene has previously been shown to be increased in nuclear extracts of c ells exposed to UV radiation or various anticancer agents, The MDR-NF1 cDNA has now been cloned by screening a human colon library with an a ctive fragment of the MDR1 promoter, The amino acid sequence encoded b y the cloned cDNA was identical to that of YB-1, Northern blot analysi s revealed that YB-1 mRNA was present in all human tissues examined, R abbit antibodies were generated against synthetic peptides correspondi ng to YB-1, and indirect immunofluorescence microscopy with these anti bodies showed that the concentration of YB-1 in all cisplatin-resistan t cell lines examined was higher than that in the respective drug-sens itive parental cells. Transfection of human epidermoid cancer KB cells with a YB-1 antisense construct established two cell Lines with reduc ed concentrations of YB-1. These transfectants showed increased sensit ivity to cisplatin, mitomycin C, and UV radiation but not to vincristi ne, doxorubicin, camptothecin, or etoposide, Thus, YB-1 may protect ce lls from the cytotoxic effects of agents that induce cross-linking of DNA, suggesting a novel function of this ancestor DNA-binding protein.