Studies on cytokines and proteases which regulate UV-induced mutation in human cells

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In order to clarify molecular mechanisms which regulate mutability in UV(mainly 250nm wavelength) - irradiated human cells, particularly those regulated by proteases and cytokines.1. Partial purification and characterization of UV-induced proteases were preformed and protease samples were obtaind from cell lysates of variant cells with increased levels of the induced protease activity. The samples contained molecules with about 20,000 Mr, the activity of which is inhibited by antipain but enhanced by ALLNa1.2. Using PCR-differential display, we have searched for genes expressed specifically in human interferon (HuIFN)- beta -treated Cockayne syndrome (CS) fibroblast cells. The cDNAs of six clones were similar to expression tagged sequences from unknown genes in databases and the remaining one was identical to the cDNA of the xeroderma pigmentosum XPG gene. Together with our previous finding of increased resistance to ultraviolet (UV) cell-killing of CS cells pretreated with HuIFN- beta … More prior to UV irradiation, suggest that XPG might be one of the genes possibly involved in the HuIFN- beta-induced UV-resistance.3. UV-resistant UV'-10 cells were established from UV-sensitive human RSb cells. We searched here for genes expressed differentially between UV'-10 and RSb cells using a differencial display method to elucidate the molecular mechanisms underlying the UV-resistance. Thirteen candidate cDNA fragment were obtained from 514 mRNA species first screened. Among the cDNA fragments, 3 revealed increased mRNA expression in UV'-10 cells compared with RSb cells by Northern analysis. Nucleotide sequence analysis identified 2 of the 3 cDNA fragments as encoding nucleophosimin (NPM) and T-plastin. The expression of NPM mRNA was induced after UV irradiation in UV'-10 cells but not in RSb cells, whereas irradiation did not affect the expression of T-plastin mRNA.UV'-10 cells transfected with antisense cDNA for NPM mRNA and possibly T-plastin genes may contribute to the incresed resistance to UV cell-killing, at least in the human cells tested. Less