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

为了阐明调节紫外线（主要是250nm波长）照射的人体细胞，特别是那些受蛋白酶和细胞因子调节的突变性的分子机制。 1．对紫外线诱导的蛋白酶进行部分纯化和表征，并从诱导蛋白酶活性水平增加的变异细胞的细胞裂解物中获得蛋白酶样品。样品中含有约 20,000 Mr 的分子，其活性被抗痛药抑制，但被 ALLNa1.2 增强。使用 PCR 差异显示，我们搜索了在人干扰素 (HuIFN)-β 处理的科凯恩综合征 (CS) 成纤维细胞中特异性表达的基因。六个克隆的 cDNA 与数据库中未知基因的表达标签序列相似，其余一个与着色性干皮病 XPG 基因的 cDNA 相同。加上我们之前发现在紫外线照射前用 HuIFN-β 预处理的 CS 细胞对紫外线 (UV) 细胞杀伤的抵抗力增强，表明 XPG 可能是可能与 HuIFN-β 诱导的紫外线抵抗有关的基因之一。3。抗紫外线 UV'-10 细胞是由对紫外线敏感的人类 RSb 细胞建立的。我们在这里使用差异显示方法搜索 UV'-10 和 RSb 细胞之间差异表达的基因，以阐明抗紫外线的分子机制。从首次筛选的514个mRNA种类中获得了13个候选cDNA片段。在 cDNA 片段中，Northern 分析显示，与 RSb 细胞相比，UV'-10 细胞中的 3 个 mRNA 表达增加。核苷酸序列分析鉴定出 3 个 cDNA 片段中的 2 个编码核磷蛋白 (NPM) 和 T-plastin。紫外线照射后，UV'-10 细胞中 NPM mRNA 的表达被诱导，但在 RSb 细胞中则不然，而照射并不影响 T-plastin mRNA 的表达。用 NPM mRNA 反义 cDNA 转染的 UV'-10 细胞和可能的 T-plastin 基因可能有助于增强对 UV 细胞杀伤的抵抗力，至少在测试的人类细胞中是这样。较少的

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