| KAI1/CD82, a recently tumor-metastasis suppressor gene, is located on human chromosome 11p 11.2 and encodes a protein of 267 amino acids. KAI1/CD82 protein belongs to a structurally distinct superfamily of membrane glycoproteins—TM4SF (transmembrane 4 superfamily or tetraspan superfamily). TM4SF proteins have four highly conserved hydrophobic regions, presumed to be transmembrane domains, and one large extracellular N-glycosylated domain. The membrane location and extensive glycosylation of these proteins suggest that they function in cell-cell interactions and cell-extracellular matrix interactions, both of which are important in tumor invasion and metastasis. KAI1/CD82 gene is a metastasis suppressor gene found in prostate cancer, while nm23 gene, with potential tumor-suppressor function, had been reported to be negatively correlated with metastasis potential in certain tumors. To further investigate metastasis suppressor effects of KAI1/CD82 gene, we applied RT-PCR, Northern blots analysis, assay for methylation status of CpG island and PCR-SSCP to analyse expression of KAI1 gene in threegroups, eight kinds of human cancer cell lines including prostate, lung and melanoma with different metastasis potential and mechanism of its reduced expression. By means of in situ hybridization , we studied the correlation between expression of KAI1/CD82 gene and metastatic behavior of esophageal cancer, we utilized Northern blots analysis to investigate expression of nm23 in the eight tumor cell lines with different metastatic potential and compared metastasis suppressor effect of KAI1/CD82 gene with that of nm23 gene. The results were summarized as follows:1. RT-PCR: The KAI1 mRNA was found expressed in all the eight human cancer cell lines PC3, PC3M, PAa, PG, WM35, WM1341b, WM983a, WM451.2. Northern blots analysis: In metastatic prostate cancer cell lines PC3 and PC3M, KAI1/CD82 mRNA expressions were obviously downregulated (integral light density value are 0.0319 and 0.0266 respectively); In nonmetastatic lung cancer cell line PAa, high expression of KAI1 mRNA was shown, but in highly metastatic lung cancer cell line PG, expression of KAI1 mRNA was low and expression level in PAa is ten times higher than that in PG (integral light density value are 0.7313 and 0.0549 respectively); Among the human melanoma cell lines with different invasive and metastatic potential, KAI1/CD82 mRNA expressions showed no obvious difference (integral light density value: WM35 0.1798, WM451 0.1800, WM983a 0.1501, WM1341b 0.1582).3. Assay for methylation status: We employed a PCR-basedapproach to assess methylation status at specific site in the promoter region of KAIl DNA. DNA was digested with a methylation-sensitive enzyme (Smal) prior to PCR amplification. But we demonstrated no presence of CCm5CGGG in a portion of the 5'-promoter region of KAIl gene.4. PCR-SSCP: Exon3-9 of KAI1/CD82 gene were analysed by nonisotopic PCR-SSCP. The results indicated that exon 7 of PC3, PC3M and PG cell lines showed abnormal bands or mutation.5. In situ hybridization: KAI1/CD82 mRNA expressions showed no difference between esophageal cancer with and without lymph node metastasis and was not correlated with differentiation of esophageal cancer.6. nm23 Northern blots analysis: nm23 mRNAs were expressed by all eight human cancer cell lines and no obvious difference in expression level among these cell lines was observed, despite they possess different metastasis potential.Conclusions are as follows:1. Decreased KAI1/CD82 expression may be associated with enhanced metastatic potential in human prostate cancer and lung cancer PG cell lines. Losing of KAI1/CD82 gene function correlates with the formation of metastasis in human prostate and lung cancer. It is shown that KAI1/CD82 gene may be a key gene in inhibiting human prostate and lung cancer metastasis.2. KAI1/CD82 gene may be not a key metastasis suppressor gene for human melanoma. That is, KAIl gene is not associated with...
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