Stokes D G, Saitta B, Timpl R, Chu M L
Department of Biochemistry and Molecular Biology, Thomas Jefferson University, Jefferson Medical College, Philadelphia, Pennsylvania 19107.
J Biol Chem. 1991 May 5;266(13):8626-33.
We recently reported the isolation and sequencing of human cDNA clones corresponding to the alpha 3 chain of type VI collagen (Chu, M.-L., Zhang, R.-Z., Pan, T.-c., Stokes, D., Conway, D., Kuo, H.-J., Glanville, R., Mayer, U., Mann, K., Deutzmann, R., and Timpl, R. (1990) EMBO J. 9, 385-393). The study indicates that the amino-terminal globular domain of the alpha 3(VI) chain consists of nine repetitive subdomains of approximately 200 amino acid residues (N1-N9) and the gene appeared to undergo alternative splicing since some clones lacked regions encoding the N9 and part of the N3 subdomains. In the present study, we report the exon structure for the region encoding the amino-terminal globular domain of the human alpha 3(VI) chain. The nine repetitive subdomains are encoded by 10 exons spanning 26 kilobase pairs of genomic DNA. Eight of the repetitive subdomains (N2-N9) were found to be encoded by separate exons of approximately 600 base pairs each. The only exception is the N1 subdomain which is encoded by two exons of 417 and 146 base pairs. Characterization of the exon/intron structure showed that the cDNA variants were the result of splicing out of exon 9 (encoding the N9 subdomain) and part of exon 3 (encoding the N3 subdomain). Nuclease S1 analysis and the polymerase chain reaction demonstrated that exon 7 (N7 subdomain) was also subject to alternative splicing in normal skin fibroblasts. Examination of these splicing events by nuclease S1 analysis in normal fibroblasts, three different human tumor cell lines, and several human tissues showed that splicing out of exon 9 is much more efficient in normal as compared to tumor cells.
我们最近报道了与VI型胶原α3链相对应的人cDNA克隆的分离和测序（朱，M.-L.，张，R.-Z.，潘，T.-c.，斯托克斯，D.，康威，D.，郭，H.-J.，格兰维尔，R.，迈耶，U.，曼，K.，多伊茨曼，R.，和蒂姆普尔，R.（1990年）《欧洲分子生物学组织杂志》9，385 - 393）。该研究表明，α3(VI)链的氨基末端球状结构域由九个约200个氨基酸残基的重复亚结构域（N1 - N9）组成，并且该基因似乎经历了可变剪接，因为一些克隆缺少编码N9和部分N3亚结构域的区域。在本研究中，我们报道了编码人α3(VI)链氨基末端球状结构域区域的外显子结构。这九个重复亚结构域由跨越26千碱基对基因组DNA的10个外显子编码。发现八个重复亚结构域（N2 - N9）分别由约600个碱基对的外显子编码。唯一的例外是N1亚结构域，它由417和146个碱基对的两个外显子编码。外显子/内含子结构的特征表明，cDNA变体是由于外显子9（编码N9亚结构域）和外显子3的一部分（编码N3亚结构域）的剪接缺失所致。核酸酶S₁分析和聚合酶链反应表明，外显子7（N7亚结构域）在正常皮肤成纤维细胞中也经历可变剪接。通过核酸酶S₁分析在正常成纤维细胞、三种不同的人肿瘤细胞系和几种人体组织中检查这些剪接事件表明，与肿瘤细胞相比，外显子9的剪接在正常细胞中效率要高得多。
