Troyanovsky S M, Leube R E, Franke W W
Division of Cell Biology, German Cancer Research Center, Heidelberg.
Eur J Cell Biol. 1992 Oct;59(1):127-37.
Among the members of the cytokeratin (CK) subfamily of intermediate filament (IF) proteins, CK 17 is remarkable as it is normally expressed in the basal cells of complex epithelia but not in stratified or simple epithelia. Because of its unusual expression pattern in normal and diseased states and because of the potential importance of CK 17 in tumor diagnosis, we have characterized the gene(s) and its cDNA-derived amino acid sequence. A cDNA clone encoding CK 17 was isolated from a HeLa cDNA library and used for the determination of the amino acid sequence, for studies of expression and for the screening of human genomic libraries. A number of lambda phage clones were isolated that covered three distinct, non-contiguous gene regions. Only one of these loci contains the functional CK 17 gene which is located only approximately 5 kbp 5'-upstream of the CK 16 gene, whereas the other two contain unprocessed CK 17 pseudogenes. Each of these genes is part of a larger CK type I gene locus the arrangement of which suggests that these genes and pseudogenes have arisen during evolution by duplication events comprising whole multigene loci. The functional CK 17 gene differs from the pseudogenes by the extent of methylation of certain DNA sequences in the 5'-upstream region. The 5 kbp CK 17 gene with 8 exons and 7 introns encodes a polypeptide of 432 amino acids with a calculated molecular weight of 48,000. Using S1-nuclease protection assays and RNAs from several cell lines we identified a single transcriptional start point 26 nucleotides down-stream from a TATA box element. Northern blot hybridization experiments showed a restricted pattern of CK 17 gene expression, supporting the notion that CK 17 synthesis is essentially regulated at the transcriptional level. From these findings and from immunohistological observations, CK 17 synthesis seems to be a marker of basal cell differentiation in complex epithelia and therefore indicative of a certain type of epithelial "stem cells".
在中间丝（IF）蛋白的细胞角蛋白（CK）亚家族成员中，CK 17很特别，因为它通常在复层上皮的基底细胞中表达，而不在复层或单层上皮中表达。由于其在正常和疾病状态下不同寻常的表达模式，以及CK 17在肿瘤诊断中的潜在重要性，我们对该基因及其cDNA推导的氨基酸序列进行了表征。从HeLa cDNA文库中分离出一个编码CK 17的cDNA克隆，并用于氨基酸序列测定、表达研究以及人类基因组文库的筛选。分离出了多个λ噬菌体克隆，它们覆盖了三个不同的、不连续的基因区域。其中只有一个位点包含功能性CK 17基因，该基因位于CK 16基因上游约5 kbp处，而另外两个包含未加工的CK 17假基因。这些基因中的每一个都是一个更大的CK I型基因位点的一部分，其排列方式表明这些基因和假基因是在进化过程中通过包含整个多基因位点的重复事件产生的。功能性CK 17基因与假基因的区别在于5'上游区域某些DNA序列的甲基化程度。这个5 kbp的CK 17基因有8个外显子和7个内含子，编码一个由432个氨基酸组成的多肽，计算分子量为48,000。使用S1核酸酶保护试验和来自几种细胞系的RNA，我们确定了一个转录起始点，位于TATA盒元件下游26个核苷酸处。Northern印迹杂交实验显示CK 17基因表达模式受限，支持CK 17合成主要在转录水平受到调控的观点。基于这些发现以及免疫组织学观察结果，CK 17合成似乎是复层上皮中基底细胞分化的一个标志物，因此可指示某种类型的上皮“干细胞”。
