THE EXPRESSION OF MUTANT EPIDERMAL KERATIN CDNAS TRANSFECTED IN SIMPLE EPITHELIAL AND SQUAMOUS-CELL CARCINOMA LINES
THE EXPRESSION OF MUTANT EPIDERMAL KERATIN CDNAS TRANSFECTED IN SIMPLE EPITHELIAL AND SQUAMOUS-CELL CARCINOMA LINES
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DOI：
10.1083/jcb.105.2.791
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发表时间：
1987-08-01
影响因子：
7.8
通讯作者：
FUCHS, E
中科院分区：
文献类型：
--
作者：
ALBERS, K;FUCHS, E
我们删除了编码人类I型表皮角蛋白K14羧基末端部分的cDNA序列，并研究了在简单上皮细胞和鳞状细胞癌细胞系中强迫这些突变体表达的分子后果。为了跟踪我们的突变产物在转染细胞中的表达，我们用编码神经肽P物质抗原结构域的序列标记了K14编码序列的3”末端。使用DNA转染和免疫组织化学（使用针对P物质的抗体），我们已经确定了一系列突变，这些突变对转染细胞的内源性角蛋白丝网络具有广泛的形态学影响。缺少K14的大部分非螺旋羧基末端结构域的突变体可以整合到内源性角蛋白丝中，而不会对网络产生任何可见的扰动。相比之下，突变体中心α - α的310个氨基酸中只有10个缺失。多肽的-螺旋结构域导致角蛋白网络的明显改变。在某些情况下，角蛋白的整个细胞骨架网络被破坏，没有留下8纳米细丝的证据。这些结果揭示了新合成的亚基和先前存在的角蛋白丝之间存在动态交换。
We have deleted cDNA sequences encoding portions of the carboxy-terminal end of a human type I epidermal keratin K14, and examined the molecular consequences of forcing the expression of these mutants in simple epithelial and squamous cell carcioma lines. To follow the expression of our mutant products in transfected cells, we have tagged the 3'' end of the K14 coding sequence with a sequence encoding an antigenic domain of the neuropeptide substance P. Using DNA transfection and immunohistochemistry (with an antibody against substance P), we have identified a collection of mutants that have a wide range of morphological effects on the endogenous keratin filament networks of transfected cells. Mutants that are missing most of the nonhelical carboxy-terminal domain of K14 incorporate into the endogenous keratin filaments without any visible perturbations on the network. In contrast, mutants that are missing as few as 10 of the 310 amino acids of the central .alpha.-helical domain of the polypeptide cause gross alterations in the keratin network. In some cases, the entire cytoskeletal network of keratins was disrupted, leaving no evidence of 8-nm filaments. These results reveal the existence of a dynamic exchange between newly synthesized subunits and preexisting keratin filaments.