CYTOPLASMIC DOMAIN OF THE 180-KD BULLOUS PEMPHIGOID ANTIGEN, A HEMIDESMOSOMAL COMPONENT - MOLECULAR AND CELL BIOLOGIC CHARACTERIZATION
CYTOPLASMIC DOMAIN OF THE 180-KD BULLOUS PEMPHIGOID ANTIGEN, A HEMIDESMOSOMAL COMPONENT - MOLECULAR AND CELL BIOLOGIC CHARACTERIZATION
复制标题
DOI：
10.1111/1523-1747.ep12616615
复制
发表时间：
1992-09-01
影响因子：
6.5
通讯作者：
JONES, JCR
中科院分区：
文献类型：
--
作者：
HOPKINSON, SB;RIDDELLE, KS;JONES, JCR
使用大疱性类天疱疮(BP)患者的血清样本，我们分离了编码半桥体180kD多肽部分的cDNA克隆，即“BP180自身抗原”。通过产生一种融合蛋白抗体来确认克隆的身份，该抗体可以识别牛舌基上皮细胞提取物和人表皮细胞提取物中的BP180。免疫电子显微镜显示，588个碱基的c DNA编码BP180的胞质片段。此外，融合蛋白的广泛物种反应性表明BP180的这一部分是高度保守的。在培养的用于共聚焦免疫荧光显微镜的人表皮细胞中，融合蛋白抗体产生与使用BP230抗体类似的点状细胞底物相关染色模式。使用原始的BP180 cDNA，我们现在已经分离到编码约1800bp的BP180的另外的cDNA克隆，其3‘序列与Giudice等人(J Clin Invest 87：734-738,199 1)中详细描述的序列重叠。对1800bp编码的氨基酸进行了二级结构分析。这些结果表明，Giudice等人描述的BP180胶原样序列(同上)从我们的融合蛋白抗体识别的BP180区域中分离出一个假定的跨膜区。事实上，BP180似乎属于一组相对罕见的蛋白质，其中N-末端位于细胞质中，C-末端位于细胞外。我们详细介绍了一些支持这一假说的初步生化实验。我们讨论了BP180和BP230在半桥粒中的可能功能。
Using a serum sample of a bullous pemphigoid (BP) patient we have isolated a cDNA clone encoding a portion of a 180-kD polypeptide component of the hemidesmosome, the "BP180 autoantigen." The identity of the clone was confirmed by the generation of a fusion protein antibody that recognizes BP180 in both a basal epithelial cell extract of bovine tongue and extracts of human epidermal cells. Immunoelectron microscopy indicates that the 588-bp cDNA encodes a cytoplasmic fragment of BP180. Furthermore, the wide species reactivity of the fusion protein suggests that this portion of BP180 is highly conserved. In cultured human epidermal cells processed for confocal immunofluorescence microscopy, the fusion protein antibody generates a punctate cell substrate-associated staining pattern that is similar to that seen using BP230 antibodies. Using the original BP180 cDNA we have now isolated additional cDNA clones encoding approximately 1800bp of BP180 the 3' sequence of which overlaps with the sequence detailed in Giudice et al (J Clin Invest 87:734 - 738, 199 1). Secondary structural analyses have been undertaken on the predicted amino acids encoded by the 1800bp. These suggest that the collagen-like sequences of BP180 described by Giudice et al (ibid.) are separated by a putative transmembrane region from the domain of BP180 recognized by our fusion protein antibody. Indeed, BP180 appears to belong to a relatively rare group of proteins in which the N-terminus is located in the cytoplasm and the C-terminus is extracellular. We detail some preliminary biochemical experiments in support of this hypothesis. We discuss possible functions of BP180 and BP230 in the hemidesmosome.