Shekels L L, Hunninghake D A, Tisdale A S, Gipson I K, Kieliszewski M, Kozak C A, Ho S B
Department of Medicine, University of Minnesota and VA Medical Center, Minneapolis, MN 55417, USA.
Biochem J. 1998 Mar 15;330 ( Pt 3)(Pt 3):1301-8. doi: 10.1042/bj3301301.
Mucin glycoproteins are a heterogeneous family of high-molecular-mass, heavily glycosylated proteins differentially expressed in epithelial tissue of the gastrointestinal, reproductive and respiratory tracts. We report here the cloning of a mouse caecal mucin (MCM). Amino acid analysis of purified MCM revealed a high content of serine (10.8%) and threonine (25.1%). Antibodies against deglycosylated MCM were prepared for immunohistochemical analysis and for screening a mouse caecal cDNA library. Immunohistochemical analysis showed strong staining of goblet cells and patchy staining of surface columnar cells in the duodenum, small intestine, caecum, colon and rectum. Screening of a mouse caecal cDNA library yielded clones containing tandem repeats of 18 bp with two predominant peptide sequences of TTTADV and TTTVVV. The tandem repeat domain is followed by 1137 bp of non-repetitive sequence and 521 bp of 3' untranslated sequence prior to the poly(A) tail. Two cysteine-rich regions lie within the 3' non-repetitive domain. The arrangement of the cysteines within these regions corresponds to epidermal growth factor-like domains. Following the second cysteine-rich region is a stretch of 19 hydrophobic amino acids which may act as a transmembrane domain or allow for interaction with hydrophobic molecules. Northern blot analysis indicates the mRNA is approximately 13.5 kb with greatest expression in the caecum and lesser amounts in the colon and small intestine. No MCM message is found in mouse stomach, trachea, lung, kidney, oesophagus or pancreas. In situ hybridization studies show that MCM message is expressed at the tips of villi in the intestine and in the upper crypts and surface cells of the caecum and colon. Chromosomal analysis assigns this gene to mouse chromosome 5 in a region of conserved linkage with human chromosome 7, the location of the human MUC3 gene. We conclude that we have identified a mouse caecal mucin which represents the mouse homologue of human MUC3. The mouse MUC3 cDNA sequence suggests that it is a novel non-polymerizing mucin which may participate in membrane or intermolecular interactions through its 3' non-repetitive region.
黏蛋白糖蛋白是一类高分子量、高度糖基化的异质性蛋白家族，在胃肠道、生殖道和呼吸道的上皮组织中差异表达。我们在此报告小鼠盲肠黏蛋白（MCM）的克隆。对纯化的MCM进行氨基酸分析显示，丝氨酸（10.8%）和苏氨酸（25.1%）含量很高。制备了针对去糖基化MCM的抗体，用于免疫组织化学分析和筛选小鼠盲肠cDNA文库。免疫组织化学分析显示，十二指肠、小肠、盲肠、结肠和直肠中的杯状细胞染色强烈，表面柱状细胞呈斑片状染色。对小鼠盲肠cDNA文库的筛选产生了含有18 bp串联重复序列的克隆，其中有两个主要的肽序列TTTADV和TTTVVV。串联重复结构域之后是1137 bp的非重复序列和521 bp的3'非翻译序列，然后是聚腺苷酸尾巴。两个富含半胱氨酸的区域位于3'非重复结构域内。这些区域内半胱氨酸的排列与表皮生长因子样结构域相对应。在第二个富含半胱氨酸的区域之后是一段19个疏水氨基酸的序列，它可能作为跨膜结构域或允许与疏水分子相互作用。Northern印迹分析表明，mRNA约为13.5 kb，在盲肠中表达最高，在结肠和小肠中表达较少。在小鼠胃、气管、肺、肾、食管或胰腺中未发现MCM信息。原位杂交研究表明，MCM信息在小肠绒毛尖端以及盲肠和结肠的上隐窝和表面细胞中表达。染色体分析将该基因定位到小鼠5号染色体上与人类7号染色体保守连锁的区域，即人类MUC3基因的位置。我们得出结论，我们已经鉴定出一种小鼠盲肠黏蛋白，它代表了人类MUC3的小鼠同源物。小鼠MUC3 cDNA序列表明，它是一种新型的非聚合黏蛋白，可能通过其3'非重复区域参与膜或分子间相互作用。
