Imbert Yoannis, Darling Douglas S, Jumblatt Marcia M, Foulks Gary N, Couzin Erica G, Steele Pamela S, Young William W
Department of Molecular, Cellular & Craniofacial Biology, School of Dentistry, University of Louisville, 501 S. Preston St., Louisville, KY 40292, USA.
Exp Eye Res. 2006 Sep;83(3):493-501. doi: 10.1016/j.exer.2006.01.031. Epub 2006 Apr 21.
Mucins are highly glycosylated proteins that are vital to the maintenance of healthy epithelial surfaces including the ocular surface. Mucins act as lubricants, protectants, and mediators of signal transduction. The majority of the O-glycosylation sites on the transmembrane mucin MUC1 are found in a highly polymorphic core region containing a variable number of tandem repeats (VNTR). MUC1 alleles can be divided into size classes that contain small (30-45) or large (60-90) numbers of repeats. Although at least 12 splice variants of MUC1 have been found in other tissues, no splice variants have been reported in human ocular surface tissues. We have used RT-PCR to identify MUC1 splice variants that were then confirmed by sequencing. We here report the presence in some samples of human cornea, conjunctiva, and lacrimal gland of MUC1/B which features canonical splicing between exons 1 and 2 and MUC1/A, a transcript that retains 27bp from the 3' end of intron 1 and is predicted to add 9 amino acids to the MUC1 sequence upstream of the tandem repeat region. Cornea and conjunctiva both contain the MUC1/SEC splice variant that lacks the transmembrane domain and, therefore, results in a soluble, secreted form of MUC1. Cornea and conjunctiva also contain MUC1/Y and MUC1/Z(X) variants that lack the tandem repeat region. Cornea, conjunctiva, and lacrimal gland also contain a previously undescribed MUC1 variant transcript, termed MUC1/YI, that retains 99bp from the 5' end and 27bp from the 3' end of the first intron, resulting in a frame shift and premature stop codon. This transcript is predicted to produce a novel 27 amino acid peptide after signal peptidase cleavage. Analysis of brush cytology samples revealed that the percentage of dry eye patients expressing the MUC1/A variant in the conjunctival epithelium is lower than in normal control donors. Western blotting confirmed that MUC1/A is associated with alleles containing the large size class of tandem repeats. Therefore, we propose that one factor in susceptibility to dry eye disease may be the lengths of the MUC1 VNTR in conjunctival epithelium based on the rationale that longer VNTR provide better lubrication and greater protection of the ocular surface against inflammation.
黏蛋白是高度糖基化的蛋白质，对维持包括眼表在内的健康上皮表面至关重要。黏蛋白充当润滑剂、保护剂和信号转导介质。跨膜黏蛋白MUC1上的大多数O-糖基化位点位于一个高度多态的核心区域，该区域包含可变数量的串联重复序列（VNTR）。MUC1等位基因可分为含有少量（30 - 45个）或大量（60 - 90个）重复序列的大小类别。尽管在其他组织中已发现至少12种MUC1的剪接变体，但在人眼表组织中尚未有剪接变体的报道。我们使用逆转录聚合酶链反应（RT-PCR）鉴定MUC1剪接变体，随后通过测序进行确认。我们在此报告，在人角膜、结膜和泪腺的一些样本中存在MUC1/B，其特征为外显子1和2之间的典型剪接，以及MUC1/A，一种从内含子1的3'端保留27bp的转录本，预计会在串联重复区域上游的MUC1序列中添加9个氨基酸。角膜和结膜都含有缺乏跨膜结构域的MUC1/SEC剪接变体，因此会产生可溶性、分泌形式的MUC1。角膜和结膜还含有缺乏串联重复区域的MUC1/Y和MUC1/Z(X)变体。角膜、结膜和泪腺还含有一种先前未描述的MUC1变体转录本，称为MUC1/YI，它从第一个内含子的5'端保留99bp，从3'端保留27bp，导致移码和过早的终止密码子。预计该转录本在信号肽酶切割后会产生一种新的27个氨基酸的肽。对刷取细胞学样本的分析显示，干眼患者结膜上皮中表达MUC1/A变体的百分比低于正常对照供体。蛋白质免疫印迹法证实MUC1/A与含有大量串联重复序列类别的等位基因相关。因此，基于较长的VNTR能提供更好的润滑以及对眼表有更强的抗炎保护作用这一原理，我们提出结膜上皮中MUC1 VNTR的长度可能是干眼疾病易感性的一个因素。
