Functions of MUC16 in corneal epithelial cells
Functions of MUC16 in corneal epithelial cells
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DOI：
10.1167/iovs.07-0430
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发表时间：
2007-10-01
影响因子：
4.4
通讯作者：
Gipson, Ilene K.
中科院分区：
文献类型：
--
作者：
Blalock, Timothy D.;Spurr-Michaud, Sandra J.;Gipson, Ilene K.
目的。膜相关黏液蛋白MUC16是一种高度o糖基化的跨膜蛋白，由眼表面上皮细胞表达，并定位于微皱襞表面的尖端。尽管MUC16在眼表糖萼中的功能尚不清楚，但人们认为MUC16为上皮膜提供了一个分离屏障。另外两种由眼表上皮表达的膜相关粘蛋白MUC1和MUC4是多功能的，分别通过其细胞质尾部和egf样结构域具有信号传导能力。MUC16细胞质尾部尚未被表征，但由于它含有一个多碱性氨基酸序列，它可能通过ezrin/radixin/moesin （ERM）肌动蛋白结合蛋白与肌动蛋白细胞骨架相互作用。通过细胞质尾肽和ERM下拉实验研究MUC16与肌动蛋白细胞骨架通过ERM的相互作用。利用RNA干扰法在永生化人角膜缘上皮（HCLE）细胞中测定MUC16的功能。采用扫描电镜和免疫电镜观察MUC16敲低对HCLE细胞微褶结构的影响。用玫瑰孟加拉染料培养HCLE细胞，测定MUC16在眼表屏障功能中的作用。测定了荧光标记的金黄色葡萄球菌与HCLE细胞的结合，以确定MUC16在保护病原体粘附在眼表上皮上的作用。MUC16细胞质尾肽与erm的n端结合，MUC1和MUC4肽未检测到结合。MUC16抑制对HCLE细胞表面膜突起无影响；然而，用玫瑰红培养的HCLE细胞显示，在MUC16抑制的细胞中，染料的排斥明显减少。此外，MUC16抑制显著增加了金黄色葡萄球菌与HCLE细胞的结合。这些结果表明MUC16是一个与肌动蛋白细胞骨架相连的多功能分子。MUC16在眼表糖萼中的表达有助于为上皮表面提供一个分离的保护屏障。
PURPOSE. The membrane-associated mucin MUC16, a heavily O-glycosylated transmembrane protein, is expressed by the ocular surface epithelia and localized on the tips of the surface microplicae. Although its functions in the ocular surface glycocalyx are unknown, it is thought that MUC16 provides a disadhesive barrier to the epithelial membrane. Two other membrane-associated mucins expressed by ocular surface epithelia, MUC1 and MUC4, are multifunctional and have signaling capabilities through their cytoplasmic tails and EGF-like domains, respectively. The MUC16 cytoplasmic tail has not been characterized, but, because it contains a polybasic amino acid sequence, it potentially interacts with the actin cytoskeleton through ezrin/radixin/moesin (ERM) actin-binding proteins.METHODS. The interaction of MUC16 with the actin cytoskeleton through ERMs was investigated using cytoplasmic tail peptides and ERM pull-down experiments. MUC16 functions were determined using RNA interference in immortalized human corneal-limbal epithelial (HCLE) cells. The effect of MUC16 knockdown on microplicae structure in HCLE cells was determined using scanning and immunoelectron microscopy. HCLE cells were incubated with rose bengal dye to measure the role of MUC16 in ocular surface barrier function. Binding of fluorescently labeled Staphylococcus aureus to HCLE cells was measured to determine the role of MUC16 in the protection of pathogen adherence on the ocular surface epithelium.RESULTS. MUC16 cytoplasmic tail peptides bound the N-terminus of ERMs, with no detectable binding of MUC1 and MUC4 peptides. No effect on surface membrane projections could be detected in HCLE cells after MUC16 suppression; however, HCLE cells incubated with rose bengal showed that exclusion of the dye was significantly reduced in cells with MUC16 suppression. In addition, S. aureus binding to HCLE cells was significantly increased with MUC16 suppression.CONCLUSIONS. These results suggest that MUC16 is a multifunctional molecule linked to the actin cytoskeleton. The expression of MUC16 in the ocular surface glycocalyx helps provide a disadhesive protective barrier for the epithelial surface.