Occludin confers adhesiveness when expressed in fibroblasts.
Occludin confers adhesiveness when expressed in fibroblasts.
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发表时间：
1997-05
影响因子：
4
通讯作者：
C. M. Itallie;James M. Anderson
中科院分区：
文献类型：
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作者：
C. M. Itallie;James M. Anderson
封闭蛋白是一种与紧密连接特异性相关的膜蛋白。以前的研究表明它可能在形成细胞间密封中起作用。在本研究中，我们表达occludin下的诱导型启动子在occludin空成纤维细胞，以确定这种蛋白质是否赋予细胞间粘附。当人闭合蛋白在NRK和Rat-1成纤维细胞中稳定表达时，其缺乏内源性闭合蛋白和紧密连接，但具有发育良好的含ZO-1的粘附蛋白样连接，闭合蛋白与ZO-1共定位于细胞-细胞接触点。相比之下，缺乏基于钙粘蛋白的粘附连接的L细胞成纤维细胞既不将ZO-1也不将occludin靶向细胞接触位点。接下来使用悬浮细胞测定来定量封闭蛋白诱导的粘附。在NRK和Rat-1细胞中，occludin表达在缺乏钙的情况下诱导粘附，因此不依赖于钙粘蛋白-钙粘蛋白接触。相比之下，L-细胞在该测定中是非粘附性的，并且在诱导闭合蛋白表达后未显示粘附增加。结合抗体的第一个假定的细胞外环的occludin证实，该序列暴露在细胞表面上，和合成肽含有该环的氨基酸序列抑制粘附occludin表达诱导。这些结果表明，occludin的细胞外表面是直接参与细胞-细胞粘附和赋予的能力与其细胞质结合蛋白，ZO-1共定位的能力相关。
Occludin is an integral membrane protein specifically associated with tight junctions. Previous studies suggest it is likely to function in forming the intercellular seal. In the present study, we expressed occludin under an inducible promotor in occludin-null fibroblasts to determine whether this protein confers intercellular adhesion. When human occludin is stably expressed in NRK and Rat-1 fibroblasts, which lack endogenous occludin and tight junctions but do have well developed ZO-1-containing adherens-like junctions, occludin colocalizes with ZO-1 to points of cell-cell contact. In contrast, L-cell fibroblasts which lack cadherin-based adherens junctions, target neither ZO-1 nor occludin to sites of cell contact. Occludin-induced adhesion was next quantified using a suspended cell assay. In NRK and Rat-1 cells, occludin expression induces adhesion in the absence of calcium, thus independent of cadherin-cadherin contacts. In contrast, L-cells are nonadhesive in this assay and show no increase in adhesion after induction of occludin expression. Binding of an antibody to the first of the putative extracellular loops of occludin confirmed that this sequence was exposed on the cell surface, and synthetic peptides containing the amino acid sequence of this loop inhibit adhesion induced by occludin expression. These results suggest that the extracellular surface of occludin is directly involved in cell-cell adhesion and the ability to confer adhesiveness correlates with the ability to colocalize with its cytoplasmic binding protein, ZO-1.