Oncogenic Raf-1 Disrupts Epithelial Tight Junctions via Downregulation of Occludin

Abstract

Occludin is an integral membrane protein of the epithelial cell tight junction (TJ). Its potential role in coordinating structural and functional events of TJ formation has been suggested recently. Using a rat salivary gland epithelial cell line (Pa-4) as a model system, we have demonstrated that occludin not only is a critical component of functional TJs but also controls the phenotypic changes associated with epithelium oncogenesis. Transfection of an oncogenic Raf-1 into Pa-4 cells resulted in a complete loss of TJ function and the acquisition of a stratified phenotype that lacked cell–cell contact growth control. The expression of occludin and claudin-1 was downregulated, and the distribution patterns of ZO-1 and E-cadherin were altered. Introduction of the human occludin gene into Raf-1–activated Pa-4 cells resulted in reacquisition of a monolayer phenotype and the formation of functionally intact TJs. In addition, the presence of exogenous occludin protein led to a recovery in claudin-1 protein level, relocation of the zonula occludens 1 protein (ZO-1) to the TJ, and redistribution of E-cadherin to the lateral membrane. Furthermore, the expression of occludin inhibited anchorage-independent growth of Raf-1–activated Pa-4 cells in soft agarose. Thus, occludin may act as a pivotal signaling molecule in oncogenic Raf- 1–induced disruption of TJs, and regulates phenotypic changes associated with epithelial cell transformation.

Crk Synergizes with Epidermal Growth Factor for Epithelial Invasion and Morphogenesis and Is Required for the Met Morphogenic Program

“…3B). This finding is consistent with a requirement for MEK1 activity in the loss of ZO-1 from tight junctions and for the maintenance of the dispersed phenotype in epithelial cells expressing activated HaRas or Raf-1 (67,68). This finding also supports previous data indicating a requirement for PI3K activity in the breakdown of adherens junctions (55,57).…”

RasMutation Impairs Epithelial Barrier Function to a Wide Range of Nonelectrolytes

“…The latter study reported leakiness, whereas the former did not, even though both reported evidence of cellular morphology and polarity changes as a result of the transformation. The finding that the downstream effector Raf-1 can also induce tight junction leakiness seems to support the contention by Chen et al (2000) that an activated Ras would in fact result in paracellular leak (Chen et al, 2000;Li and Mrsny, 2000). However, although Chen et al (2000) and Li and Mrsny (2000) reported Ras-and Raf-1-associated tight junction leak, it was not clear that the leak was on a molecular level, rather than a random loss of cells resulting in actual holes in the cell sheet.…”

“…The finding that the downstream effector Raf-1 can also induce tight junction leakiness seems to support the contention by Chen et al (2000) that an activated Ras would in fact result in paracellular leak (Chen et al, 2000;Li and Mrsny, 2000). However, although Chen et al (2000) and Li and Mrsny (2000) reported Ras-and Raf-1-associated tight junction leak, it was not clear that the leak was on a molecular level, rather than a random loss of cells resulting in actual holes in the cell sheet. Either diffusion through tight junctions or leak across multicellular holes in a cell sheet will both permit transepithelial flux of mannitol, PEG, or inulin at increased rates with similar first order kinetics.…”

Zonula Occludens-1, Occludin and E-cadherin Expression and Organization in Salivary Glands with Sjögren’s Syndrome

“…Previous studies using a polarized rat parotid gland epithelial cell line have shown that the functional role of occludin in salivary epithelium is a critical component in TJ organization (Li and Mrsny 2000). Additionally, it may control phenotypic changes associated with epithelium oncogenesis (Li and Mrsny 2000).…”