Claudin-2 is expressed in the tight junctions of leaky epithelia, where it forms cation-selective and water permeable paracellular channels. Its abundance is under fine control by a complex signaling network that affects both its synthesis and turnover in response to various environmental inputs. Claudin-2 expression is dysregulated in many pathologies including cancer, inflammation, and fibrosis. Claudin-2 has a key role in energy-efficient ion and water transport in the proximal tubules of the kidneys and in the gut. Importantly, strong evidence now also supports a role for this protein as a modulator of vital cellular events relevant to diseases. Signaling pathways that are overactivated in diseases can alter claudin-2 expression, and a good correlation exists between disease stage and claudin-2 abundance. Further, loss- and gain-of-function studies showed that primary changes in claudin-2 expression impact vital cellular processes such as proliferation, migration, and cell fate determination. These effects appear to be mediated by alterations in key signaling pathways. The specific mechanisms linking claudin-2 to these changes remain poorly understood, but adapters binding to the intracellular portion of claudin-2 may play a key role. Thus, dysregulation of claudin-2 may contribute to the generation, maintenance, and/or progression of diseases through both permeability-dependent and -independent mechanisms. The aim of this review is to provide an overview of the properties, regulation, and functions of claudin-2, with a special emphasis on its signal-modulating effects and possible role in diseases.

The tight junctional pore-forming protein claudin-2 (CLDN-2) mediates paracellular Na ؉ and water transport in leaky epithelia and alters cancer cell proliferation. Previously, we reported that tumor necrosis factor-␣ time-dependently alters CLDN-2 expression in tubular epithelial cells. Here, we found a similar expression pattern in a mouse kidney injury model (unilateral ureteral obstruction), consisting of an initial increase followed by a drop in CLDN-2 protein expression. CLDN-2 silencing in LLC-PK 1 tubular cells induced activation and phosphorylation of guanine nucleotide exchange factor H1 (GEF-H1), leading to Ras homolog family member A (RHOA) activation. Silencing of other claudins had no such effects, and re-expression of an siRNA-resistant CLDN-2 prevented RHOA activation, indicating specific effects of CLDN-2 on RHOA. Moreover, kidneys from CLDN-2 knockout mice had elevated levels of active RHOA. Of note, CLDN-2 silencing reduced LLC-PK1 cell proliferation and elevated expression of cyclin-dependent kinase inhibitor P27 (P27KIP1) in a GEF-H1/RHOA-dependent manner. P27KIP1 silencing abrogated the effects of CLDN-2 depletion on proliferation. CLDN-2 loss also activated myocardin-related transcription factor (MRTF), a fibrogenic RHOA effector, and elevated expression of connective tissue growth factor and smooth muscle actin. Finally, CLDN-2 downregulation contributed to RHOA activation and smooth muscle actin expression induced by prolonged tumor necrosis factor-␣ treatment, because they were mitigated by re-expression of CLDN-2. Our results indicate that CLDN-2 suppresses GEF-H1/RHOA. CLDN-2 down-regulation, for example, by inflammation, can reduce proliferation and promote MRTF activation through RHOA. These findings suggest that the initial CLDN-2 elevation might aid epithelial regeneration, and CLDN-2 loss could contribute to fibrotic reprogramming.

Claudin-2 (Cldn-2) is a channel-forming tight junction (TJ) protein in the proximal tubules that mediates paracellular Na transport and has also emerged as a regulator of proliferation and migration. Expression of Cldn-2 is altered by numerous stimuli, but the underlying mechanisms remain incompletely understood. Here we show that Cldn-2 protein and mRNA expression were low in subconfluent tubular cells and increased during junction maturation. Cldn-1 or occludin did not exhibit similar confluence-dependence. Conversely, disruption of TJs by Ca removal or silencing of zonula occludens-1 (ZO-1) or ZO-2 induced a large drop in Cldn-2 abundance. Immunofluorescent staining revealed a more uneven Cldn-2 staining in nascent, Cldn-1-positive TJs. Subconfluence and ZO-1 silencing augmented Cldn-2 degradation and reduced Cldn-2 promoter activity, suggesting that insertion into the TJs slows Cldn-2 turnover. Indeed, blocking endocytosis or lysosomal degradation increased Cldn-2 abundance. Cell confluence increased expression of the junctional adapters ZO-1 and -2, and the small GTPase Rac, and elevated Rac activity and p21-activated kinase (Pak) phosphorylation, suggesting that they might mediate confluence-dependent Cldn-2 regulation. Indeed, Rac silencing or Pak inhibition strongly reduced Cldn-2 protein abundance, which was likely the combined effect on turnover, as these interventions reduced Cldn-2 promoter activity and augmented Cldn-2 degradation. Taken together, our data suggest that TJ integrity and maturity, ZO-1 expression/TJ localization, and Rac/Pak control Cldn-2 degradation and synthesis. A feedback mechanism connecting Cldn-2 expression with junction remodeling, e.g., during wound healing, epithelial-mesenchymal transition, or tumor metastasis formation, may have important downstream effects on permeability, proliferation, and migration.