Claudin-7 Modulates Cl- and Na+ Homeostasis and WNK4 Expression in Renal Collecting Duct Cells.

Claudin-7 knockout (CLDN7-/-) mice display renal salt wasting and dehydration phenotypes. To address the role of CLDN7 in kidneys, we established collecting duct (CD) cell lines from CLDN7+/+ and CLDN7-/- mouse kidneys. We found that deletion of CLDN7 increased the transepithelial resistance (TER) and decreased the paracellular permeability for Cl- and Na+ in CLDN7-/- CD cells. Inhibition of transcellular Cl- and Na+ channels has no significant effect on TER or dilution potentials. Current-voltage curves were linear in both CLDN7+/+ and CLDN7-/- CD cells, indicating that the ion flux was through the paracellular pathway. The impairment of Cl- and Na+ permeability phenotype can be rescued by CLDN7 re-expression. We also found that WNK4 (its mutations lead to hypertension) expression, but not WNK1, was significantly increased in CLDN7-/- CD cell lines as well as in primary CLDN7-/- CD cells, suggesting that the expression of WNK4 was modulated by CLDN7. In addition, deletion of CLDN7 upregulated the expression level of the apical epithelial sodium channel (ENaC), indicating a potential cross-talk between paracellular and transcellular transport systems. This study demonstrates that CLDN7 plays an important role in salt balance in renal CD cells and modulating WNK4 and ENaC expression levels that are vital in controlling salt-sensitive hypertension.

Claudin-7 increases chemosensitivity to cisplatin through the upregulation of caspase pathway in human NCI-H522 lung cancer cells.

Claudins are a family of tight junction (TJ) integral membrane proteins that play a crucial role in maintaining cell polarity, adhesion, and paracellular permeability. Changes in expression levels of claudin proteins have been associated with human lung cancer. Previously, we have reported that claudin-7 expression is significantly downregulated in human lung carcinomas. To investigate the role of claudin-7 in lung cancer cells after anti-cancer drug treatments, we transfected claudin-7 cDNA into human NCI-H522 lung cancer cells, which have no detectable expression of claudin-7 protein. Flow cytometry analysis demonstrated that cells transfected with claudin-7 had a significantly higher percentage of cell apoptosis when compared to that of vector transfected cell population. The cell viability assayed by MTT and Annexin V was significantly decreased and cell apoptosis was dramatically increased in claudin-7 transfected cells compared to that of vector transfected cells after cisplatin treatment. Cisplatin is an anti-cancer drug clinically used to treat tumors in several tissues including lung tumors. Most importantly, after cisplatin treatment, the expression levels of cleaved caspase-3, -8, and poly adenosine 5'-diphosphate ribose polymerase (PARP) were much higher in claudin-7 transfected cells than in control cells. Furthermore, using the site-directed mutagenesis approach, we identified that claudin-7 was phosphorylated at serine 204 by protein kinase C. Non-phosphorylated claudin-7 mutant showed increased cell viability, suggesting that phosphorylation increases chemosensitivity to cisplatin treatment. We concluded that claudin-7 expression in H522 lung cancer cells increases chemosensitivity to cisplatin through the increased activation of caspase pathway.

Claudin-7 inhibits human lung cancer cell migration and invasion through ERK/MAPK signaling pathway.

Tight junctions are the most apical component of the junctional complex critical for epithelial cell barrier and polarity functions. Although its disruption is well documented during cancer progression such as epithelial-mesenchymal transition, molecular mechanisms by which tight junction integral membrane protein claudins affect this process remain largely unknown. In this report, we found that claudin-7 was normally expressed in bronchial epithelial cells of human lungs but was either downregulated or disrupted in its distribution pattern in lung cancer. To investigate the function of claudin-7 in lung cancer cells, we transfected claudin-7 cDNA into NCI-H1299, a human lung carcinoma cell line that has no detectable claudin-7 expression. We found that claudin-7 expressing cells showed a reduced response to hepatocyte growth factor (HGF) treatment, were less motile, and formed fewer foot processes than the control cells did. In addition, cells transfected with claudin-7 dramatically decreased their invasive ability after HGF treatment. These effects were mediated through the MAPK signaling pathway since the phosphorylation level of ERK1/2 was significantly lower in claudin-7 transfected cells than in control cells. PD98059, a selective inhibitor of ERK/MAPK pathway, was able to block the motile effect. Claudin-7 formed stable complexes with claudin-1 and -3 and was able to recruit them to the cell-cell junction area in claudin-7 transfected cells. When control and claudin-7 transfected cells were inoculated into nude mice, claudin-7 expressing cells produced smaller tumors than the control cells. Taken together, our study demonstrates that claudin-7 inhibits cell migration and invasion through ERK/MAPK signaling pathway in response to growth factor stimulation in human lung cancer cells.