Activation of the calcium sensing receptor increases claudin-14 expression via a PLC -p38-Sp1 pathway.

Activation of the basolateral calcium sensing receptor (CaSR) in the renal tubular thick ascending limb (TAL) increases claudin-14 expression, which reduces paracellular calcium (Ca2+ ) permeability, thus increasing urinary Ca2+ excretion. However, the upstream signaling pathway contributing to altered CLDN14 gene expression is unknown. To delineate this pathway, we identified and then cloned the CaSR responsive region including the promoter of mouse Cldn14 into a luciferase reporter vector. This 1500 bp sequence upstream of the 5' UTR of Cldn14 variant 1, conferred increased reporter activity in the presence of high extracellular Ca2+ (5 mM) relative to a lower (0.5 mM) concentration. Assessment of Cldn14 reporter activity in response to increased extracellular Ca2+ in the presence or absence of specific inhibitors confirmed signaling through PLC and p38, but not JNK. Overexpression of SP1 attenuated Cldn14 reporter activity in response to CasR signaling. SP1 is expressed in the TAL and phosphorylation was attenuated by CaSR signaling. Finally, activating mutations in the CaSR increased Cldn14 reporter activity while a dominant negative mutation in the CaSR inhibited it. Together, these studies suggest that basolateral activation of the CASR leads to increased Cldn14 expression via a PLC- stimulated p38 pathway that prevents Sp1 mediated repression.