Michlig, Stéphanie Harris, Michael Loffing, Johannes Rossier, Bernard C. Firsov, Dmitri 2005-09-19 Activation of the mitogen-activated protein (MAP) kinase cascade by progesterone in <i>Xenopus</i> oocytes leads to a marked down-regulation of activity of the amiloride-sensitive epithelial sodium channel (ENaC). Here we have studied the signaling pathways involved in progesterone effect on ENaC activity. We demonstrate that: (i) the truncation of the C termini of the αβγENaC subunits results in the loss of the progesterone effect on ENaC; (ii) the effect of progesterone was also suppressed by mutating conserved tyrosine residues in the Pro-<i>X</i>-<i>X</i>-Tyr (PY) motif of the C termini of the βand γ ENaC subunits (β_Y618A and γ_Y628A); (iii) the down-regulation of ENaC activity by progesterone was also suppressed by co-expression ENaC subunits with a catalytically inactive mutant of Nedd4-2, a ubiquitin ligase that has been previously demonstrated to decrease ENaC cell-surface expression via a ubiquitin-dependent internalization/degradation mechanism; (iv) the effect of progesterone was significantly reduced by suppression of consensus sites (β_T613A and γ_T623A) for ENaC phosphorylation by the extracellular-regulated kinase (ERK), a MAP kinase previously shown to facilitate the binding of Nedd4 ubiquitin ligases to ENaC; (v) the quantification of cell-surface-expressed ENaC subunits revealed that progesterone decreases ENaC open probability (whole cell <i>P</i>_o, wc<i>P</i> _o) and not its cell-surface expression. Collectively, these results demonstrate that the binding of active Nedd4-2 to ENaC is a crucial step in the mechanism of ENaC inhibition by progesterone. Upon activation of ERK, the effect of Nedd4-2 on ENaC open probability can become more important than its effect on ENaC cell-surface expression. application/pdf https://folia.unifr.ch/global/documents/299883 https://folia.unifr.ch/documents/299883/files/1_loffing_pdr.pdf eng info:eu-repo/semantics/altIdentifier/doi/10.1074/jbc.M506308200 info:eu-repo/semantics/openAccess License undefined Journal of Biological Chemistry. - 2005, vol. 280, no. 46, p. 38264-38270 info:eu-repo/classification/udc/57 Progesterone down-regulates the open probability of the amiloride-sensitive epithelial sodium channel via a Nedd4-2-dependent mechanism http://purl.org/coar/resource_type/c_6501