title: Synergistic activation of ENaC by three membrane-bound channel-activating serine proteases (mCAP1, mCAP2, and mCAP3) and serum- and glucocorticoid-regulated kinase (Sgk1) in Xenopus Oocytes.
abstract: Sodium balance is maintained by the precise regulation of the activity of the epithelial sodium channel (ENaC) in the kidney. We have recently reported an extracellular activation of ENaC-mediated sodium transport (I(Na)) by a GPI-anchored serine protease (mouse channel-activating protein, mCAP1) that was isolated from a cortical collecting duct cell line derived from mouse kidney. In the present study, we have identified two additional membrane-bound serine proteases (mCAP2 and mCAP3) that are expressed in the same cell line. We show that each of these proteases is able to increase I(Na) 6-10-fold in the Xenopus oocyte expression system. I(Na) and the number (N) of channels expressed at the cell surface (measured by binding of a FLAG monoclonal I(125)-radioiodinated antibody) were measured in the same oocyte. Using this assay, we show that mCAP1 increases I(Na) 10-fold (P < 0.001) but N remained unchanged (P = 0.9), indicating that mCAP1 regulates ENaC activity by increasing its average open probability of the whole cell (wcP(o)). The serum- and glucocorticoid-regulated kinase (Sgk1) involved in the aldosterone-dependent signaling cascade enhances I(Na) by 2.5-fold (P < 0.001) and N by 1.6-fold (P < 0.001), indicating a dual effect on N and wcP(o). Compared with Sgk1 alone, coexpression of Sgk1 with mCAP1 leads to a ninefold increase in I(Na) (P < 0.001) and 1.3-fold in N (P < 0.02). Similar results were observed for mCAP2 and mCAP3. The synergism between CAPs and Sgk1 on I(Na) was always more than additive, indicating a true potentiation. The synergistic effect of the two activation pathways allows a large dynamic range for ENaC-mediated sodium regulation crucial for a tight control of sodium homeostasis.
authors.affiliation: Institut de Pharmacologie et de Toxicologie, Université de Lausanne, 1015 Lausanne, Switzerland.
authors.forename: Grégoire
authors.initials: G
authors.lastname: Vuagniaux
authors.forename: Véronique
authors.initials: V
authors.lastname: Vallet
authors.forename: Nicole Fowler
authors.initials: NF
authors.lastname: Jaeger
authors.forename: Edith
authors.initials: E
authors.lastname: Hummler
authors.forename: Bernard C
authors.initials: BC
authors.lastname: Rossier
mesh_terms: D000818:Animals; D002460:Cell Line / Q000737:chemistry; D004347:Drug Interactions; D004357:Drug Synergism; D053503:Epithelial Sodium Channels; D017874:Immediate-Early Proteins; D008565:Membrane Proteins / Q000302:isolation & purification / Q000494:pharmacology / Q000502:physiology*; D051379:Mice; D009687:Nuclear Proteins; D009865:Oocytes; D017346:Protein Serine-Threonine Kinases / Q000494:pharmacology / Q000502:physiology*; D051381:Rats; D012697:Serine Endopeptidases / Q000302:isolation & purification / Q000494:pharmacology / Q000502:physiology*; D015843:Serpins; D015222:Sodium Channels / Q000187:drug effects / Q000378:metabolism*; D014981:Xenopus; D000099184:Serum-Glucocorticoid Regulated Kinases; D000099271:Prostasin
chemical_list: D053503:Epithelial Sodium Channels; D017874:Immediate-Early Proteins; D008565:Membrane Proteins; D009687:Nuclear Proteins; D017346:Protein Serine-Threonine Kinases; D012697:Serine Endopeptidases; D015843:Serpins; D015222:Sodium Channels; D000099184:Serum-Glucocorticoid Regulated Kinases; D000099271:Prostasin; C106610:Serpinb9 protein, mouse; C470745:Tmprss4 protein, mouse