title: Novel role of Rac1/WAVE signaling mechanism in regulation of the epithelial Na+ channel.
abstract: The epithelial Na(+) channel (ENaC) is an essential channel responsible for Na(+) reabsorption in the aldosterone-sensitive distal nephron. Consequently, ENaC is a major effector impacting systemic blood volume and pressure. We have shown recently that Rac1 increases ENaC activity, whereas Cdc42 fails to change channel activity. Here we tested whether Rac1 signaling plays a physiological role in modulating ENaC in native tissue and polarized epithelial cells. We found that Rac1 inhibitor NSC23766 markedly decreased ENaC activity in freshly isolated collecting ducts. Knockdown of Rac1 in native principal cells decreased ENaC-mediated sodium reabsorption and the number of channels at the apical plasma membrane. Members of the Wiskott-Aldrich syndrome protein (WASP) family play a central role in the control of the actin cytoskeleton. N-WASP functions downstream of Cdc42, whereas WAVEs are effectors of Rac1 activity. N-WASP and all 3 isoforms of WAVE significantly increased ENaC activity when coexpressed in Chinese hamster ovary cells. However, wiskostatin, an inhibitor of N-WASP, had no effect on ENaC activity. Immunoblotting demonstrated the presence of WAVE1 and WAVE2 and absence of N-WASP and WAVE3 in mpkCCD(c14) and M-1 principal cells. Immunohistochemistry analysis also revealed localization of WAVE1 and WAVE2 but not N-WASP in the cortical collecting duct of Sprague-Dawley rat kidneys. Moreover, patch clamp analysis revealed that Rac1 and WAVE1/2 are parts of the same signaling pathway with respect to activation of ENaC. Thus, our findings suggest that Rac1 is essential for ENaC activity and regulates the channel via WAVE proteins.
authors.affiliation: Department of Physiology, Medical College of Wisconsin, 8701 Watertown Plank Rd, Milwaukee, WI 53226, USA.
authors.forename: Alexey V
authors.initials: AV
authors.lastname: Karpushev
authors.forename: Vladislav
authors.initials: V
authors.lastname: Levchenko
authors.forename: Daria V
authors.initials: DV
authors.lastname: Ilatovskaya
authors.forename: Tengis S
authors.initials: TS
authors.lastname: Pavlov
authors.forename: Alexander
authors.initials: A
authors.lastname: Staruschenko
mesh_terms: D000634:Aminoquinolines / Q000494:pharmacology; D000704:Analysis of Variance; D000818:Animals; D015153:Blotting, Western; D016466:CHO Cells; D002478:Cells, Cultured; D006224:Cricetinae; D003412:Cricetulus; D004594:Electrophysiology; D053503:Epithelial Sodium Channels / Q000378:metabolism*; D007150:Immunohistochemistry; D007685:Kidney Tubules, Collecting / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D011743:Pyrimidines / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley; D015398:Signal Transduction / Q000502:physiology*; D012964:Sodium / Q000378:metabolism*; D051316:Wiskott-Aldrich Syndrome Protein / Q000378:metabolism; D051300:Wiskott-Aldrich Syndrome Protein Family / Q000378:metabolism*; D020830:rac1 GTP-Binding Protein / Q000037:antagonists & inhibitors / Q000378:metabolism*
chemical_list: D000634:Aminoquinolines; D053503:Epithelial Sodium Channels; C487513:NSC 23766; D011743:Pyrimidines; D051316:Wiskott-Aldrich Syndrome Protein; D051300:Wiskott-Aldrich Syndrome Protein Family; D012964:Sodium; D020830:rac1 GTP-Binding Protein