Evidence for endocytosis of ROMK potassium channel via clathrin-coated vesicles

Wei Zhong Zeng, Victor Babich, Bernardo Ortega, Raymond Quigley, Stanley J. White, Paul A. Welling, Chou Long Huang

Research output: Contribution to journalArticle

63 Scopus citations

Abstract

ROMK channels are present in the cortical collecting ducts of kidney and are responsible for K+ secretion in this nephron segment. Recent studies suggest that endocytosis of ROMK channels is important for regulation of K+ secretion in cortical collecting ducts. We investigated the molecular mechanisms for endocytosis of ROMK channels expressed in Xenopus laevis oocytes and cultured Madin-Darby canine kidney cells. When plasma membrane insertion of newly synthesized channel proteins was blocked by incubation with brefeldin A, ROMK currents decreased with a half-time of ∼6 h. Coexpression with the Lys44→Ala dominant-negative mutant dynamin, but not wild-type dynamin, reduced the rate of reduction of ROMK in the presence of brefeldin A. Mutation of Asn371 to Ile in the putative NPXY internalization motif of ROMK1 abolished the effect of the Lys44→Ala dynamin mutant on endocytosis of the channel. Coimmunoprecipitation study and confocal fluorescent imaging revealed that ROMK channels associated with clathrin coat proteins in Madin-Darby canine kidney cells. These results provide compelling evidence for endocytosis of ROMK channels via clathrin-coated vesicles.

Original languageEnglish (US)
Pages (from-to)F630-F639
JournalAmerican Journal of Physiology - Renal Physiology
Volume283
Issue number4 52-4
StatePublished - Oct 1 2002

Keywords

  • Brefeldin A
  • Dominant-negative dynamin
  • Madin-Darby canine kidney cells
  • Tyrosine-based consensus motif
  • Xenopus laevis oocytes

ASJC Scopus subject areas

  • Physiology
  • Urology

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