title: Evidence for functional and dynamic microcompartmentation of Cav-1/TRPV4/K(Ca) in caveolae of endothelial cells.
abstract: Ca(2+)-activated K(+) channels (KCa) play a pivotal role in the endothelium-dependent hyperpolarization and regulation of vascular tone and blood pressure. For activation, KCa depend on an increase of intracellular calcium which is substantially mediated by Ca(2+)-permeable cation channels including the transient receptor potential V4 (TRPV4). It has been proposed that KCa and Ca(2+)-permeable cation channels may be clustered in localized positions within the cell membrane to form functional units and that caveolae may constitute the scaffolding for such microcompartmental organization. Here, we sought to elucidate the composition and functional relevance of these microcompartments in vitro and in vivo. We show that TRPV4 and small-conductance KCa2.3 are enriched in caveolae of human microvascular endothelial cells. Using immunoprecipitation, immunocytology and superresolution microscopy, we found a caveolae-dependent association between caveolin-1, TRPV4 and small conductance KCa2.3, but not intermediate conductance KCa3.1, in endothelial cells under static condition. Mechanical stimulation of cells via exposure to shear stress led to a partial de-novo colocalization of KCa3.1 with Cav-1 and TRPV4. In a mouse model of genetic Cav-1 deficiency, we found significantly reduced KCa-mediated currents as determined by patch-clamping in carotid artery endothelial cells (CAEC) from Cav-1(-/-) mice compared to wildtype. Functionally, Cav-1 deficiency was associated with impaired endothelium-derived hyperpolarizing factor (EDHF)-mediated vasodilation in response to shear stress and acetylcholine. In summary, our findings provide evidence for a dynamic microcompartmentation of TRPV4/KCa in caveolae of endothelial cells and highlight the importance of Cav-1 for endothelial KCa functions and flow-induced vasodilation.
authors.affiliation: Department of Internal Medicine and Nephrology, University Hospital Giessen and Marburg, Philipps-University Marburg, 35033 Marburg, Germany.
authors.forename: Sybelle
authors.initials: S
authors.lastname: Goedicke-Fritz
authors.affiliation: Department of Internal Medicine and Nephrology, University Hospital Giessen and Marburg, Philipps-University Marburg, 35033 Marburg, Germany.
authors.forename: Anuradha
authors.initials: A
authors.lastname: Kaistha
authors.affiliation: Department of Internal Medicine and Nephrology, University Hospital Giessen and Marburg, Philipps-University Marburg, 35033 Marburg, Germany.
authors.forename: Michael
authors.initials: M
authors.lastname: Kacik
authors.affiliation: Department of Internal Medicine and Nephrology, University Hospital Giessen and Marburg, Philipps-University Marburg, 35033 Marburg, Germany.
authors.forename: Sabrina
authors.initials: S
authors.lastname: Markert
authors.affiliation: Department of Internal Medicine and Nephrology, University Hospital Giessen and Marburg, Philipps-University Marburg, 35033 Marburg, Germany.
authors.forename: Andreas
authors.initials: A
authors.lastname: Hofmeister
authors.affiliation: Department of Internal Medicine and Nephrology, University Hospital Giessen and Marburg, Philipps-University Marburg, 35033 Marburg, Germany.
authors.forename: Christoph
authors.initials: C
authors.lastname: Busch
authors.affiliation: Department of Cell Biology and Cell Pathology, Philipps-University Marburg, 35037 Marburg, Germany.
authors.forename: Sebastian
authors.initials: S
authors.lastname: Bänfer
authors.affiliation: Department of Cell Biology and Cell Pathology, Philipps-University Marburg, 35037 Marburg, Germany.
authors.forename: Ralf
authors.initials: R
authors.lastname: Jacob
authors.affiliation: Department of Internal Medicine and Nephrology, University Hospital Giessen and Marburg, Philipps-University Marburg, 35033 Marburg, Germany. Electronic address: grgic@med.uni-marburg.de.
authors.forename: Ivica
authors.initials: I
authors.lastname: Grgic
authors.affiliation: Department of Internal Medicine and Nephrology, University Hospital Giessen and Marburg, Philipps-University Marburg, 35033 Marburg, Germany. Electronic address: hoyer@med.uni-marburg.de.
authors.forename: Joachim
authors.initials: J
authors.lastname: Hoyer
mesh_terms: D000818:Animals; D002118:Calcium / Q000378:metabolism; D021941:Caveolae / Q000502:physiology*; D051242:Caveolin 1 / Q000235:genetics / Q000378:metabolism*; D002451:Cell Compartmentation; D042783:Endothelial Cells / Q000502:physiology*; D006801:Humans; D051660:Intermediate-Conductance Calcium-Activated Potassium Channels / Q000378:metabolism; D051379:Mice; D018345:Mice, Knockout; D018408:Patch-Clamp Techniques; D051640:Small-Conductance Calcium-Activated Potassium Channels / Q000378:metabolism*; D050916:TRPV Cation Channels / Q000378:metabolism*; D014664:Vasodilation / Q000502:physiology
chemical_list: D051242:Caveolin 1; D051660:Intermediate-Conductance Calcium-Activated Potassium Channels; C496923:KCNN4 protein, human; D051640:Small-Conductance Calcium-Activated Potassium Channels; D050916:TRPV Cation Channels; C416986:TRPV4 protein, human; D002118:Calcium
keywords: Ca(2+)-activated K(+) channels; Caveolae; Endothelium-derived hyperpolarizing factor; Microcompartmentation; Transient receptor potential V4; Vasoregulation