title: Kv2.1 potassium channels are retained within dynamic cell surface microdomains that are defined by a perimeter fence.
abstract: Ion channel localization to specific cell surface regions is essential for proper neuronal function. The Kv2.1 K+ channel forms large clusters on the plasma membrane of hippocampal neurons and transfected human embryonic kidney (HEK) cells. Using live cell imaging, we address mechanisms underlying this Kv2.1 clustering in both HEK cells and cultured hippocampal neurons. The Kv2.1-containing surface clusters have properties unlike those expected for a scaffolding protein bound channel. After channel is delivered to the plasma membrane via intracellular transport vesicles, it remains localized at the insertion site. Fluorescence recovery after photobleaching (FRAP) and quantum dot tracking experiments indicate that channel within the surface cluster is mobile (FRAP, tau = 14.1 +/- 1.5 and 11.5 +/- 6.1 s in HEK cells and neurons, respectively). The cluster perimeter is not static, because after fusion of adjacent clusters, green fluorescent protein (GFP)-Kv2.1 completely exchanged between the two domains within 60 s. Treatment of hippocampal neurons expressing GFP-Kv2.1 with 5 microM latrunculin A resulted in a significant increase in average cluster size from 0.89 +/- 0.16 microm2 to 12.15 +/- 1.4 microm2 with a concomitant decrease in cluster number. Additionally, Kv2.1 was no longer restricted to the cell body, suggesting a role for cortical actin in both cluster maintenance and localization. Thus, Kv2.1 surface domains likely trap mobile Kv2.1 channels within a well defined, but fluid, perimeter rather than being tightly bound to a scaffolding protein-containing complex. Channel moves directly into these clusters via trafficking vesicles. Such domains allow for efficient trafficking to the cell surface while sequestering channel with signaling proteins.
authors.affiliation: Department of Biomedical Sciences, Colorado State University, Ft. Collins, Colorado 80523, USA.
authors.forename: Kristen M S
authors.initials: KM
authors.lastname: O'Connell
authors.forename: Annah S
authors.initials: AS
authors.lastname: Rolig
authors.forename: Jennifer D
authors.initials: JD
authors.lastname: Whitesell
authors.forename: Michael M
authors.initials: MM
authors.lastname: Tamkun
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D002478:Cells, Cultured; D006801:Humans; D021962:Membrane Microdomains / Q000737:chemistry* / Q000378:metabolism*; D008969:Molecular Sequence Data; D009474:Neurons / Q000737:chemistry / Q000378:metabolism; D021381:Protein Transport / Q000502:physiology; D051381:Rats; D051669:Shab Potassium Channels / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
chemical_list: D051669:Shab Potassium Channels