title: Interaction of ASIC1 and ENaC subunits in human glioma cells and rat astrocytes.
abstract: Glioblastoma multiforme (GBM) is the most common and aggressive of the primary brain tumors. These tumors express multiple members of the epithelial sodium channel (ENaC)/degenerin (Deg) family and are associated with a basally active amiloride-sensitive cation current. We hypothesize that this glioma current is mediated by a hybrid channel composed of a mixture of ENaC and acid-sensing ion channel (ASIC) subunits. To test the hypothesis that ASIC1 interacts with αENaC and γENaC at the cellular level, we have used total internal reflection fluorescence microscopy (TIRFM) in live rat astrocytes transiently cotransfected with cDNAs for ASIC1-DsRed plus αENaC-yellow fluorescent protein (YFP) or ASIC1-DsRed plus γENaC-YFP. TIRFM images show colocalization of ASIC1 with both αENaC and γENaC. Furthermore, using TIRFM in stably transfected D54-MG cells, we also found that ASIC1 and αENaC both localize to a submembrane region following exposure to pH 6.0, similar to the acidic conditions found in the core of a glioblastoma lesion. Using high-resolution clear native gel electrophoresis, we found that ASIC1 forms a complex with ENaC subunits which migrates at ≈480 kDa in D54-MG glioma cells. These data suggest that different ENaC/Deg subunits interact and could combine to form a hybrid channel that likely underlies the amiloride-sensitive current seen in human glioma cells.
authors.affiliation: Dept. of Physiology and Biophysics, University of Alabama at Birmingham, 1918 University Blvd., Birmingham, AL 35294, USA.
authors.forename: Niren
authors.initials: N
authors.lastname: Kapoor
authors.forename: William
authors.initials: W
authors.lastname: Lee
authors.forename: Edlira
authors.initials: E
authors.lastname: Clark
authors.forename: Rafal
authors.initials: R
authors.lastname: Bartoszewski
authors.forename: Carmel M
authors.initials: CM
authors.lastname: McNicholas
authors.forename: Cecelia B
authors.initials: CB
authors.lastname: Latham
authors.forename: Zsuzsanna
authors.initials: Z
authors.lastname: Bebok
authors.forename: Vladimir
authors.initials: V
authors.lastname: Parpura
authors.forename: Catherine M
authors.initials: CM
authors.lastname: Fuller
authors.forename: Cheryl A
authors.initials: CA
authors.lastname: Palmer
authors.forename: Dale J
authors.initials: DJ
authors.lastname: Benos
mesh_terms: D062565:Acid Sensing Ion Channels; D000818:Animals; D001253:Astrocytes / Q000166:cytology / Q000378:metabolism*; D001932:Brain Neoplasms / Q000378:metabolism* / Q000473:pathology; D045744:Cell Line, Tumor; D053503:Epithelial Sodium Channels / Q000235:genetics / Q000378:metabolism*; D005910:Glioma / Q000378:metabolism* / Q000473:pathology; D006801:Humans; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism*; D018408:Patch-Clamp Techniques; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D015222:Sodium Channels / Q000235:genetics / Q000378:metabolism*
chemical_list: C568266:ASIC1 protein, human; D062565:Acid Sensing Ion Channels; C568268:Asic1 protein, rat; D053503:Epithelial Sodium Channels; D008164:Luminescent Proteins; D009419:Nerve Tissue Proteins; D020033:Protein Isoforms; D011993:Recombinant Fusion Proteins; D015222:Sodium Channels