title: Membrane raft actin deficiency and altered Ca2+-induced vesiculation in stomatin-deficient overhydrated hereditary stomatocytosis.
abstract: In overhydrated hereditary stomatocytosis (OHSt), the membrane raft-associated stomatin is deficient from the erythrocyte membrane. We have investigated two aspects of raft structure and function in OHSt erythrocytes. First, we have studied the distribution of other membrane and cytoskeletal proteins in rafts by analysis of detergent-resistant membranes (DRMs). In normal erythrocytes, 29% of the actin was DRM-associated, whereas in two unrelated OHSt patients the DRM-associated actin was reduced to <10%. In addition, there was a reduction in the amount of the actin-associated protein tropomodulin in DRMs from these OHSt cells. When stomatin was expressed in Madin-Darby canine kidney cells, actin association with the membrane was increased. Second, we have studied Ca2+-dependent exovesiculation from the erythrocyte membrane. Using atomic force microscopy and proteomics analysis, exovesicles derived from OHSt cells were found to be increased in number and abnormal in size, and contained greatly increased amounts of the raft proteins flotillin-1 and -2 and the calcium binding proteins annexin VII, sorcin and copine 1, while the concentrations of stomatin and annexin V were diminished. Together these observations imply that the stomatin-actin association is important in maintaining the structure and in modulating the function of stomatin-containing membrane rafts in red cells.
authors.affiliation: Institute of Molecular and Cellular Biology, Faculty of Biological Sciences, University of Leeds, Leeds LS2 9JT, UK.
authors.forename: D Katie
authors.initials: DK
authors.lastname: Wilkinson
authors.forename: E Jane
authors.initials: EJ
authors.lastname: Turner
authors.forename: Edward T
authors.initials: ET
authors.lastname: Parkin
authors.forename: Ashley E
authors.initials: AE
authors.lastname: Garner
authors.forename: Penny J
authors.initials: PJ
authors.lastname: Harrison
authors.forename: Mark
authors.initials: M
authors.lastname: Crawford
authors.forename: Gordon W
authors.initials: GW
authors.lastname: Stewart
authors.forename: Nigel M
authors.initials: NM
authors.lastname: Hooper
mesh_terms: D000199:Actins / Q000172:deficiency* / Q000378:metabolism; D000743:Anemia, Hemolytic / Q000378:metabolism*; D000818:Animals; D001798:Blood Proteins / Q000737:chemistry / Q000378:metabolism; D002118:Calcium / Q000494:pharmacology*; D002460:Cell Line; D003598:Cytoskeletal Proteins / Q000378:metabolism; D003902:Detergents / Q000494:pharmacology; D004285:Dogs; D004591:Electrophoresis, Polyacrylamide Gel; D004912:Erythrocytes / Q000187:drug effects / Q000473:pathology / Q000648:ultrastructure; D005089:Exocytosis / Q000187:drug effects*; D006801:Humans; D021962:Membrane Microdomains / Q000187:drug effects / Q000378:metabolism*; D008565:Membrane Proteins / Q000172:deficiency* / Q000378:metabolism; D018625:Microscopy, Atomic Force; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D051348:Tropomodulin / Q000378:metabolism
chemical_list: D000199:Actins; D001798:Blood Proteins; D003598:Cytoskeletal Proteins; D003902:Detergents; D008565:Membrane Proteins; C036860:STOM protein, human; D051348:Tropomodulin; D002118:Calcium