title: The Transmembrane Domains of β and IX Subunits Mediate the Localization of the Platelet Glycoprotein Ib-IX Complex to the Glycosphingolipid-enriched Membrane Domain.
abstract: We have previously reported that the structural elements of the GP Ib-IX complex required for its localization to glycosphingolipid-enriched membranes (GEMs) reside in the Ibβ and IX subunits. To identify them, we generated a series of cell lines expressing mutant GP Ibβ and GP IX where 1) the cytoplasmic tails (CTs) of either or both GP Ibβ and IX are truncated, and 2) the transmembrane domains (TMDs) of GP Ibβ and GP IX were swapped with the TMD of a non-GEMs associating molecule, human transferrin receptor. Sucrose density fractionation analysis showed that the removal of either or both of the CTs from GP Ibβ and GP IX does not alter GP Ibα-GEMs association when compared with the wild type. In contrast, swapping of the TMDs of either GP Ibβ or GP IX with that of transferrin receptor results in a significant loss (∼ 50%) of GP Ibα from the low density GEMs fractions, with the largest effect seen in the dual TMD-replaced cells (> 80% loss) when compared with the wild type cells (100% of GP Ibα present in the GEMs fractions). Under high shear flow, the TMD-swapped cells adhere poorly to a von Willebrand factor-immobilized surface to a much lesser extent than the previously reported disulfide linkage dysfunctional GP Ibα-expressing cells. Thus, our data demonstrate that the bundle of GP Ibβ and GP IX TMDs instead of their individual CTs is the structural element that mediates the β/IX complex localization to the membrane GEMs, which through the α/β disulfide linkage brings GP Ibα into the GEMs.
authors.affiliation: From the XinHua Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200092, China, the Cardiovascular Research Section, Department of Medicine, Baylor College of Medicine, Houston, Texas 77030.
authors.forename: Guofeng
authors.initials: G
authors.lastname: Xu
authors.affiliation: the Department of Vascular Surgery, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, China, the Cardiovascular Research Section, Department of Medicine, Baylor College of Medicine, Houston, Texas 77030.
authors.forename: Dan
authors.initials: D
authors.lastname: Shang
authors.affiliation: the School of Basic Medicine, Central South University, Changsha 410013, China, the Cardiovascular Research Section, Department of Medicine, Baylor College of Medicine, Houston, Texas 77030.
authors.forename: Zuping
authors.initials: Z
authors.lastname: Zhang
authors.affiliation: the Cardiovascular Research Section, Department of Medicine, Baylor College of Medicine, Houston, Texas 77030.
authors.forename: Tanner S
authors.initials: TS
authors.lastname: Shaw
authors.affiliation: the Cardiovascular Research Section, Department of Medicine, Baylor College of Medicine, Houston, Texas 77030.
authors.forename: Yali
authors.initials: Y
authors.lastname: Ran
authors.affiliation: the Department of Medicine, Puget Sound Blood Center, Division of Hematology, University of Washington, Seattle, Washington 98195, and.
authors.forename: José A
authors.initials: JA
authors.lastname: López
authors.affiliation: the Cardiovascular Research Section, Department of Medicine, Baylor College of Medicine, Houston, Texas 77030 Yuandong@bcm.edu.
authors.forename: Yuandong
authors.initials: Y
authors.lastname: Peng
mesh_terms: D000818:Animals; D001665:Binding Sites / Q000235:genetics; D015153:Blotting, Western; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D006028:Glycosphingolipids / Q000378:metabolism*; D006801:Humans; D021962:Membrane Microdomains / Q000378:metabolism*; D009154:Mutation; D019038:Platelet Glycoprotein GPIb-IX Complex / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D021122:Protein Subunits / Q000235:genetics / Q000378:metabolism; D011990:Receptors, Transferrin / Q000235:genetics / Q000378:metabolism; D013314:Stress, Mechanical; D014841:von Willebrand Factor / Q000378:metabolism
chemical_list: D006028:Glycosphingolipids; D019038:Platelet Glycoprotein GPIb-IX Complex; D021122:Protein Subunits; D011990:Receptors, Transferrin; D014841:von Willebrand Factor
keywords: lipid raft; platelet glycoprotein Ib; protein expression; shear stress; transmembrane domain