title: Membrane-Associated RING-CH proteins associate with Bap31 and target CD81 and CD44 to lysosomes.
abstract: Membrane-associated RING-CH (MARCH) proteins represent a family of transmembrane ubiquitin ligases modulating intracellular trafficking and turnover of transmembrane protein targets. While homologous proteins encoded by gamma-2 herpesviruses and leporipoxviruses have been studied extensively, limited information is available regarding the physiological targets of cellular MARCH proteins. To identify host cell proteins targeted by the human MARCH-VIII ubiquitin ligase we used stable isotope labeling of amino-acids in cell culture (SILAC) to monitor MARCH-dependent changes in the membrane proteomes of human fibroblasts. Unexpectedly, we observed that MARCH-VIII reduced the surface expression of Bap31, a chaperone that predominantly resides in the endoplasmic reticulum (ER). We demonstrate that Bap31 associates with the transmembrane domains of several MARCH proteins and controls intracellular transport of MARCH proteins. In addition, we observed that MARCH-VIII reduced the surface expression of the hyaluronic acid-receptor CD44 and both MARCH-VIII and MARCH-IV sequestered the tetraspanin CD81 in endo-lysosomal vesicles. Moreover, gene knockdown of MARCH-IV increased surface levels of endogenous CD81 suggesting a constitutive involvement of this family of ubiquitin ligases in the turnover of tetraspanins. Our data thus suggest a role of MARCH-VIII and MARCH-IV in the regulated turnover of CD81 and CD44, two ubiquitously expressed, multifunctional proteins.
authors.affiliation: Department of Molecular Genetics and Microbiology, College of Medicine, University of Florida, Gainesville, Florida, United States of America.
authors.forename: Eric
authors.initials: E
authors.lastname: Bartee
authors.forename: Craig A
authors.initials: CA
authors.lastname: Eyster
authors.forename: Kasinath
authors.initials: K
authors.lastname: Viswanathan
authors.forename: Mandana
authors.initials: M
authors.lastname: Mansouri
authors.forename: Julie G
authors.initials: JG
authors.lastname: Donaldson
authors.forename: Klaus
authors.initials: K
authors.lastname: Früh
mesh_terms: D015703:Antigens, CD / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism; D011992:Endosomes / Q000378:metabolism; D005347:Fibroblasts / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D018960:Hyaluronan Receptors / Q000378:metabolism*; D006820:Hyaluronic Acid / Q000737:chemistry; D008247:Lysosomes / Q000378:metabolism*; D008562:Membrane Glycoproteins / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism*; D008954:Models, Biological; D020543:Proteome; D040901:Proteomics / Q000379:methods; D053719:Tandem Mass Spectrometry / Q000379:methods; D060225:Tetraspanin 28; D060245:Tetraspanin 29
chemical_list: D015703:Antigens, CD; C091247:BCAP31 protein, human; C554798:CD81 protein, human; C554869:CD9 protein, human; D018960:Hyaluronan Receptors; D008562:Membrane Glycoproteins; D008565:Membrane Proteins; D020543:Proteome; D060225:Tetraspanin 28; D060245:Tetraspanin 29; D006820:Hyaluronic Acid