In situ chemical cross-linking on living cells reveals CD9P-1 cis-oligomer at cell surface.
Tetraspanins are integral membrane proteins involved in a variety of physiological and pathological processes. They associate with each other in multimolecular complexes containing numerous membrane proteins. As a first step towards the study of the supramolecular organization of tetraspanin complexes, we have implemented a proteomic approach based on in situ protein cross-linking on living cells followed by affinity purification of tetraspanin complexes. This allowed observing the presence of high molecular weight protein complexes that were characterized as containing CD9P-1/CD315 using LC-MS/MS. Western blot analyses and the use of different tags demonstrated the presence of CD9P-1 oligomer in cis-association at cell surface. A significant amount of CD9P-1 oligomer was observed on various cell types. We have shown that CD9P-1 self-associates independently from its association with tetraspanins. However, the expression level of CD9 or CD81 that associate directly and specifically with CD9P-1, positively modulates the cross-linking efficiency of CD9P-1. Thus, tetraspanins can play a role on CD9P-1 oligomerization status.
André
Magali
M
Inserm, U602, F-94807 Villejuif, France.
Chambrion
Célia
C
Charrin
Stéphanie
S
Soave
Sabrina
S
Chaker
Joëlle
J
Boucheix
Claude
C
Rubinstein
Eric
E
Le Naour
François
F
D000595:Amino Acid Sequence; D015703:Antigens, CD / Q000378:metabolism; D000954:Antigens, Surface / Q000737:chemistry / Q000378:metabolism; D002462:Cell Membrane / Q000187:drug effects* / Q000378:metabolism; D002478:Cells, Cultured / Q000187:drug effects*; D003432:Cross-Linking Reagents / Q000494:pharmacology*; D006801:Humans; D007536:Isomerism; D020014:K562 Cells; D008562:Membrane Glycoproteins / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism; D008954:Models, Biological; D046912:Multiprotein Complexes / Q000378:metabolism; D009363:Neoplasm Proteins / Q000737:chemistry / Q000378:metabolism*; D011485:Protein Binding; D025941:Protein Interaction Mapping; D055503:Protein Multimerization / Q000187:drug effects; D060225:Tetraspanin 28; D060245:Tetraspanin 29; D059470:Tetraspanins
D015703:Antigens, CD; D000954:Antigens, Surface; C554798:CD81 protein, human; C554869:CD9 protein, human; D003432:Cross-Linking Reagents; D008562:Membrane Glycoproteins; D008565:Membrane Proteins; D046912:Multiprotein Complexes; D009363:Neoplasm Proteins; C486331:PTGFRN protein, human; C508125:TSPAN1 protein, human; D060225:Tetraspanin 28; D060245:Tetraspanin 29; D059470:Tetraspanins