Hierarchical regulation of CTLA-4 dimer-based lattice formation and its biological relevance for T cell inactivation.
CTLA-4 is an activation-induced, homodimeric inhibitory receptor in T cells. Recent crystallographic reports have suggested that it may form lattice-like arrays on the cell surface upon binding B7.1/B7.2 (CD80, CD86) molecules. To test the biological relevance of these CTLA-4-B7 lattices, we introduced a C122A point mutation in human CTLA-4, because this residue was shown to be essential for dimerization in solution. Surprisingly, we found that up to 35% of C122A CTLA-4 dimerized in human T lymphocytes. Moreover, C122A CTLA-4 partitioned within lipid rafts, colocalized with the TCR in the immunological synapse, and inhibited T cell activation. C122-independent dimerization of CTLA-4 involved N-glycosylation, because further mutation of the N78 and N110 glycosylation sites abrogated dimerization. Despite being monomeric, the N78A/N110A/C122A triple mutant CTLA-4 localized in the immunological synapse and inhibited T cell activation. Such functionality correlated with B7-induced dimerization of these mutant molecules. Based on these data, we propose a model of hierarchical regulation of CTLA-4 oligomerization by which B7 binding ultimately determines the formation of dimer-dependent CTLA-4 lattices that may be necessary for triggering B7-dependent T cell inactivation.
Darlington
Peter J
PJ
FOCIS Center for Clinical Immunology and Immunotherapeutics, Robarts Research Institute, 100 Perth Drive, London, Ontario, Canada N6A 5K8.
Kirchhof
Mark G
MG
Criado
Gabriel
G
Sondhi
Jitin
J
Madrenas
Joaquín
J
D000409:Alanine / Q000235:genetics; D015703:Antigens, CD / Q000737:chemistry / Q000378:metabolism* / Q000502:physiology; D000943:Antigens, Differentiation / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D018122:B7-1 Antigen / Q000737:chemistry / Q000378:metabolism* / Q000502:physiology; D051940:B7-2 Antigen; D060908:CTLA-4 Antigen; D002461:Cell Line, Transformed; D003545:Cysteine / Q000235:genetics; D019281:Dimerization; D004220:Disulfides / Q000737:chemistry; D006031:Glycosylation; D006801:Humans; D007166:Immunosuppressive Agents / Q000737:chemistry / Q000378:metabolism*; D019169:Jurkat Cells; D008024:Ligands; D008213:Lymphocyte Activation / Q000235:genetics / Q000276:immunology*; D008562:Membrane Glycoproteins / Q000737:chemistry / Q000378:metabolism* / Q000502:physiology; D021962:Membrane Microdomains / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017354:Point Mutation; D011485:Protein Binding / Q000235:genetics / Q000276:immunology; D013601:T-Lymphocytes / Q000737:chemistry / Q000276:immunology* / Q000378:metabolism*
D015703:Antigens, CD; D000943:Antigens, Differentiation; D018122:B7-1 Antigen; D051940:B7-2 Antigen; C497964:CD86 protein, human; D060908:CTLA-4 Antigen; C556706:CTLA4 protein, human; D004220:Disulfides; D007166:Immunosuppressive Agents; D008024:Ligands; D008562:Membrane Glycoproteins; D003545:Cysteine; D000409:Alanine