Characterization of CTLA-4 structure and expression on human T cells.
CTLA-4 is an adhesion receptor expressed on activated T cells. The amino acid sequence of CTLA-4 is related to CD28, and although the function of CTLA-4 remains unknown, it shares several features with CD28, including a common counter-receptor, B7, that is present on Ag-presenting cells. In a recent study we found that CD28 and CTLA-4 were coexpressed at the mRNA level on activated T cells but that only CD28 was expressed on resting T cells. Here we show that within the T cell population, CTLA-4 expression is restricted to the subset of T cells that also express cell surface CD28. CTLA-4 mRNA expression can be induced on quiescent T cells via phorbol ester-mediated activation of protein kinase C but not with calcium ionophore treatment alone. Phorbol ester-induced expression of CTLA-4 mRNA could be enhanced with calcium ionophore treatment, and treatment of cells in this manner resulted in a reciprocal decrease in expression of CD28 mRNA. Ligation of CD28 with monoclonal antibody also resulted in the specific and rapid induction of CTLA-4 mRNA. To study the expression of CTLA-4 at the protein level, a rabbit antiserum against a recombinant protein derived from CTLA-4 cDNA was generated. When activated T cells were labeled with [35S]methionine, the rabbit antiserum precipitated a 41- to 43-kDa protein from whole cell lysates. Similar results were found when detergent-soluble lysates from 125I surface-labeled resting and activated T cells were analyzed by SDS-PAGE. Surprisingly, under the conditions tested, CTLA-4 migrated primarily as a monomer at the cell surface, and could not be shown to exist as a disulfide-bonded homodimer or as a heterodimer consisting of CTLA-4 and CD28. These results suggest that B7 can bind to T cells via distinct receptor complexes consisting of either CD28 or CTLA-4, and that these complexes may potentially mediate distinct biologic functions. Further, the present results suggest that noncovalent interactions might mediate association of CTLA-4 and/or CD28 at the cell surface.
Department of Pathology, University of Michigan, Ann Arbor 48109.
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T
Lindsten
K P
KP
Lee
E S
ES
Harris
B
B
Petryniak
N
N
Craighead
P J
PJ
Reynolds
D B
DB
Lombard
G J
GJ
Freeman
L M
LM
Nadler
G S
GS
Gray
D000069594:Abatacept; D000818:Animals; D015703:Antigens, CD / Q000032:analysis; D000943:Antigens, Differentiation / Q000032:analysis / Q000737:chemistry / Q000235:genetics*; D000945:Antigens, Differentiation, T-Lymphocyte / Q000032:analysis; D001483:Base Sequence; D018106:CD28 Antigens; D060908:CTLA-4 Antigen; D002460:Cell Line; D015870:Gene Expression; D006801:Humans; D007106:Immune Sera / Q000276:immunology; D018796:Immunoconjugates; D008969:Molecular Sequence Data; D012333:RNA, Messenger / Q000032:analysis; D011817:Rabbits; D013601:T-Lymphocytes / Q000378:metabolism*
D015703:Antigens, CD; D000943:Antigens, Differentiation; D000945:Antigens, Differentiation, T-Lymphocyte; D018106:CD28 Antigens; D060908:CTLA-4 Antigen; C556706:CTLA4 protein, human; D007106:Immune Sera; D018796:Immunoconjugates; D012333:RNA, Messenger; D000069594:Abatacept