Functional CD2 mutants unable to bind to, or be stimulated by, LFA-3.
To define epitopes on the CD2 (T11, the T cell erythrocyte receptor) molecule that are necessary for interaction with lymphocyte function-associated Ag-3 (LFA-3), we have expressed the human wild-type CD2 cDNA and mutant CD2 cDNA in a murine Ag-specific T cell hybridoma that responds to human HLA-DR Ag. Here we have expressed mutations at amino acid 91 and 92 of CD2 in the T cell hybridoma. The mutated CD2 molecules were functional in that pairs of anti-CD2 mAb that continued to bind were able to stimulate IL-2 production by the hybridomas. However, CD2 mutants with either the 91 or 92 amino acid substitution had lost the ability to bind to or be activated by either SRBC, which bear an LFA-3 homologue, or by murine L cells expressing human LFA-3. Unlike hybridomas expressing the wild-type CD2 molecule, there was no enhanced response to Ag stimulation. Taken together, these data suggest that the mutated CD2 molecules were no longer able to bind to, or to utilize, LFA-3 for activation. We have previously demonstrated that a mutation at amino acid 51 of CD2 results in loss of binding to LFA-3. Whether these two regions of CD2, discrete and separable by amino acid sequence, form one or more binding sites for LFA-3 remains to be determined.
Division of Pediatric Oncology, Dana-Farber Cancer Institute, Boston, MA 02115.
H L
HL
Wolff
S J
SJ
Burakoff
B E
BE
Bierer
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D015703:Antigens, CD / Q000502:physiology*; D000945:Antigens, Differentiation, T-Lymphocyte / Q000502:physiology*; D000954:Antigens, Surface / Q000502:physiology*; D018801:CD2 Antigens; D018968:CD58 Antigens; D004252:DNA Mutational Analysis; D004912:Erythrocytes / Q000276:immunology; D005434:Flow Cytometry; D006801:Humans; D006825:Hybridomas; D007376:Interleukin-2 / Q000096:biosynthesis; D008213:Lymphocyte Activation; D008562:Membrane Glycoproteins / Q000502:physiology*; D051379:Mice; D011485:Protein Binding; D011971:Receptors, Immunologic / Q000502:physiology*; D012756:Sheep; D013329:Structure-Activity Relationship
D000911:Antibodies, Monoclonal; D015703:Antigens, CD; D000945:Antigens, Differentiation, T-Lymphocyte; D000954:Antigens, Surface; D018801:CD2 Antigens; D018968:CD58 Antigens; D007376:Interleukin-2; D008562:Membrane Glycoproteins; D011971:Receptors, Immunologic