Clonal expansion of freshly isolated CD4T cells by randomized peptides and identification of peptide ligands using combinatorial peptide libraries.
We synthesized Xn (n = 9 -- 19) peptides that consist of 9 to 19 residues with random sequences. X19 is considered to deliver antigenic stimuli to CD4 T cells, because: (a) X19 induces proliferation of peripheral blood mononuclear cells (PBMC), in the presence of IL-2, which is abrogated by monoclonal antibodies to class II HLA; (b) X19 + IL-2 induces proliferation of CD4 T cell clones of distinct specificities; and (c) T cell clones recognizing the same TCR ligands with distinct V beta usage are equally stimulated by X19 + IL-2. We next co-cultured single peripheral CD4 T cells with X19 and mitomycin-treated autologous PBMC. Indeed, single T cells of CD45RA(-) memory phenotype exhibited clonal expansion, with variable rates of proliferation, when IL-4, IL-7, IL-9, IL-15 and agonistic antibody to CD29 were included in the culture. These T cell clones showed heterogeneous proliferation patterns against KGXXXXXXXXXGK-based and KGXXXXXXXXXGKGKK-based combinatorial peptides libraries, in the presence of IL-2. Pattern-match search on a T cell clone resulted in peptide ligand candidates, one of which induced proliferation, as did protein molecules carrying the corresponding sequence. These results indicate that X19 can induce proliferation of peripheral memory T cells, the peptide ligands of which can be determined using combinatorial peptide libraries.
Matsushita
S
S
Division of Immunogenetics, Department of Neuroscience and Immunology, Kumamoto University Graduate School of Medical Sciences, Kumamoto, Japan. imgshom@gpo.kumamoto-u.ac.jp
Tanaka
Y
Y
Matsuoka
T
T
Nakashima
T
T
D000595:Amino Acid Sequence; D000941:Antigens / Q000737:chemistry* / Q000235:genetics / Q000276:immunology*; D015496:CD4-Positive T-Lymphocytes / Q000187:drug effects / Q000276:immunology*; D002455:Cell Division / Q000187:drug effects; D002478:Cells, Cultured; D002999:Clone Cells / Q000187:drug effects / Q000276:immunology; D020650:Combinatorial Chemistry Techniques; D019012:Integrin beta1 / Q000276:immunology; D007378:Interleukins / Q000276:immunology; D007963:Leukocytes, Mononuclear / Q000187:drug effects / Q000276:immunology; D008024:Ligands; D008213:Lymphocyte Activation / Q000187:drug effects; D016685:Mitomycin / Q000494:pharmacology; D008969:Molecular Sequence Data; D019151:Peptide Library; D010455:Peptides / Q000737:chemistry / Q000235:genetics / Q000276:immunology*
D000941:Antigens; D019012:Integrin beta1; D007378:Interleukins; D008024:Ligands; D019151:Peptide Library; D010455:Peptides; D016685:Mitomycin