Recently, the generation of Ova- and mCD1-specific T cells, by immunization of mice with naked plasmid DNA encoding B7C1, CD1

Recently, the generation of Ova- and mCD1-specific T cells, by immunization of mice with naked plasmid DNA encoding B7C1, CD1.1, and chicken Ova, was reported (5). was not inhibited by excess Ova or the peptide epitope derived from it. This suggests that, although both lipid and peptide presentation may occur naturally, mCD1 may interact differently with these two types of antigens. CD1 molecules are a family of antigen-presenting molecules distantly related to major histocompatibility complex (MHC)-encoded class I and class II molecules. CD1 molecules have a number of distinct features, including a lack of polymorphism, nearly equal levels of homology with both class I and class II molecules (1), and expression that is impartial of either the peptide transporter associated with antigen presentation (TAP) (2, 3) or the invariant chain (Ii). These distinct features, and the conservation of CD1 Rabbit Polyclonal to IRAK2 molecules through out much of the mammalian order, justify their categorization as a separate, third family of antigen-presenting molecules. Two different T cell subsets reactive to wild-type mouse CD1.1 (mCD1) have been reported. T cells Erythropterin reactive with peptides or proteins that have a hydrophobic mCD1 binding motif have been described by our groups (4, 5). These T cells are TCR +, CD8+ lymphocytes that exhibit cytolytic activity. The other subset of mCD1 specific T lymphocytes, which are either CD4+ or double unfavorable, have been shown to be mCD1 autoreactive (6, 7). More recently, a major populace of these mCD1 autoreactive T cells, namely those that express the NK1.1 molecule and an invariant V14 TCR, have been shown to recognize the glycolipid -galactosylceramide (-GalCer) presented by mCD1 (8, 9). Erythropterin These so-called NK T cells rapidly secrete large amounts of cytokines, and they have been reported to play important immunoregulatory roles in a variety of situations (10, 11). mCD1 molecules are unique in their ability to present chemically well defined peptide and nonpeptide antigens, although the nonclassical H-2M3 molecule also may be capable of this duality of function (12). It has been shown recently, however, that the great majority of mCD1 molecules purified from mammalian cells are bound to glycophosphatidyl inositol containing compounds and that bound peptides could not be detected (13). This raises the questions regarding whether peptide antigen presentation by mCD1 is a phenomenon that occurs naturally and, if so, whether mCD1 molecules are capable of acquiring peptide antigens intracellularly. The experiments described here demonstrate that mCD1 indeed can acquire peptide antigens from processed proteins intracellularly, and, furthermore, they suggest that peptide and nonpeptide antigens may bind to mCD1 differently. MATERIALS AND METHODS Mice, Cell Lines, and Hybridomas. C57BL/6, BALB/c, Erythropterin and C57BL/6 BALB/c F1 (CB6F1) mice were obtained from The Jackson Laboratory or were bred in our animal facility. TAP 1?/? mice were bred from stock provided by Luc van Kaer (Vanderbilt University, Nashville, TN). Mice 8C10 weeks old of both sexes were used. Transfectants expressing wild-type CD1, and transfectants expressing mouse CD1 with the cytoplasmic tail deleted (CD1.1TD), have been described elsewhere (5, 14). Antigens and Antibodies. HPLC-grade chicken Ova was purchased from Sigma. HPLC-purified mCD1 binding peptides, p99 (YEHDwere harvested. For cytokine detection, 1 105 T cells/well in 96-well plates were cultured with 5 104 APC, which were either mCD1+ transfectants, control parental lines, splenocytes, or thymocytes. APC were pulsed for 2C3 hr at 37C with either Ova or peptide p18 and then were washed and irradiated before being added to the cultures. To obtain -GalCer reactive T cells, fresh spleen cells from naive CB6F1 mice were cultured at 2 105 cells/well with 6 104.

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