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Molecules encoded by a single major histocompatibility complex class I gene can associate with any one of a large number of peptide ligands. T-cell receptors have the capacity to discriminate among these peptide-class I complexes and in many cases bind only a single peptide-class I complex with sufficient affinity to trigger effector function. In contrast, it is generally assumed that class I-specific alloantibodies are indifferent to peptide heterogeneity, being directed toward allele-specific determinants on the molecule. In this report, three monoclonal antibodies were used to precipitate Kb molecules from cell lysates. Surprisingly, in each case a different set of peptides was found to be associated with Kb as detected by peptide-dependent Kb-specific alloreactive cytolytic T lymphocytes or by biochemical resolution. These results demonstrate that the affinity of binding by alloantibodies can be affected by the endogenous peptide ligand.

The role of TCR-gamma delta T lymphocytes in immune responses is currently not well understood. TCR-gamma delta cells have a limited repertoire suggesting that TCR-gamma delta T a limited number of evolutionarily conserved Ag such as nonpolymorphic MHC and heat shock proteins. TCR-gamma delta T lymphocytes appear in enhanced numbers in skin lesions produced by Mycobacterium leprae and in the synovial fluid of joints affected by rheumatoid arthritis, raising the possibility that this subset of T lymphocytes may play a role in control of infectious processes and in autoimmune diseases. We report the identification of a TCR-gamma delta T cell clone isolated from a HSV-infected mouse that recognizes glycoprotein I of HSV type 1. Clone recognition of glycoprotein I does not appear to require the expression of MHC class I or class II gene products. These data suggest that TCR-gamma delta lymphocytes may play an important role in the immune response to viral infections.

We have analyzed the functional significance of the four amino acid differences between the parental H-2Kb and mutant H-2Kbm8 glycoproteins. Six bm8 variants including single substitutions at residues 22, 23, 24, and 30 as well as paired substitutions at residues 23, 30 and 22, 24 were generated and transfected into L cells. Surface expression of these H-2Kb variants was analyzed using monoclonal antibodies which bind to well-defined H-2Kb epitopes. No alterations introduced into the conformational structure of H-2Kb by the amino acid substitutions were detected. The effect of these substitutions on CTL recognition was initially analyzed using the following bulk CTL: either H-2Kb anti-H-2Kbm8, H-2Kbm8 anti-H-2Kb, or third party anti-H-2Kb. The alloreactivity between H-2Kb and H-2Kbm8 is dominated by the amino acid substitution at residue 24 (Glu----Ser). The complete bm8 phenotype, however, also requires the additional substitution at residue 22 (Tyr----Phe). The H-2Kbm8 anti-Kb bulk CTL reacted with both variant H-2Kbm8 molecules containing single substitutions at amino acid positions 22 or 24 but not the variant molecule containing both substitutions. Further analysis using three individual H-2Kbm8 anti-Kb CTL clones indicated the complexity of the self Kbm8 phenotype. Clone 8B1.20 did not react to changes in residues 22 or 24. The 8B1.32 clone reacted with the change at residue 22 but not with the change at residue 24, although the 8B1.54 clone reacted with the change at residue 24 but not with the change at residue 22. The changes in residues 23 (Met----Ile) and/or 30 (Asp----Asn) did not impact significantly on the alloantigenic properties of Kbm8 as determined by both the bulk and cloned CTL populations. According to the three-dimensional class I structure the substitution at amino acid 24 is inaccessible to the TCR. The location of this substitution within the Ag recognition site implies that altered peptide binding, and not a disruption of MHC residues that interact with the TCR, is responsible for the alloreactivity between H-2Kb and H-2Kbm8.

Interleukin-4 (IL-4) has been demonstrated to be an important lymphokine for the generation of cytotoxic T lymphocytes (CTLs). Here we describe an IL-4 inducible gene specifically expressed in CTLs. By sequence homology, this gene is likely to be the mouse homolog of pancreatic lipase. Oocyte translation of in vitro transcribed mRNA results in the expression of a protein with lipase activity, and Northern analysis of various tissues and a large panel of hematopoietic cell types demonstrates that this gene is expressed only in the pancreas and CTLs. Lysates of CTLs grown in IL-4, but not in IL-2, exhibit lipase activity. Furthermore, Northern analysis of CTLs grown in the presence of IL-4 for as little as 5 days demonstrates a marked induction of lipase mRNA, which correlates with enhanced cytolysis by these cells. These results suggest that this lipase may have an important role in CTL effector function.

In vitro PFC responses to the thymus-independent (TI) antigen Streptococcus pneumoniae R36a require T cell replacing factor(s) (TRF). This requirement for TRF is as significant as for the thymus-dependent (TD) antigen SRBC. TRF is shown to be distinct from IL 2 by the following observations: 1) culture supernatants from the cloned T cell line L2, collected over an 8-day period after allogeneic stimulation, transiently contain IL 2 activity but maintain high levels of TRF activity throughout 192 hr; 2) L2V, a variant subclone of L2, produces much higher levels of TRF activity than the parental line but no detectable IL 2 activity; 3) the addition of IL2+, TRF- supernatants from the T cell hybridoma FS6-14.13 does not affect the L2V SF-driven PFC responses to R36a or SRBC; and 4) the addition of contaminating T cells to cultures containing T cell-depleted spleen cells, L2V SF, and antigen does not affect the PFC response. TRF does appear to be indistinguishable from polyclonal B cell stimulating factor (BCSF), which stimulates polyclonal PFC responses in the absence of antigen, mitogen, or anti-Ig. The TRF and BCSF activities of L2V SF could not be separated by ion-exchange, hydrophobic-interaction, and gel-filtration chromatography. TRF and BCSF have an apparent m.w. of approximately 40,000.

Two distinct CD3-associated T cell receptors (TCR alpha beta and TCR gamma delta) are expressed in a mutually exclusive fashion on separate subsets of T lymphocytes. While the specificity of the TCR alpha beta repertoire for major histocompatibility complex (MHC) antigens is well established, the diversity of expressed gamma delta receptors and the ligands they recognize are less well understood. An alloreactive CD3+CD4-CD8- T cell line specific for murine class II MHC (Ia) antigens encoded in the I-E subregion of the H-2 gene complex was identified, and the primary structure of its gamma delta receptor heterodimer was characterized. In contrast to a TCR alpha beta-expressing alloreactive T cell line selected for similar specificity, the TCR gamma delta line displayed broad cross-reactivity for multiple distinct I-E-encoded allogeneic Ia molecules.

The CD8 (Lyt 2) molecule is a phenotypic marker for T lymphocytes that recognize and react with major histocompatibility complex (MHC) class I molecules. Antibody blocking experiments and gene transfection studies indicate that CD8 binds to a determinant on MHC class I molecules on the target cells, facilitating interaction between effector T lymphocytes and the target cell. The CD8 molecule may also be involved in transmembrane signalling during T-cell activation. The existence of CD8- cytotoxic T lymphocytes (CTL) and class I-reactive CTL that are not inhibited by antibody to CD8 suggests that at least some CTL do not require the CD8 molecule to interact with and lyse target cells. We have recently demonstrated that cells transfected with an H-2Dd gene that carries a mutation at residue 227 are not killed by primary CTL8. Here we show that although this mutation abrogates recognition by primary CTL, it does not affect recognition by CD8-independent CTL, suggesting that residue 227 of class I molecules might contribute to a determinant that is the ligand of the CD8 molecule.

(CBA/N x BALB/c)F1 hybrid male mice are unable to mount anti-phosphorylcholine (PC) plaque-forming cell (PFC) responses because they carry the CBA/N X-linked immune defect of B lymphocyte differentiation. Transplantation of splenic B cells from BALB/c mice restores responsiveness to thymus-dependent and thymus-independent PC antigens up to 8 months after cell transfer. Cytotoxicity studies demonstrate the donor origin of PFC generated in reconstituted (CBA/N x BALB/c)F1 mice. Although responsiveness to PC is restored permanently, a shift in idiotype expression that leads to the loss of T 15 idiotypic dominance 3 months after cell transfer can be detected. This shift originates from Ig- cells because Ig+ splenic cells purified in a fluorescence-activated cell sorter maintain T 15 dominance. Therefore, the Ig+ cells have a remarkable capacity to maintain responsiveness to antigens and can perpetuate idiotypic dominance if the stem cell pool is removed.