Diversity of alpha and beta subunits of T-cell receptors specific for the H4 minor histocompatibility antigen

 The mouse H4 minor histocompatibility antigen (HA) includes a single H2K^b-bound peptide that stimulates rejection of skin allografts and generation of cytolytic T lymphocytes (CTL). We evaluated the diversity of the CTL response to this single minor HA peptide by sequencing alpha and beta chains of T-cell receptors (Tcr) from H4-specific CTLs as a first step toward understanding the diversity of Tcrs specific for single minor HA. We selected H4-specific CTL clones from short-term lines that were restricted by K^b (19 clones), K^bm5 (7 clones), and K^bm11 (10 clones). Whereas multiple V_α and V_β family members were identified in the panel of K^b-restricted CTLs, five VB genes and one VA subfamily were predominant in CTLs derived from multiple individuals. Similar distributions were observed with K^bm5- and K^bm11-restricted CTLs, suggesting that these two mutants did not alter Tcr gene usage observable at the VA and/or VB gene levels. Negatively charged residues were present in the CDR3 regions of 12/13 unique K^b-restricted beta chains. A comparable observation was made with K^bm5-restricted CTLs, and the distributions of these residues among CDR3 positions were similar in the two CTL panels. However, the K^bm11 mutation dramatically altered the distribution of these residues resulting in their presence in positions 10–12 of CDR3 regions in all CTLs. These results indicate that the Tcrs expressed by CTLs specific for this single minor HA peptide are oligoclonal and characterized by the presence of negatively charged residues in beta CDR3 regions, the distribution of which is profoundly altered by the K^bm11 mutation.     

Cross-reactivity patterns of vaccinia-specific cytotoxic T lymphocytes from H-2K b mutants

Limit-dilution cultures were used to select vaccinia-immune T-cell populations from bml and bm3 mutant mice that were not lytic for virus-infected targets expressing the K^b and D^b MHC glycoprotein. Approximately 30% of virus-immune CTL were restricted in each case to K^bm1 and K^bm3, rather than to D^b. Evidence of extensive cross-reactivity was found for these virus-immune CTL. Bm3 and bmll mice sharing one amino acid mutation from wild-type but differing by a second mutation seen only in bm3 are the most cross-reactive pair in their presentation of vaccinia. The bm1 and bm10 pair with dissimilar mutations from wild-type affecting the same CNBr fragment are also largely cross-reactive. However, 30% cross-reactivity is also found for bm1 and bm3, which differ in separate CNBr fragments. That mutants expressing amino acid substitutions in the same region of the peptide tend to show more evidence of cross-reactivity does not necessarily mean the T cells see linear arrays of amino acids on the MHC glycoprotein. For instance, K^bm1 and K^bm10 differ for three amino acids, but bm1 T cells are highly lytic for bm10 virus-infected targets. However, there is no cross-reactivity for K^bm1 and K^b, which differ at only two amino acids. The key to further understanding may rest with defining the nature of the conformational differences among the K^bm1, K^bm10, and K^b glycoproteins.     

Adoptive transfer of delayed-type hypersensitivity to sendai virus

TheH-2 restriction pattern of cytolytic T lymphocytes (Tc) and T lymphocytes which mediate a delayed-type hypersensitivity response (Td) directed against infectious Sendai virus was investigated usingH-2 mutant mice. Td and Tc lymphocytes exhibit the same fine specificity for self-recognition, for example, B6.C-H- 2^bm1 effector T cells were unable to recognize viral antigens in association with wild-type K^b and vice versa, B6.-H- 2^bm6 effector cells did not mediate a reaction against virus plus wild-type K^b but, on the other hand, T cells of wild-type K^b recognized virus plus K^bm6.BALB/c-H- 2^dm2 T cells lacked reactivity against virus in association with wild-type D^d, but again wild-type D^d effector cells recognized virus plus D^dm2.Abbreviations used in this paper DTH delayed-type hypersensitivity - EID₅₀ mean egg infective dose - FCS fetal calf serum - HAU hemagglutinating units - LPS lipopolysaccharide - Ly^(–) low amount of Ly antigens - MHC major histocompatibility complex - 2-ME 2-mercaptoethanol - PBS phosphate-buffered saline - Tc cytolytic T cell - Td T cell which mediates a delayed-type hypersensitivity reaction     

Effect of anH-2 mutation on cytotoxic T cell recognition. Specific antigen can determine the pattern ofH-2 restriction

Hz1 (H-2 ^bm1 ) mice, an H-2 mutant strain derived from C57BL/6(H-2 ^b ), were either injected with vaccinia virus or had their spleen cells sensitized in vitro with syngeneic TNP-modified cells. The cytotoxic cells generated were tested for their activity against target cells that were either infected with vaccinia virus, TNP-modified, or both vaccinia infected and TNP-modified.Hz1 anti-TNP cytotoxic cells specifically lysed syngeneic target cells that were trinitrophenylated but not infected with vaccinia virus, while anti-vaccinia cells specifically lysed vaccinia infected target cells but not TNP-cells. Hz1 (H-2K ^bm1 D ^b ) anti-TNP effector cells killed B10.A(5R)-TNP (H-2K ^b D ^d ) targets, indicating that there is cross-reactivity between TNP-H-2K^b and TNP-H-2K^bm1. On the other hand, there is no cross-reactivity between vaccinia-H-2K^b and H-2K^bm1, since Hz1 anti-vaccinia effector cells did not kill vaccinia infected B10.A(5R) targets.Since Hz1 anti-TNP effector cells lysed B10.A(5R) target cells that were first infected with vaccinia virus and then derivatized with TNP, virus does not mask cross-reactive determinants shared by TNP-H-2K^b and H-2K^bm1. Additional experiments showed that Hz1 anti-TNP effector cells lysed TNP-modified and vaccinia infected B10.A(5R) target cells irrespective of the virus concentration used for infection or the time of addition of virus. Further, there are no detectable quantitative differences between C57BL/6 and Hz1 anti-TNP effector cells in their ability to kill TNP-5R targets.The cytotoxic effect of Hz1 anti-TNP effector cells on B10.A(5R)-TNP targets could not be blocked with TNP derivatized inhibitor cells that carry theH-2D ^d region allele. Thus, the ability of anti-TNP H-2K^b effector cells to cross-react with H-2K^bm1 cannot be explained by a cross-reaction between H-2K^bm1 and H-2D^d.Abbreviations used in this paper TNP trinitrophenol - PFU plaque forming unit - Con A Concanavalin A - BSS balanced-salt-solution - FCS fetal calf serum - TNBS trinitrobenzene sulfonic acid - PBS phosphate-buffered-saline     

Neonatal tolerance induction acrossH-2 mutational disparity: Induction and specificity of tolerance

Mutational disparities derived from alleles of theH-2K andH-2D loci vary widely in their ability to induce neonatal tolerance. The more subtle mutations, such asK ^bm5 andK ^bm8, proved to be excellent tolerogens, but theK ^bm3 mutant (M505) turned out to be the poorest tolerogen yet studied of all H-2 alloantigens. By challenging tolerant animals with skin grafts from related mutants, it was found that expression of tolerance was highly specific. Although a minority of tolerant animals failed to discriminate between the K^b, K^bm5 and K^bm8 antigens, they never failed to discern K^b, K^bml and K^bm3 as distinctly different alloantigens.     

Biochemical studies on the H-2K antigens of the MHC mutantsbm3 andbm11

Biochemical analyses of the H-2K-gene products of the C57BL/6 mutant strains, bm3 and bm11, have been carried out in order to characterize the structural relationships among these antigens. From comparative tryptic peptide mapping of cyanogen bromide fragments from the mutant and parent K^b glycoproteins and from preliminary amino-acid sequence analyses, a number of discrete differences have been discerned. Two sites of difference at amino residues 77 and 89 in the K^bm3 glycoprotein are noted relative to K^b. An alteration at residue 77 similar or identical to that seen in K^bm3 is present in the K antigen of thebm11 mutant. Because our techniques sample only 75 to 80 percent of the extracellular portion of H-2K^b, other undetected changes are possible. However, our present findings are most consistent with the conclusion that only very limited differences exist between mutant and parent molecules. Further, taken together with CML (cell-mediated-lymphocytotoxicity) reaction patterns (Melief et al. 1980), the biochemical data support the hypothesis that the proposed structural alterations in theK ^b mutant glycoproteins are directly related to their observed immunological specificity.     

Variable spread of X inactivation affecting the expression of different epitopes of the Hya gene product in mouse B-cell clones

Cloned B-cell lines from a female T16H/XSxr mouse in which Tdy expression was suppressed due to X inactivation and from a male X/XSxr mouse, both of the (kxb)F₁ haplotype, were examined for H-Y expression. This was determined both by their ability to act as targets for H-2^k and H-2^b-restricted H-Y-specific cytotoxic T cells and by their ability to stimulate the proliferation of H-2K^k, H-2D^b (class I) and A^b (class II)-restricted T-cell clones. In B-cell clones from the T16H/XSxr mouse, expression of H-Y/D^b exhibited partial X inactivation and only a proportion ( 30%) of the cells were targets for or stimulated H-2D^b-restricted H-Y-specific T cells. In contrast, H-Y eiptopes restricted by H-2^k (H-Y/K^k, H-Y/D^k) and A^b (H-Y/A^b) exhibited no X inactivation. Furthermore, no inactivation of H-Y/D^b, H-Y/A^b, or H-Y^k was observed in the male X/XSxr mouse. These results indicate that the T16H/XSxr female is a mosaic, as a result of the variable spread of X inactivation into the Sxr region. They further suggest that the H-Y antigen recognized in association with H-2^k and H-2D^b class I molecules and A^b class II molecules may be the product of more than one gene.     

Studies ofH-2K b mutant mice

Three newH-2 ^b mutant strains, B6.C-H-2 ^bm9 , B6.C-H-2 ^bm10 and B6.C-H–2 ^bm11 , are described. The three mutant strains are of the gain and loss type as they reject skin grafts reciprocally with the parental C57BL/6Kh. The mutations, which arose independently, are all allelic at the same locus as 11 other mutant strains already described. By complementation and other studies the mutated gene has been shown to beH-2K ^b . The strains were typed directly and by absorption with antisera specific for H-2K^b and H-2D^b private and public specificities and for Ia^b specificities. Each strain typed differently with these sera. The strain B6.C-H-2 ^bm9 was found to be serologically identical with C57BL/6. The strains B6.C-H-2 ^bm10 and B6.C-H-2 ^bm11 were found to have alterations in the private H-2K^b specificity, H-2.33, and in the public specificity, H-2.5, but to a different extent. B6.C-H- 2^bm10 had a marked decrease in the amount of H-2.33 expressed on the splenic cell surface as compared to C57BL/6 and also has a marked decrease in the expression of H-2.5 on both spleen and red blood cells. In comparison, B6.C-H-2 ^bm11 has a decrease in the expression of H-2.33 but an increase in the expression of H-2.5 on both splenic and red blood cells. The other H-2^b specificities appeared to be unaltered as compared with C57BL/6.     

Biochemical studies on the H-2K mutant B6.C-H-2 bm10

The H-2K glycoprotein from the MHC mutant bm10 was analyzed biochemically to determine where primary structural differences distinguished it from the parental standard molecule, K^b. Comparative peptide maps showed differences in two peptides known to be part of the parental CNBr fragment spanning amino acids 139 to 228. Partial sequence analyses of CNBr fragments and tryptic peptides identified two tightly clustered amino acid substitutions at amino acids 165 (Val to Met) and 173 (Lys to unknown). The substitutions in bm10 represent the most carboxy-terminal substitutions characterized in the K^b molecules of the spontaneous, histogenically active H-2 mutants.     

H-2 class I loci determine sensitivity to MCMV in macrophages and fibroblasts

Peritoneal (PM) and bone marrow-derived (BMM) macrophages and lung fibroblasts (LF) from inbred, intra-H-2 recombinant, H-2 mutant, and hybrid mice were infected with murine cytomegalovirus (MCMV) under centrifugal enhancement. At the concentration of virus employed, peritoneal macrophages from strains carrying Kd, Kb, D^d, KS and/or D^s, K4 and/or D4 alleles could be infected to a level of 80%–100%, as assessed by viral antigen expression or loss of Fc receptors. Cells lacking these haplotypes and carrying K^k, K^f, D^k, Df, or Db were resistant, yielding levels of infection below 20% . The background (non-H-2) and class II genotype and the S allele did not influence the proportions of cells infected. Furthermore, sensitivity was dominant in the F, progeny of H-2 b x H-2 k and H-2d x H-2 k crosses, and was not compromised by thebm1, bm3, bm10, or bm14 mutations in the al or2 regions of Kb orD ^b. The proportions of cells able to release infectious virus were low, but paralleled the frequencies of viral antigen expression. The class I genotype also determined susceptibility to MCMV infection in BMM and LF, although up to 35% of H-2 k BMM and 46% of H-2 k LF could be infected. The findings are consistent with an association between K and D antigens and a cellular receptor for MCMV on all three cell types.     

Studies of twoH-2D b mutants: B6.C-H-2 bm13 and B6.C-H-2 bm14

Two new C57BL/6H-2 mutants,B6.C-H- 2^bm13 and B6.C-H- 2^bm14 are described. They arose independently in C57BL/6 as spontaneous mutations of the gain and loss type. Complementation studies map the mutations in both bm13 and bm14 to theH-2D ^b gene. How ever, these two mutant strains are not identical, but occurred as independent mutations at the same locus, as shown by reciprocal graft rejection and by the inability of the (bm13 × bm114)F₁ hybrid to accept C57BL/6 grafts. Serological studies by direct testing (cytotoxicity and hemagglutination) and by quantitative absorption demonstrated a decrease in the H-2D^b private specificity H-2.2 in both bm13 and bm14 when compared to C57BL/6. This was confirmed by SDS-PAGE analysis using antisera detecting the H-2.2 specificity. Attempts to produce antibodies to either the gained or lost specificities of the two mutant strains failed.     

Class I MHC molecules rather than other mouse genes dictate influenza epitope recognition by cytotoxic T cells

Influenza nucleoprotein (NP) is an important target antigen for influenza A virus cross-reactive cytotoxic T cells (Tc). Here we examine the NP epitope recognized by cloned and polyclonal BALB/c Tc and the genetics of this recognition pattern. We can define NP residues 147–161 as the epitope seen in conjunction with K ^d , the only H-2^d class I responder allele for NP restriction. H-2 ^d /H-2 ^b F₁ mice (C57BL × DBA/2) primed by influenza infection lyse only H-2^d target cells treated with peptide 147–161 while H-2^b targets are recognized only after treatment with NP residues 365–379 (previously found to be recognized by D^b restricted Tc cells). Tc cell recognition of NP peptide 147–161 is entirely dictated by expression of K ^d and not by other B10 or OH background genes of congenic mice. Restriction of a unique NP sequence by each responder class I major histocompatibility complex (MHC) allele suggests that antigen and class I MHC interact for Tc recognition.     

Human helper T-cell clones that recognize different influenza hemagglutinin determinants are restricted by different HLA-D region epitopes

Human T-lymphocyte clones (TLCs) were generated against the hemagglutinin (HA) of A/Texas/1/77 influenza virus by limiting dilution. TLCs were then screened for antigen specificity on chemically synthesized peptides representing the HA1 molecule. It has been hypothesized that different T cells that recognize the identical antigenic determinant are controlled by (restricted by) the same class II epitope. Two TLCs, HA1.4 and HA1.7, both recognized the same HA peptide and in proliferation studies exhibited identical restriction patterns. Two other clones, HA 1.9 and HA 2.43, recognized different HA determinants and also had distinct restriction patterns. Proliferation inhibition studies with monoclonal antibodies against human class II molecules demonstrated three unique patterns of blocking with the clones, suggesting that clones may be restricted to a unique class II epitope depending on the HA determinant recognized. These data can be interpreted as supporting the argument that human immune responses to influenza hemagglutinin are under Ir gene control exerted at the level of the viral antigenic determinant recognized in association with particular D-region restricting elements. The determinant selection and clonal deletion theories are compared for their capacity to best explain these findings.Abbreviations used in this paper ³HTdR tritiated methyl thymidine - MHC major histocompatibility complex - HLA human MHC - PBLs peripheral blood lymphocytes - APCs antigen-presenting cells - TLCs T-lymphocyte clones - TCGF T-cell growth factor - MoAbs monoclonal antibodies     

Target-cell recognition by cloned lines of influenza virus-specific cytotoxic T lymphocytes. Selective inhibition by a monoclonal H-2-specific antibody

A monoclonal H-2^d-specific antibody markedly inhibits target-cell lysis mediated by two influenza virus A/JAP/57-specific, H-2K ^d -restricted cloned CTL lines. Three other A/JAP/57-specific, H-2 ^d -restricted CTL clones (two of which are also restricted to H-2K ^d in target-cell recognition) are only minimally inhibited by this monoclonal antibody. The inhibitory effect of the antibody is not due to selective binding to certain cloned CTL lines but rather is due to blocking of a determinant on the target cell. The monoclonal antibody produces partial inhibition of lysis mediated by a heterogeneous population of A/JAP/57-specific, H-2 ^d -restricted CTL. Likewise the profound, selective inhibition of cytolysis produced by the H-2^d-specific monoclonal antibody could not be reproduced with a conventional H-2^d alloantiserum. These observations suggest that more than one site on a particular H-2K or H-2D molecule can serve as a determinant for H-2-restricted CTL recognition. They furthermore imply that there is more than one recognition structure (receptor) for self MHC products clonally distributed among a population of H-2-restricted CTL directed to a particular antigen.     

Mutations in H-2K b influence the specificity of alloreactive effector cells included in the repertoire of H-2D b -restricted cytotoxic T lymphocytes against Moloney leukemia virus

Moloney leukemia virus-specific cytotoxic T lymphocytes (CTL), generated by secondary in vitro stimulation of spleen cells with syngeneic virus-infected cells, frequently lysed not only syngeneic virus-infected cells, but also noninfected allogeneic target cells. This phenomenon was studied with B6(H-2 ^b ) responder cells and a series of H-2K ^b -mutant responder cells. Thus, B6 Moloney-specific CTL lysed noninfected K ^b -mutant cells, but not B6 cells, whereas K ^b -mutant Moloney-specific CTL lysed noninfected B6 cells and not noninfected cells of the same mutant. Cold-target-inhibition studies showed that the CTL reactions against different allogeneic cells were mediated by different subpopulations of virus-specific CTL: lysis of allogeneic target cells was fully inhibited only by the same allogeneic and by syngeneic virus-infected cells, but not by another allogeneic cell, also lysed by the same effector-cell population. Lysis of syngeneic virus-infected cells could not be inhibited by allogeneic target cells. These data imply that a minority of virus-specific CTL shows cross-reactivity with a given allogeneic target cell. It is concluded that limited amino acid substitutions in the K^b molecule alter the repertoire of Moloney virus-specific CTL, as reflected in alloreactive CTL populations, even though the virus-specific CTL response. of B6 and all K ^b mutants is mainly D^b-restricted. Thus, the development of tolerance to self class-I major histocompatibility complex (MHC) molecules affects the repertoire of self-restricted cytotoxic T cells.     

Nonequivalence of Classical MHC Class I Loci in Ability to Direct Effective Antiviral Immunity

Structural diversity in the peptide binding sites of the redundant classical MHC antigen presenting molecules is strongly selected in humans and mice. Although the encoded antigen presenting molecules overlap in antigen presenting function, differences in polymorphism at the MHC I A, B and C loci in humans and higher primates indicate these loci are not functionally equivalent. The structural basis of these differences is not known. We hypothesize that classical class I loci differ in their ability to direct effective immunity against intracellular pathogens. Using a picornavirus infection model and chimeric H-2 transgenes, we examined locus specific functional determinants distinguishing the ability of class I sister genes to direct effective anti viral immunity. Whereas, parental FVB and transgenic FVB mice expressing the H-2K^b gene are highly susceptible to persisting Theiler''s virus infection within the CNS and subsequent demyelination, mice expressing the D^b transgene clear the virus and are protected from demyelination. Remarkably, animals expressing a chimeric transgene, comprised primarily of K^b but encoding the peptide binding domain of D^b, develop a robust anti viral CTL response yet fail to clear virus and develop significant demyelination. Differences in expression of the chimeric K^bα1α2D^b gene (low) and D^b (high) in the CNS of infected mice mirror expression levels of their endogenous H-2^q counterparts in FVB mice. These findings demonstrate that locus specific elements other than those specifying peptide binding and T cell receptor interaction can determine ability to clear virus infection. This finding provides a basis for understanding locus-specific differences in MHC polymorphism, characterized best in human populations.     

H-2 effects on cell-cell interactions in the response to single non-H-2 alloantigens

Individual mice were tested for their proliferative T-cell response to H-Y- and H-3-incompatible stimulator cells in secondary mixed lymphocyte culture. Responders expressing the H-2 ^bhaplotype were restricted in their response to stimulators presenting H-Y and H-3 in the context of H-2 ^b. Lymphocytes from individual B10 females proliferated in response to H-Y presented with I-A ^band D ^b. The ratio of I-A ^b/D ^b-restricted responses varied between individual responders, indicating significant qualitative variation between genetically identical responders. The majority of the proliferative response in all tested mice was restricted to the entire H-2 ^bhaplotype suggesting complementation of I-A ^b- and D ^b-region genes in presenting the H-Y antigen. Similar observations were made in the response of individual B10.LP mice to the H-3 antigen. H-3-specific, proliferating T cells were restricted to H-3 antigen presented with K ^bA^band D ^bwith significant variation between individuals in their preference for H-3 plus K ^bA^band D ^b. In contrast to the response to H-Y, the proliferative response to H-3 plus H-2 ^bcould be accounted for by the summation of the proliferative responses to H-3. plus K ^bA^band D ^b. These observations demonstrate that the proliferative response to non-H-2 H antigens in the context of I-region determinants is not a sine qua non for the T-cell response to these antigens. Further, the individual qualitative and quantitative variation observed with individual genetically identical mice has strong implications for our knowledge of intrastrain variation in immune responsiveness and the characterization of inbred strains for immune responsiveness.     

Fine specificity and T-cell receptor β-chain gene rearrangements of five H-2Db-specific cytotoxic T-cell clones

A panel of cytotoxic T lymphocyte clones that recognize H-2^b target cells has been established. Six different clones were distinguished according to the following criteria. First, the fine specificity of the clones was determined by testing proliferation and cytotoxicity on target cells of recombinant mice. Clone 221 recognized H-2K^b, and five other clones recognized H-2D^b. Clone 433 distinguished itself from the other five D^b-specific clones by cross-reacting with an antigen on H-2^k cells. Second, the presence of an idiotypic determinant as defined by the 3179 clone-specific monoclonal antibodies was investigated in cytotoxicity inhibition experiments. One of the D^b-specific clones, 653, was inhibited by these antibodies and was therefore clearly different from the other D^b-specific clones. The third criterion involved the rearrangement pattern of the DNA coding for the chain of the T-cell receptor. Southern blot analysis showed that each clone had a unique pattern. Interestingly, clone 653 , which expresses the same idiotypic determinant as clone 3F9, had deleted the C 1 gene cluster, whereas this gene is functionally expressed in clone 3179.Abbreviations used in this paper C constant gene segment - Con A concanavalin A - CTLs cytotoxic T lymphocytes - D diversity gene segment - ³H-dThd tritiated thymidine - J joining gene segment - kb kilobase pairs - LPS lipopolysaccharide - MHC major histocompatibility complex - MLC mixed lymphocyte culture - SDS sodium dodecyl sulfate - V ariable gene segment     

Influenza virus-specific human cytotoxic T cell clones: Heterogeneity in antigenic specificity and restriction by class II MHC products

Human cytotoxic T lymphocytes specific for A/JAP/57 (H2N2) influenza virus were cloned from in vitro stimulations of peripheral blood lymphocytes. Analysis of the viral specificity in cytotoxic function revealed one clone that killed all type A influenza-infected targets, another clone that was specific for the hemagglutinin subtype of the immunizing influenza virus, and the third clone that demonstrated cytotoxicity restricted to the hemagglutinin of A/JAP/57 and A/JAP/62 (H2N2) and not other type A influenza strains with the H2N2 subtypes. The phenotype of these three clones was Leu 2^?, Leu 3⁺, Leu 4⁺; MHC restriction of their cytotoxic function was mapped to HLA-DR by a panel of target cells as well as by inhibition of cytotoxicity with monoclonal antibodies. Proliferation of these clones, examined in a tritiated thymidine incorporation assay, was found to be driven by antigen in the absence of exogenous lymphokines. For all three clones antigen-dependent production and secretion of lymphokines with IL-2 activity was demonstrated. The antigen specificity of proliferation and factor production was shown to be identical to the pattern that each clone revealed in its cytotoxic function.     

Biochemical studies of H-2K antigens from a group of related mutants. II. Identification of a shared mutation in B6-H-2 bm6 , B6.C-H-2 bm7 , and B6.C-H-2 bm9

In an earlier paper, we presented evidence that two independent mutants of the bg series, B6-H-2 ^bm5 (bm5) and B6-H-2 ^bm16 (bm16) carry identical mutations such that tyrosine at residue number 116 of the H-2K^b molecule from the parent strain C57BL/6Kh is replaced by a phenylalanine in each of the two mutant molecules. In this paper, we demonstrate, using similar techniques, that the independent bg series mutants B6-H-2 ^bm6 (bm6), B6.C-H-2 ^bm7 (bm7), and B6.C-H-2 ^bm9 (bm9), which share biological properties with bm5 and bm16, can be grouped together because they share two identical mutations, one of which is common to bm5 and bm16, a Tyr to Phe interchange at residue number 116. In addition, a second mutation is at residue number 121, where a Cys in the H-2K molecule from 136 is substituted with an Arg in the mutant. Since all of the bg series mutants arose independently and share biological and biochemical characteristics, it is anticipated that study of these mutants could lead to some understanding of the high mutation rate in the K^b molecule.