Mice Of Haplotypes Research Articles

Biosynthesis of C4 by mouse peritoneal macrophages. I. Characterization of an in vitro culture system and comparison of C4 synthesis by "low" vs "high" C4 strains.

C4 biosynthesis and secretion in mouse resident peritoneal macrophage cultures was examined by functional and antigenic analysis. The rate of secretion of functional and antigenic C4 decreased in parallel in short-term culture despite constant total protein secretion and increasing factor B secretion. Feedback inhibition and protease degradation did not account for the decreasing rate of C4 secretion. Despite a 10-fold difference in plasma C4 levels between high C4 (H-2d haplotype) and low C4 (H-2k haplotype) mouse strains, resident peritoneal macrophages from both strains synthesized and secreted similar amounts of antigenic and functional C4 in short-term culture. The C4 secreted by H-2d and H-2k macrophages was relatively stable, because it exhibited only approximately 2% loss of hemolytic activity per hour in culture. These data suggest that an unidentified environmental signal leads to the specific decrease in C4 synthesis and secretion by peritoneal macrophages and that a regulatory defect may not be present, at least in macrophages, to account for the low C4 levels in the H-2k haplotype.

Genetic structure and origin of t haplotypes of mice, analyzed with H-2 cDNA probes

We investigated the genetic organization and evolutionary origin of t chromosomes of mice by examining the restriction fragment patterns of DNA from t haplotypes and normal chromosomes with cDNA probes to H-2 class I genes. On genomic DNA blots, the restriction fragments containing H-2-related sequences were highly variable among different inbred strains of mice, whereas they were very similar among different t haplotypes even when the t haplotypes carried serologically different H-2 haplotypes. These observations suggest that all t haplotypes have a common origin and are not products of independent mutational events. We also mapped the position of several restriction fragments characteristic of t DNA by using a battery of recombinant t haplotypes, defined with respect to their t-lethal factors and H-2 haplotypes. We thus show that restriction fragments containing H-2-related sequences map to the left of the H-2 class I genes in t chromosomes, a region in which the t w32 b-lethal factor also maps. The cloning of these fragments can be expected to provide an entry for the structural analysis of t DNA.

Ia antigens are the stimulating determinants in the syngeneic mixed lymphocyte reaction.

The murine syngeneic mixed lymphocyte reaction (sMLR) is a T cell proliferative response to self-antigens on non-T stimulator cells. The nature and specificity of the stimulating antigens was investigated using secondary MLR. Lymph node cells were stimulated in vitro by syngeneic spleen cells. After optimal proliferation occurred, the responding cell population was recovered and restimulated with spleen cells from mice of various haplotypes. The secondary response was found to be specific for the haplotype of the primary stimulator (a self-restricted response). The use of recombinant mice in the secondary stimulation localized the determinant to the I-A and/or I-E subregions of the major histocompatibility complex (MHC). This result was confirmed by the ability of monoclonal antibodies directed to the I-A or I-E subregion products to significantly inhibit the response while alloantiserum directed to products of the K or D regions had no effect.

Genetic control of the cytolytic T lymphocyte response to influenza viruses: H-2 genes influence the response to H-2Kb plus virus.

Genetic control of the cytolytic T lymphocyte responses to influenza virus was examined. Mice bearing the H-2b haplotype and F1 hybrid mice derived from C57BL/6 parents failed to recognize H-2Kb plus type A or type B influenza viruses. Congenic mice that shared H-2Kb but had either H-2Dd of H-2Dq genotypes responded in in vitro secondary cultures to H-2Kb plus virus. Mapping studies suggest the gene(s) controlling the ability to respond to H-2Kb plus virus can be localized in or near the H-2D region. Several interpretations of these observations are discussed.

Identification of D-locus alleles in mouse B10.W lines.

B10.W lines carry H-2 haplotypes of wild mice on the C57BL/10Sn (=B10) background (reviewed by Klein and Figueroa 1981). The serological analysis of these lines has been carried out in steps. In the first step, the wild-derived H-2 haplotypes had been identified with the help of antibodies specific for known class I antigens and antisera produced in (A x B10)F1 hybrids against the individual lines (Klein 1972, Zaleska-Rutczynska and Klein 1977, Duncan and Klein 1980). However, this first-step analysis has left D alleles in several of the lines unidentified because, for some unknown reason, most of the (A x B10)F1 anti-B10.W sera reacted with K allomorphs (products of K alleles) but not with the D allomorphs. In the second step we have selected new strain combinations (Table 1), some of which should differ at the D locus, and have attempted to produce antisera in these combinations. The antisera were produced by multiple injections of donor-strain spleen, lymph-node, and thymus cells according to the protocol described elsewhere (Klein et al. 1975). They were analyzed using the two-stage microcytotoxicity test with lymph-node lymphocytes as target cells and a 1:1:8 mixture of normal rabbit serum, normal guinea pig serum and Hanks' balanced salt solution containing 10% fetal calf serum as the source of complement (Klein et al. 1975). For absorptions, an antiserum was incubated for 30 rain at room temperature with equal volume of packed lymph-node and spleen cells, and then again, under the same conditions, with fresh cells. Each antiserum was first tested against the panel of the classical inbred strains (H-2 haplotypes b, d, f , j , k, p, q, r, s, u, v, and z) and if it reacted with any of these strains, it was absorbed to remove this reactivity. The preabsorbed antiserum was then tested against the panel of 31 B 10.W lines and checkerboardtype absorptions were done with all positive strains. The nine antisera analyzed in this fashion (Tables 2 and 3) have permitted the identification of six D alleles in the B10.W lines: D wl, D wS, D wv, D r, D w16, and D z. The identification of the individual alleles will now be described.

H-2-associated differences in replicated strains of mice divergently selected for body weight.

A random-bred strain (Q) was established and divided into six replicates. Each replicate was divergently selected for 6-week weight (for over 30 generations) and each had an unselected control. We have investigated the H-2 haplotype of individual mice of the 18 selected Q strains to determine whether selection for size had also selected for H-2 or H-2-linked genes. From the results it appeared that only the H2b and H-2q haplotypes were present in the foundation stock. A large number of individuals of the six small sublines were of H-2b haplotype, while the majority of those of the six large sublines were of the H-2q haplotype. Individuals in the six control strains were H-2b, H-2q or both (i.e., H-2 heterozygotes and/or H-2 recombinants). These results suggest that control of body size is associated with H-2 or an H-2-linked gene(s).

Variable synthesis and expression of E alpha and Ae (E beta) Ia polypeptide chains in mice of different H-2 haplotypes.

The genetic and molecular requirements for cell-surface expression of Ia antigens precipitated by anti-I-E subregion sera have been examined. Inbred mice f the d, k, p, and r haplotypes synthesize and express on their lymphocytes the two I-region products normally found in anti-I-E-subregion immunoprecipitates, E alpha and Ae (E beta). Cells from mice of the b and s haplotypes fail to synthesize E alpha chains but do synthesize Ae chains, which remain in the cytoplasm as partially glycosylated precursors. Cells of the f and q haplotypes fail to synthesize either the Ae or E alpha polypeptide chains, as shown by both genetic complementation tests and analyses of total cell proteins by two-dimensional polyacrylamide gel electrophoresis. The patterns of expression of the intact E alpha: Ae complex are consistent with the theory that both the Ae and E alpha polypeptide chains must be present in the cells for either chain to be expressed in normal amounts on the cell surface. The implications of these observations for the genetics of I-region-controlled functions are discussed.

The expression of anti-poly(LGlu60, LPhe40) idiotypic determinants dictated by the gene products in the major histocompatibility complex.

Antibodies raised in SWR/J mice (H-2q, Igc) to the random copolymer poly(LGlu60, LPhe40) (GPhe) were purified by immunoadsorbent chromatography and used to immunize a New Zealand red rabbit. The rabbit anti-idiotypic antiserum thus produced strongly inhibited the binding of 125I-GPhe by anti-GPhe antisera produced only in mice of H-2q haplotype, and had no effect on the binding of GPhe by anti-GPhe antisera produced in mice of other haplotypes, namely, H-2k and H-2p. The anti-idiotypic antiserum also inhibited the binding of GPhe by anti-GPhe-methylated bovine serum albumin antisera produced only in mice of H-2q haplotype. No linkage to Ig allotype was observed. The anti-GPhe antisera produced in F1 mice the anti-idiotypic antiserum demonstrating the dominant presence in these F1 mice of idiotypic determinants whose expression is dictated by the H-2q major histocompatibility complex (MHC). The anti-idiotypic antiserum also inhibited the binding of 125I-poly)LGlu56, LLys35, LPhe9) and 125I-GPhe antisera produced only in mice of H-2q haplotype. These specificities were also confirmed by the inhibition of the plaque-forming cells. It was concluded that the antibodies produced in mice of H-2q haplotype against GPhe and GLPhe share common idiotypic determinants that are recognized by the anti-idiotypic antiserum. A possible explanation for the unique findings that the expression of anti-GPhe idiotypic determinants in mice of H-2q haplotype are dictated by the gene product in the MHC is that the macrophages in mice of H-2q haplotype present unique determinants of GPhe polymer in the response process to GHphe.

GENETIC CONTROL OF THE IMMUNE RESPONSE TO HAEMOGLOBIN: I. DEMONSTRATION OF SEPARATE GENETIC CONTROL OF THE RESPONSES TO THE α‐ AND β‐SUBUNITS BY IN VITRO LYMPHOCYTE PROLIFERATION

These studies were undertaken to analyse the genetic control of the immune response to an oligomeric protein and the role of individual subunits in the regulation of the response. Human adult haemoglobin (Hb) was selected as a model for these studies because it is a well-characterized protein and its antigenic structure is being determined in our laboratories. Mice of various congenic strains were immunized with Hb and the lymph node cells from Hb-primed mice were challenged in vitro with Hb, and its alpha-chain and beta-chains as well as an appropriate control antigen. Lymphocyte proliferation was determined by 3H-thymidine incorporation. The data collected indicated that mice of the H-2b and H-2d haplotypes were high responders while H-2k, H-2s, H-2q and H-2j haplotype mice were low responders to Hb. Studies with H-2 recombinant strains indicated that the immune response to Hb and its subunits is determined by genes in the I-A subregion and the D end of the H-2 complex. The significance of these findings in terms of control and regulation of the overall response to native Hb are discussed.

A model of T-cell unresponsiveness using the male-specific antigen, H-Y

The immune response to H-Y, a simple cell surface determinant, is clearly extraordinarily complex. It is controlled by H-2 genes mapping at the K and D loci, as well as by I-region genes. It is manifest by skin graft rejection, H-2 restricted cytotoxicity, delayed-type hypersensitivity, and host-versus-graft responses. Suppressor cells may very well be responsible both for neonatally induced tolerance and for nonresponsiveness in adult mice of certain haplotypes. The in vivo responses of graft rejection and DTH have so far been seen only in mice of the H-2 b haplotype whereas the in vivo host-versus-graft response and the in vitro cytotoxic response have been found in a wider range of H-2 types, although a variety of strategies need to be used in order to reveal responsiveness in some haplotypes. We do not yet know which of these responses are important in antitumour immunity, but the findings so far have alerted us to the possibility that MHC antigens of the responder are very important and that certain refractory states of nonresponsiveness can be reversed. Understanding of the underlying mechanisms and cell-cell interation during the generation of immune responses is vital for a rational approach to the control of antiallograft and antitumour responses.

IgD allotypic determinants. II. Determinants expressed on IgD of mice of the b, e, and o Igh-C haplotypes.

Allotypic determinants expressed on membrane IgD of 27 strains of mice representing each of the major Igh-C haplotypes of inbred mice were examined with reagents specific for the IgD of mice of the b, e, and o Igh-C haplotypes. The distribution of two specificities, IgD.44 (Ig5.2) and IgD.37 (Ig5.3), recognized by two monoclonal anti-delta reagents was determined. In addition, a new specificity, IgD.46, expressed on mice of the o Igh-C type, but not of the b or a types, was detected by a BALB/c anti-AL/N spleen cell antiserum. The results also confirmed the description of the new Igh-C haplotype o, by demonstrating that the mice of this group express the IgD.37 determinant. This determinant is also expressed by mice of the b and e types, but not by mice of the d haplotype, from which the o haplotype was subdivided.

Aberrant Ae (E beta) Ia polypeptide chain in H-2g2 haplotype mice. Possible result of an intragenic recombination or mutation.

Mice of strains D2.GD and B10.GD, which carry the recombinant haplotype H-2g2 (previously typed as H-2Kd I-Ad / I-Bb I-Jb I-Eb I-Cb Sb H-2Db), have an Ae (E beta) polypeptide chain electrophoretically distinct from the Ae chains of both b and d haplotype mice, including the progenitor strains from which the recombinant H-2g2 chromosome was derived. The evidence presented suggests that the altered molecular properties of the Aeg2 chain may be a consequence of an intragenic recombination event in the I-A subregion within the structural gene for this polypeptide chain. Because the A alpha and A beta chains controlled by H-2g2 appear to be d haplotype in origin, this finding would map the gene for Ae to the right of the loci controlling the A alpha and A beta chains

Variable expression and immunogenicity of an H-2K-coded alloantigen on murine tumours.

The K region of the H-2 major histocompatibility complex (MHC) of mice of H-2Kk haplotype codes for two distinct alloantigens. One of these alloantigens, designated k-common, is expressed by C3HfB/HeN mice (C3Hf). The other alloantigen, designated k-unique, is not expressed by C3Hf mice. The H-2 haplotype of C3Hf mice has been classified as kv1 and the variant antigen distinguishing this strain from mice of H-2Kk haplotype has been designated kv1-unique. Several transplacentally-induced lung tumours of C3Hf mice express the k-unique, rather than the expected kv1-unique, antigen. The immunogenicity of the k-unique antigen on C3Hf-derived lung tumours varies with different tumours. In particular, the capacity of the k-unique antigen to induce radioresistant immunity in C3Hf mice appears to be lost on long term cultured tumour cells even though the tumour remains susceptible to in vivo immune responses directed against the k-unique antigen. Alterations in expression and in immunogenicity of unique H-2-coded antigens may dictate the nature and efficacy of immune surveillance of autochthanous tumours.

Complementation ofH-2-linkedIr genes: Use of helper factor to analyze responses to GLPhe

Responses to GLPhe5 were generated in vitro, using the GLPhe helper factor. A plaque assay, using GLPhe-coated SRC, was used to measure antibody responses to GLPhe. Mice of differentH-2 haplotypes were tested for the ability to produce the GLPhe helper factor or to respond to a known GLPhe helper factor. We report here defects in nonresponder mice of different strains in both the production of helper factor and in the capacity to respond to it.

Two-Gene Control of the Expression of Murine Ia Antigens

Abstract Two-dimensional polyacrylamide gel electrophoresis of Ia antigens obtained from 35S-methionine-labeled lymphocytes has revealed that the expression on the cell surface of an antigen coded for by a locus in the I-A subregion may be controlled by an I-E subregion product. Three distinct polypeptide chains (Aa, Ab, and Ae) coded for by loci in I-A have been identified in gels of immunoprecipitated Ia antigens; one of these (Ae) is coprecipitated with an I-E antigen by anti-I-E antibodies and probably is complexed to the I-E chain in the cell. Mice of haplotypes which do not have serologically-detectable I-E antigens (such as b and s haplotypes) appear not to synthesize and I-E product. These mice also do not express Ae on their cell surfaces, although precursors of surface Ae can be detected in the cytoplasm. Some haplotypes, however, contain a locus mapping between Ia.4 in I-J and H-2D which can complement I-A in either the cis or trans position to allow the cell surface expression of Ae.

Lung tumor-associated derepressed alloantigen coded for by the K region of the H-2 major histocompatibility complex

Transplacental induction of lung tumors in C3HfeB/HeN (C3Hf) strain mice can be readily achieved with the carcinogen 1-ethyl-1-nitrosourea. Several of these tumors express, as a tumor-associated transplantation antigen (TATA), a normal tissue alloantigen present in strain A and C3H/HeN (C3H) mice. In the present study it was shown that the tumor-associated alloantigen on the C3Hf-derived lung tumor 85 was present in all mice of H-2(a) and H-2(k) haplotypes tested and in CBA (532) strain mice (H-2(ka) haplotype). Studies using congenic-resistant and recombinant strains of mice indicated that the genetic locus controlling the expression of this antigen was either within or to the left of the H-2K region of the major histocompatibility complex (MHC). Thus the antigen was expressed in B10.A (4R) mice (kkbbbb MHC haplotype) but not in B10 (bbbbbb) or B10.AQR mice (qkkddd). The antigen was expressed in all tissues tested of C3H and A strain mice. It was not detected on any tissue tested including embryo tissue of C3Hf mice or mice of MHC haplotype other than H-2(k) or H-2(a). Because C3Hf strain mice were originally derived from C3H strain mice (H-2(k)), the MHC haplotype of C3Hf mice has been provisionally designated H-2(kb). The finding of a tumor-associated change in the expression of a H-2K region-coded antigen is consistent with the concept that MHC-coded antigens may act as targets for immunological surveillance of tumors.

Host Defenses in Experimental Scrub Typhus: Genetics of Natural Resistance to Infection

Genetic resistance to lethal infection with Rickettsia tsutsugamushi was studied in over 30 inbred strains, inbred hybrids, and outbred stocks of mice. Inbred mice infected intraperitoneally with the Gilliam strain of R. tsutsugamushi showed three patterns of response: susceptible (A/HeJ, C3H/HeDub, C3H/HeJ, C3H/HeN, C3H/St, CBA/J, DBA/1J, DBA/2J, and SJL/J), resistant (AKR/J, BALB/cDub, BALB/cJ, C57BL/6J, C57L/J, and SWR/J), and selectively resistant (A/J). The selectively resistant pattern was characterized by random deaths occurring throughout the titration range and was also observed in three of the six outbred mouse stocks surveyed. No correlation was evident between the H-2 haplotype of inbred mice and their response to Gilliam infection. The progeny from five different Gilliam-resistant by Gilliam-susceptible inbred parental crosses were all resistant. Study of F(1), F(2), and parental backcross generations of BALB/cDub (resistant) and C3H/HeDub (susceptible) hybrids indicated resistance was dominant and was controlled by a single gene or a closely linked cluster of genes that were autosomal and not linked to coat color. The resistance of BALB/cDub mice was not due to an inability of host cells to support rickettsial growth, since C3H/HeDub and BALB/cDub embryo cell cultures supported similar growth of Gilliam organisms. C3H/HeDub mice, although susceptible to intraperitoneal Gilliam infection, were capable of mounting an immune response to Gilliam antigens, since subcutaneous infection was not lethal and did protect animals against subsequent intraperitoneal challenge with either the Gilliam or Karp strains of R. tsutsugamushi.

Genetic control of the antibody response to sperm whale myoglobin in mice.

The antibody response to the main chromatographic component IV of sperm whale myoglobin in mice has been shown to be under the control of both H-2-linked and non-H-2-linked genes, using an assay which measures antibody concentration independent of affinity. The effect of non-H-2 background genes was such that for a given H-2 haplotype, mice of the A background were higher responders than those of the B10 background. Among congenic mice of all the same background, mice of haplotypes H-2d,s were high responders, while those of haplotypes H-2k,b,q were low responders. Use of intra-H-2 recombinant strains B10.A, B10.A(5R), A.TL, and D2.GD allowed mapping of at least two H-2-linked Ir genes, both within the I region, one tentatively mapping in I-A, designated Ir-Mb-1, and one mapping to the right of I-E and left of H-2D, designated Ir-Mb-2. Strains bearing both genes or only Ir-Mb-1 were high responders, while those bearing Ir-Mb-2 were intermediate responders. When the concentration of antimyoglobin antibodies which bound to [14C]-fragment (132-153) of myoglobin was measured in the same sera described above, levels in the B10.A sera were low like those of the B10.BR rather than intermediate between the levels of the B10.BR and the high responding B10.D2, as for myoglobin, whereas levels in the D2.GD sera were still high. It is therefore suggested that the 2 or more genetically defined H-2-linked Ir genes each control the response to a different chemically defined determinant or group of determinants on myoglobin. Further studies with other fragments are in progress to test this hypothesis.

Unidirectional skin graft incompatibility between theH-2 i5 andH-2 i7 mouse haplotypes

Unidirectional skin graft incompatibility between theH-2 i5 andH-2 i7 mouse haplotypes

Immunosuppressive Factor(s) Specific for L-Glutamic Acid50-L-Tyrosine50 (GT)

Abstract The random synthetic copolymer of L-glutamic acid50-L-tyrosine50 (GT) fails to elicit a GT-specific antibody response in all inbred strains of mice tested. Preimmunization with GT specifically inhibits a GT-MBSA response in certain, H-2d,k,s, but not other, H-2a,b,q, nonresponder mice. This unresponsiveness is mediated by GT-specific suppressor T cells. Extracts prepared from lymphoid cells of GT-primed suppressor haplotype mice inhibit the development of primary GT-specific antibody responses to GT-MBSA in normal syngeneic mice. Nonsuppressor haplotype mice do not produce GT-specific suppressor factor. The GT-suppressive extract has affinity for antigen and a m.w. of less than 50,000 daltons, thus, resembling antigen-specific immunosuppressive factors already described. However, the GT-suppressive extract does not appear to have H-2 restrictions since it works across allogeneic barriers. Evidence is presented that two genes are required for factor-mediated suppression.