Demonstration of a new genetic locus in the major histocompatibility system of the rat

A new recombination within the major histocompatibility complex (RT1) of the rat has been detected. The recombination occurred between a wild-derived haplotype, provisionally designated p1, and the RT1 haplotype of the BN strain. The recombinant haplotype, designated p3, carries the RT1.A locus (classical histocompatibility antigens) of the BN strain, a locus from the BN strain that codes for the expression of an Ia antigen and strong mixed lymphocyte response (MLR), and a second locus derived from the p1 haplotype that controls the expression of a second Ia antigen, the ability to elicit a strong MLR and the immune response to poly(G1u⁵²Lys³³Tyr¹⁵). This recombinant therefore demonstrates the division of the RT1.B region into two loci, tentatively designated RT1.B and RT1.D, and provides evidence for the existence of at least four loci in the MHC of the rat.     

A three-locus model for the chicken major histocompatibility complex

The major histocompatibility complexes (B complexes) of chickens of various origins have been studied by serological and biochemical methods. TwoB complexes are of particular interest:B ^R1, a recombinant haplotype derived from theB complexes of the inbred CB and CC strains, andB ^G-B1 , theB haplotype of the G-B1 strain. TheB ^R1 haplotype differs detectably from theB ^CB haplotype only at a locus controlling the synthesis of an antigen, B-G, which (in peripheral blood) is present only on red cells. Anti-B-G sera precipitate, from¹²⁵I-labeled red cell lysates, two chains of apparent molecular weights 42,000 and 31,000 (measured under reducing conditions); the smaller is perhaps derived by proteolysis from the larger. TheB ^G-B1 haplotype differs detectably from theB ^CC haplotype only at a locus controlling the synthesis of an antigen whose tissue distribution and biochemical and biological properties are identical to those of B-G. The chicken major histocompatibility complex therefore contains at least three loci—those controlling synthesis of the B-G, and of the previously defined B and B-L antigens.The following abbreviations are used in this paper MHC major histocompatibility complex - SDS-PAGE sodium dodecyl sulfate polyacrylamide gel electrophoresis - MLR mixed leukocyte reaction - GVH graft-versus-host reaction     

Ia antigens in inbred rats and their relationship to the MLR phenotype.

Three different alloantisera were raised by using Ag-B/MLR disparate rats, and the cytotoxic activity remaining after absorption with erythrocytes to remove anti-Ag-B antibodies was examined. The alloantisera detected surface antigens present only on B cells and segregation studies demonstrated that the genes that code for these antigenic specificities were linked to the major histocompatibility complex. The reactivity of the alloantisera with splenic lymphocytes from a panel of strains representative of the currently known Ag-B groups showed that multiple specificities were present in two of the three antisera and that these specificities were shared by many inbred strains. The appropriate absorption studies showed, however, that each antiserum detected an unique specificity that was found only in those inbred strains that shared the same mixed lymphocyte reactivity (MLR) phenotype as the donor strain. The alloantiserum produced against the KGH strain inhibited the MLR reactions involving this strain only when it was used as the stimulating cell population. The antigens detected by the three alloantisera described here have the characteristics of Ia antigens, and they have tentatively been designated Ia.1 (ACI anti-KGH), Ia.3 (B3 anti-BN) and Ia.4 (MNR anti-DA).     

Polymorphism of the major histocompatibility locus in the wild Norway rat

Specific alloantisera against the eight Ag-B groups found in inbred strains of rats were capable of reacting with all wild Norway rats (Rattus norvegicus) tested. Absorption studies, antisera production, and progeny testing involving wild rats showed that the antigenic specificities detected in the wild rat population were similar, if not identical, to the Ag-B antigens present in inbred strains. Xenoantisera prepared in rabbits against rat erythrocyte antigens (Ag-C1 and/or C2) reacted with erythrocytes from each wild rat tested. Progeny testing involving these erythrocyte antigens was identical to that observed in inbred strains. The restricted genetic polymorphism of theAg-B alleles in the wild rat population suggests that the functional and evolutionary significance of the major histocompatibility complex in the rat may not depend upon a high degree of genetic variability.     

Genetic analyses of alloreactions between recently wild and classical inbred strains of syrian hamsters: Evidence in favor of a major histocompatibility complex

Cytotoxic alloantisera were raised between recently wild and classical inbred strains of Syrian hamsters. Antisera produced by immunizing the classical inbred strains with tissue from the partially inbred, recently wild hamsters detect several specificities shared between the classical and recently wild strains. Reciprocal mixed lymphocyte reactions between the two different groups of hamsters suggest that the new source of hamsters possesses several unique MLR phenotypes which may represent new Hm-1 haplotypes. Moreover, several recently wild strains express MLR phenotypes quite similar if not identical to the Hm-1 ^a haplotype of the inbred strain, MHA. Genetic analyses of alloreactions between domestic inbred and recently wild strains suggest that a single locus or chromosomal region encodes the allodeterminants that induce strong MLR reactivity. Six unique MLR phenotypes have been defined which most likely represent haplotypes of the hamster MHC equivalent, Hm-1. Genetic linkage studies indicate that some alloantisera detect determinants encoded by loci closely linked to the MLR locus, and therefore define Hm-1 determinants. Moreover, other alloantisera recognize determinants encoded by a locus that is unlinked to Hm-1. These studies suggest that Syrian hamsters express a polymorphic MHC equivalent, Hm-1, which encodes determinants that induce both cell-mediated and humoral alloreactivity.     

Segregation of allotypes in a strain of chickens homozygous for theB locus

Sublines homozygous for alleles controlling IgM and IgG allotypes at two linked loci (M1 andG1) were established in the B14 strain of chickens, which is homozygous for theB locus controlling major histocompatibility antigens. Transfer of lymphoid cells resulted in a permanent donor-type IgG synthesis in irradiated 12-day-old recipients, but was followed by protracted rejection in chickens injected embryonally.     

Murine responses to (Tyr-Glu-Ala-Gly)n

A series of known sequential polypeptides is being synthesized and used in our laboratory to study the contribution of antigen structure, i. e., amino acid sequence and conformation in antigen recognition and specificity of the immune response. The capacity to respond to one such -helical polypeptide (T-G-A-Gly)_n, is T-cell dependent and restricted to mice of theH- 2^b haplotype. The response is controlled by anIr gene mapping to theK region and/or theIA subregion which allows the animal to make both a T-cell mediated response, as well as a humoral response to the polypeptide. The response of three mutant strains at theK end of the major histocompatibility locus (MHC) need not differ from that of the responder parental haplotype.PETLES obtained from mice possessing a responder haplotype proliferate when cultured in vitro with (T-G-A-Gly)_n. The antibody level of individual inbred mice of a given strain at a given time differs significantly (from 80% binding to less than 10% antigen bound in 3 out of 57 mice). There is also great individual variability in time of appearance of the antibody response and where peak optimal levels are seen. Possible explanations for the variation in the antibody expression include: (a) the polymer is a weak immunogen, (b) the presence of modifier gene(s) outside of the major histocompatibility complex controlling the magnitude of the antibody level, (c) the possible effect of the polymer which is a B cell mitogen as a generator of suppressor T cells and, (d) a feedback mechanism effect on B cells controlling the antibody level.This work was supported in part by the following grants: The National Institute of Allergy and Infectious Diseases A107825; American Cancer Society Grant IM-5F; National Foundation-March of Dimes 1-492.     

Immune response to the Slp alloantigen in the mouse:H-2-linked qualitative and quantitative control

Immunization of inbred mouse strains lacking the Slp allotype results in the production of Slp antibodies in some strains but elicits no detectable response in other strains. Analysis of standard inbred and congenic resistant strains reveals that both the qualitative and quantitative ability to respond to the Slp allotype is associated with theH-2 haplotype of the recipient. Three different response phenotypes can be identified utilizing complement fixation and quantitative immunodiffusion tests. Strains which carry theH-2 ^q haplotype are high responders,H-2 ^k strains are intermediate in response, whileH-2 ^b andH-2 ^v strains produce no detectable antibody. The characteristic response patterns of high and intermediate responders were manifest by day 35 of immunization and continued as discrete response types after a second booster. Quantitative data in the immune response of the intra-H-2 recombinant B10.A(4R) suggest that the recombination event which established theH-2 ^h4 chromosome disturbs the proper function of the genetic determinant controlling response to Slp.     

Gene order in the major histocompatibility complex of the rat

The loci in the major histocompatibility complex (MHC) of the rat which code for class I and class II antigens—RT1.A and RT1.B, respectively — have previously been separated by laboratory-derived recombinants and by observations in inbred and wild rats. Closely linked to the MHC is the growth and reproduction complex (Grc) which contains genes influencing body size (dw-3) and fertility (ft). These phenotypic markers were used in this study to orient the A and B loci of the MHC. Two recombinants were used for mapping. The BIL(R1) animal is a recombinant between the MHC and Grc, and it carries the haplotype RT1.A ^lB^lGrc⁺. The r10 animal is an intra-MHC recombinant, and it has the haplotype RT1.A ⁿB¹ Grc. These recombinants were characterized serologically, by mixed lymphocyte reactivity, by immune responsiveness to poly (Glu⁵²Lys³³Tyr¹⁵) and by the presence of the dw-3 gene. The data demonstrate that the gene order of the loci is: dw-3-RT1.B-RT1.A.     

Ly-m11: TheH-3 region of mouse chromosome 2 controls a new surface alloantigen

Spleen cells from an SJL mouse immunized with B10.S spleen cells were fused with the nonsecretor myeloma line NS.1. One established hybrid cell line continuously secreted antibody that recognized a new antigenic specificity, tentatively called Ly-mll. This newly found antigen is detectable on nearly 100 percent of spleen and lymph-node cells, 70 percent of bone-marrow cells, and 20 percent of thymus cells by direct cytotoxicity assays, and on the cells derived from kidney and liver. Strains that are Ly-mll (+) include C57BL/6, C57BL/10J, B10.S, C57BR/cdJ, C57L/J, and C57BL/KsJ. Other mouse strains so far tested are Ly-m11 (–). The strain distribution pattern distinguished Ly-mll from any known murine lymphocyte alloantigens, but it follows theH-3 ^a haplotype which is defined by skin transplantation. Linkage tests of nine congenic strains ofH-3 and/orH-13/a loci and five recombinant inbred lines including CXB, BXH, AKXL, SWXL, and BXD revealed no recombinations betweensH-3 andLy-m11 loci on chromosome 2. This newly discovered Ly-m11 alloantigen could itself constitute a minor histocompatibility antigen detectable by serological means.Abbreviations used in this paper RI recombinant inbred - H histocompatibility - a non-agouti - B10 C57BL/10Sn The prefix m (monoclonal) is used following a suggestion by Klein and co-workers (1979).     

Considerable haplotypic diversity in the RT1-CE class I gene region of the rat major histocompatibility complex

The major histocompatibility complex (MHC) class I region extending between the Bat1 and Pou5f1 genes shows considerable genomic plasticity in mouse and rhesus macaque but not in human haplotypes. In the rat, this region is known as the RT1-CE region. The recently published rat MHC sequence gave rise to a complete set of class I gene sequences in a single MHC haplotype, namely the RT1ⁿ haplotype of the widely used BN inbred strain. To study the degree of genetic diversity, we compared the RT1-CE region-derived class I genes of the RT1ⁿ haplotype with class I sequences of other rat haplotypes. By using phylogenetic tree analyses, we obtained evidence for extensive presence and absence polymorphisms of single loci and even small subfamilies of class I genes in the rat. Alleles of RT1-CE region class I genes could also be identified, but the rate of allelic nucleotide substitutions appeared rather low, indicating that the diversity in the RT1-CE region is mainly based on genomic plasticity.     

BALB/c-H-2 db : A newH-2 mutant in BALB/cKh that identifies a locus associated with theD region

A newH-2 mutant, BALB/c-H-2 ^db , is described. This mutant originated in BALB/c, is inbred, and is coisogenic with the parental BALB/cKh strain. The mutation is of the loss type since BALB/c-H- ^db rejects BALB/c, but not vice versa. Complementation studies have localized the mutation to theD region of theH-2 complex. A cross between BALB/c-H-2 ^db and B10.D2-H-2 ^da failed to complement for either BALB/c or B10.D2 skin grafts, indicating that these are two separate mutations at the same locus (Z2). Direct serological analysis and absorption studies revealed that, with one exception, theH-2 andIa specificities of BALB/c and BALB/c-H-2 ^db are identical. In particular,H-2.4, the H-2D^d private specificity, is quantitatively and qualitatively identical in the two strains. The exception is that of the specificities detected by antiserum D28b: (k×r)F₁ anti-h, which contains anti-H-2.27, 28, and 29. These specificities appear to be absent from theH-2 ^db mutant since they are not detected directly or by absorption. Other public specificities are present in normal amounts,e.g., the reaction with antisera to H-2.3, 8, 13, 35, and 36. The reaction with antiserum D28 (f×k)F₁ anti-s, which contains antibodies to H-2.28, 36, and 42, is the same in both strains. Antiserum made between the two strains (H-2 ^db anti-H-2 ^d ) reacts like an anti-H-2 serum, in that it reacts with both T and B cells by cytotoxicity, but is not a hemagglutinating antibody. The serum reacts as does the D28b serum in both strain distribution and in cross-absorption studies. We conclude that theH-2 ^db mutation occurred at a locus in theD region, resulting in the loss of the H-2.28 public serological specificity and of a histocompatibility antigen. Whether these are one and the same antigen is not yet known. The data, in view of other evidence, imply that the public and private specificities are coded for by separate genes.Abbreviations used in this paper are as follows CML cell-mediated lysis - MLR mixed lymphocyte reaction - GVHR graft-versus-host reaction - RFC rosette-forming cells - RAM-Ig rabbit anti-mouse IgG     

H-2-linked recessiveIr gene regulation of high antibody responsiveness to TNP hapten conjugated to autogenous albumin

The antibody response to the hapten 2,4,6-trinitrophenyl (TNP) conjugated to autogenous mouse serum albumin (MSA) is regulated by anIr gene(s) located within the major histocompatibility complex (MHC). Both the qualitative and quantitative ability of congenic strains to produce TNP-specific antibodies are functions of theH-2 haplotype. Thus, mouse strains may be classified as high (H-2 ^d), intermediate (H-2 ^b,H-2 ^s), and low responders (H-2 ^a,H-2 ^k,H-2 ⁿ,H-2 ^p,H-2 ^q). Antibody responses, as measured by antigen-binding capacities in modified Farr assays, were compared among strains carrying recombinantH-2 haplotypes and their hybrid progenies. Distinct high- and low-responder phenotypes were evident throughout the time course of both primary and secondary antibody responses. The gene locus controlling specific responsiveness to TNP-MSA, now designatedIr-6, was mapped within theI-B subregion of theH-2 complex. Recessive inheritance of high responsiveness was confirmed in hybrid progenies of three different low × high-responder crosses.     

Genetic analysis of resistance to Type-1 Diabetes in ALR/Lt mice,a NOD-related strain with defenses against autoimmune-mediated diabetogenic stress

ALR mice are closely related to type-1 diabetes mellitus (T1DM)-prone NOD mice. The ALR genome confers systemically elevated free radical defenses, dominantly protecting their pancreatic islets from free radical generating toxins, cytotoxic cytokines, and diabetogenic T cells. The ALR major histocompatibility complex (MHC) (H2^gx haplotype) is largely, but not completely identical with the NOD H2^g7 haplotype, sharing alleles from H2-K through the class II and distally into the class III region. This same H2^gx haplotype in the related CTS strain was linked to the Idd16 resistance locus. In the present study, ALR was outcrossed to NOD to fine map the Idd16 locus and establish chromosomal regions carrying other ALR non-MHC-linked resistance loci. To this end, 120 (NOD×ALR)×NOD backcross progeny females were monitored for T1DM and genetic linkage analysis was performed on all progeny using 88 markers covering all chromosomes. Glucosuria or end-stage insulitis developed in 32 females, while 88 remained both aglucosuria and insulitis free. Three ALR-derived resistance loci segregated. As expected, one mapped to Chromosome 17, with peak linkage mapping just proximal to H2-K. A novel resistance locus mapped to Chr 8. A pairwise scan for interactions detected a significant interaction between the loci on Chr 8 and Chr 17. On Chr 3, resistance segregated with a marker between previously described Idd loci and coinciding with an independently mapped locus conferring a suppressed superoxide burst by ALR neutrophils (Susp). These results indicate that the Idd16 resistance allele, defined originally by linkage to the H2^gx haplotype of CTS, is immediately proximal to H2-K. Two additional ALR-contributed resistance loci may be ALR-specific and contribute to this strain's ability to dissipate free-radical stress.     

Molecular analysis of the MHC class II region inDR4, DR7, andDR9 haplotypes

Within the class 11 region of the major histocompatibility complex (MHC) the amount of DNA in theDR-DQ interval has been shown to be haplotype dependent, with those carrying the DR4, DR7, and DR9 specificities having been reported to contain 110–160 kilobases (kb) more DNA than haplotypes carrying the DR3 specificity. Certain subtypes of haplotypes carrying particular DR specificities are more closely associated with autoimmune diseases than others. With the prospect of the DNA perhaps containing a disease susceptibility locus, we have mapped eight DR4 and two DR7 homozygous cell lines and a DR7/9 heterozygous cell line together with a control DR3 cell line using pulsed field gel electrophoresis (PFGE) with the enzymesBss H II,Pvu I, andNot I/Nru I. Our results, however, show that the presence and amount of the extra DNA is constant irrespective of the subtype. We have also tried to narrow down the position of insertion of the extra DNA using eight further rare-cutting enzymes but, due to the polymorphic nature of sites and/or differences in methylation in this region, it was not possible to refine it further than between DRA and DQA1/B1. This polymorphic nature of theDR-DQ region is unusual, considering the uniformity of rare cutter sites that has been observed within the rest of the class II, and class III, regions. The presence of this, and other, haplotype dependent variations in the DNA content of theDR subregion may be important with respect to recombination and will be particularly interesting if the additional DNA is found to contain novel genes.     

Rat interstrain antibody response and crossidiotypic specificity

Interstrain analysis of the humoral response of rats to streptococcal group A carbohydrate (SACHO) 1, employing seven inbred strains representing six histocompatibility haplotypes at the Ag-B locus, suggests that the immune response genes to SACHO are not linked to the major rat histocompatibility locus. The low-precipitin response of all seven inbred rat strains was similar to the precipitin response of F₇ Sprague-Dawley rats selectively bred for a low-precipitin response to SACHO. Although strain differences were not apparent in the magnitude of the precipitin response to SACHO, the qualitative expression of anti-SACHO antibodies with restricted heterogeneity was more frequently observed in the August strain of rats than in the six other inbred strains examined. Cross-idiotypic specificity was demonstrated for anti-SACHO antisera obtained from nine inbred rat strains. The observations on idiotypy favor the importance of germ-line genes coding for rat antibody variable region determinants in response to SACHO.In this paper, the following abbreviations are used SACHO streptococcal group A carbohydrate - Aug August 2887 - W/Fu Wistar Furth - M520 Marshall 520 - Cop Copenhagen - F344 Fisher 344 - Buf Buffalo/Cr - BN Brown Norway - GASV group A streptococcal vaccine - DEAE diethylaminoethyl-cellulose - RIA radioimmunoassay     

Genetic linkage between immune response to GAT and the fate of RSV-induced tumors in chickens

Recent studies suggest that the gene locus controlling the fate of tumors induced by Rous sarcoma virus (RSV) is linked to theB histocompatibility complex. Birds carrying the dominant allele regress the tumor; homozygous recessives being unable to do so, develop large tumors and die. These are called progressors.The Bryan strain of RSV was inoculated into 220 6 week old Leghorns homozygous forB ¹ B ¹,B ² B ², orB ¹⁹ B ¹⁹ of which the percentages of progressors were 79, 22 and 56, respectively. The balance of each were regressors and survived.TheB ¹ B ¹ test birds were derived from special matings, i.e., high and low immune responders to the amino acid polymer, GAT. Of 67 tests progeny of theB ¹ B ¹ GAT-low mating, 63 or 94% proved to be progressors, and 6% were regressors. Of 84 test progeny of theB ¹ B ¹ GAT-high matings, 67% were progressors, and 33% were regressors. The difference between the high and low GAT responders is highly significant and indicates that the locus controlling the fate of RSV-induced tumors is closely linked to the locus controlling immune response to GAT. The latter maps within theIr region of theB histocompatibility complex.     

Genetic influences on spontaneous autoimmune thyroiditis in (CS x OS)F2 chickens

Genetic effects on spontaneous autoimmune thyroiditis in chickens were assessed by measuring phenotypic symptoms, the titer of circulating antibody to thyroglobulin, and the pathological change in the thyroids of young chicks. One or more loci within theB complex (the major histocompatibility complex of the chicken) are responsible for the expression of autoimmunity, and evidence is provided for an interaction of theB haplotype with genes at other loci. The influence of theB complex component on genetic susceptibility is more visible in animals with limited susceptibility at other loci and becomes indistinguishable as the frequency of other genes determining thyroid autoimmunity increases.     

Preliminary evidence of genetic control of the immune response to the Thy-1.2 antigen in mice

The primary immune response to the Thy-1.2 antigen was measured by the plaque assay, detecting cells (PFC) producing antibodies lytic for thymocytes carrying this antigen. On the basis of statistically significant differences in response, the inbred strains studied could be classified as low (producing fewer than 10³ PFC/spleen) or high (producing more than 10³ PFC/spleen) responders. The data collected from these inbred strains and segregating generations were consistent with the concept that the primary immune response to the Thy-1.2 antigen is under genetic control. This control appeared to be exerted primarily by alleles at a locus linked to theH-2 complex but involvement of alleles at other loci could not be ruled out. Contrary to the commonly described experimental models, the high responsiveness to the Thy-1.2 antigen in some strain combinations seemed to be a recessive rather than a codominant trait.     

Sexual preference of meiotic recombination within the H-2 complex

The recombination frequency between the H-2K and H-2D marker loci in male mice was measured using heterozygotes that carry the H-2 ^wm7 haplotype derived from the Japanese wild mouse and common H-2 haplotypes derived from inbred mice. Previous mating experiments in which backcross progeny of heterozygous females were screened demonstrated that the H-2 ^2m7 displays marked enhancement of recombination within the H-2 complex. In contrast to recombination in female mice, no enhancement of recombination was observed during male meiosis in the present study. Thus, it appeared that enhancement of recombination is specific to female mice. A genealogical study of recombination indicated that the postmeiotic stage is not involved in the generation of sexual preference of enhancement of recombination, suggesting that the preference is meiotic-drive and that a female-specific mechanism is involved in meiotic recombination mediated by the H-2 ^wm7 haplotype.