Class II transplantation antigens are involved in physiological interactions between cells of var... more Class II transplantation antigens are involved in physiological interactions between cells of various types within the immune system [1]. Although class II antigens have been implied in several discrete immunological events, little is as yet known about their functions at the molecular level. This is at least partly due to the fact that class II antigens are relatively difficult to isolate and characterize. However , recombinant D N A techniques now offer new means to characterize the class II antigens, in regards to structure, genetic polymorphism, and, eventually, function. Using cloned cDNA, it has already been possible to gain some insight into the number of class II antigen loci, the distribution of variant amino acids among allelic forms of class II antigens, and the organization of archetypal genes for class II antigens. In this communication, we will briefly summarize some of our data on these points.
Summary. We have developed a semi-nested PCR assay which measures gene conversion in mouse MHC cl... more Summary. We have developed a semi-nested PCR assay which measures gene conversion in mouse MHC class genes. Detectable frequencies occur in DNA from sperm, but not in somatic DNA, such as liver or fibroblast DNA. The frequency of gene conversion events in MHC class II genes varies strongly from one MHC haplotype to another, with the highest detected frequencies as high as 1140 000 for an individual heterozygous for both donor and acceptor sequences. Deletions or insertions in one gene relative to the other seem to lower the efficiency of gene conversion considerably. However, not all the variation in gene conversion frequency can be readily ascribed to the MHC genes themselves, implying that variation in the non-MHC background genes also might be of importance. Stretches within MHC genes amenable to gene conversion are located in CpG clusters, whereas MHC genes not involved in gene conversion have background CpG levels. This feature extends to some, but not all, of the non-MHC genes that have been reported to undergo gene conversion in mammals. DNA damage, either chern ical or radiation-induced, increases the frequency of gene conversion of MHC class II genes in cultured cells of the fibroblastoid lineage. The effect of chemical DNA damage seems roughly dose-dependent, whereas irradiation has maximal effect at low doses.
Mutation Research: Fundamental And Molecular Mechanisms Of Mutagenesis, 1999
We have explored the effects of γ-irradiation and etoposide on the gene conversion frequency betw... more We have explored the effects of γ-irradiation and etoposide on the gene conversion frequency between the endogenous major histocompatibility complex class II genes Abk and Ebd in a mouse testis cell line of non-germline origin with a polymerase chain reaction assay. Both γ-rays and etoposide were shown to increase the gene conversion frequency with up to 15-fold compared to untreated
Class II genes of the bovine major histocompatibility complex have been investigated by Southern ... more Class II genes of the bovine major histocompatibility complex have been investigated by Southern blot analysis using human cDNA probes for DQ alpha, DQ beta, DR alpha and DR beta. In this report restriction fragment length polymorphisms of DR alpha and DR beta are described. The polymorphisms were interpreted genetically by analysing five paternal half-sib families of the Swedish Red and White Breed, comprising altogether 28 offspring. Using the restriction enzymes BamHI, EcoRI and PvuII, three DR alpha and three DR beta allelic fragment patterns were resolved. The DR alpha and DR beta genes thus appear to be much less polymorphic than the previously described DQ alpha and DQ beta genes. Also, the observed linkage disequilibrium between DR genes was less pronounced than that between DQ genes, whereas the association between DR and DQ haplotypes was very strong. The family data available indicated strongly that the DQ alpha, DQ beta, DR alpha and DR beta genes are all closely linked.
The I-E molecule of the major histocompatibility complex (MHC) can prevent the spontaneous develo... more The I-E molecule of the major histocompatibility complex (MHC) can prevent the spontaneous development of diabetes in nonobese diabetic (NOD) mice. The mechanism of this protection has been investigated by breeding wild-type and promoter-mutated E kappa alpha transgenes onto the NOD genetic background. Animals carrying the various mutated transgenes expressed I-E on different subsets of immunocompetent cells, and thus cells important for the I-E protective effect could be identified. Although the wild-type transgene prevented the infiltration of lymphocytes into pancreatic islets, none of the mutants did. However, all of the transgenes could mediate the intrathymic elimination of T cells bearing antigen receptors with variable regions that recognize I-E. Thus, the I-E molecule does not protect NOD mice from diabetes simply by inducing the deletion of self-reactive T cells.
The nonobese diabetic (NOD) mouse is a useful model for human autoimmune diabetes. The gene for t... more The nonobese diabetic (NOD) mouse is a useful model for human autoimmune diabetes. The gene for the anti-apoptotic protein Bcl-2 has previously been suggested as a probable susceptibility candidate for the NOD mouse disease. In this study, we investigated how overexpression of Bcl-2 in lymphocytes might affect insulitis in NOD mice. A bcl-2 transgene expressed constitutively under the SV40-promoter and the 5'Igh enhancer, Emu, was bred onto NOD background. Two bcl-2 transgenic NOD strains were produced and analysed, one with overexpression of Bcl-2 on only B cells and the other with overexpression of Bcl-2 on both B and T cells. Subsequent to verification of expression pattern and functionality of the transgene, insulitis intensity was investigated in different backcross generations of the two transgenic strains. Overexpression of Bcl-2 on both B and T cells leads to a statistically significant protection of the mice from insulitis compared with normal littermates. Overexpression of Bcl-2 on only B cells, on the other hand, does not have any statistically significant effect on insulitis. Possible mechanisms for the effect of Bcl-2 on insulitis in NOD mice are discussed.
The NOD mouse is an animal model for insulin-dependent diabetes with many similarities to the hum... more The NOD mouse is an animal model for insulin-dependent diabetes with many similarities to the human disease. NOD mice which are transgenic for the Ea gene, allowing expression of the E molecule, are protected from diabetes and rarely develop insulitis. We have constructed bone marrow chimeras between transgenic and non-transgenic NOD mice to study the correlation of E expression on bone marrow derived cells and thymic epithelium vs the production of IL-4 and IFN-gamma. We show that NOD-E-->NOD-E and NOD-E-->NOD chimeras have elevated levels of IL-4 compared to NOD-->NOD and NOD-->NOD-E chimeras in the thymus. However, in the periphery the protected NOD-E-->NOD-E show much higher IL-4 levels than any of the other chimeras. This drop in peripheral IL-4 production seen in NOD-E-->NOD, NOD-->NOD-E and NOD-->NOD chimeras correlates with the increased insulitis seen in these mice compared to NOD-E-->NOD-E. In contrast, there were no differences in IFN-gamma production between the chimeras. We suggest that the precommitted, regulatory T cells, selected in an E-expressing thymic environment, need continuous interaction with E-expressing primary antigen presenting cells in the periphery for optimal IL-4 production. Decrease in IL-4 production correlates with increased insulitis.
Probes isolated from DR and DQ beta cDNA and genomic clones were used in hybridizations to restri... more Probes isolated from DR and DQ beta cDNA and genomic clones were used in hybridizations to restriction enzyme-digested DNA from human homozygous typing cells (HTC) as well as other DR homozygous cells in order to estimate the number of beta genes in the DR/DQ class II region. Varying numbers of DR beta genes were found in HTC of different DR specificities, from possibly one in DR 8 cells to three in cells of DR 2 to 7. The DR beta genes of different specificities seem to be related to one another in a distinct fashion. In contrast, all HTC contain two DQ beta genes per chromosome. The restriction site polymorphism of DQ beta genes is considerably more extensive than that of DQ serology, although one of the genes seems to be nonpolymorphic. In addition to the two DP beta genes identified previously, a minimum of three to five DQ and DR beta genes exist in the human haploid genome.
Class II transplantation antigens are involved in physiological interactions between cells of var... more Class II transplantation antigens are involved in physiological interactions between cells of various types within the immune system [1]. Although class II antigens have been implied in several discrete immunological events, little is as yet known about their functions at the molecular level. This is at least partly due to the fact that class II antigens are relatively difficult to isolate and characterize. However , recombinant D N A techniques now offer new means to characterize the class II antigens, in regards to structure, genetic polymorphism, and, eventually, function. Using cloned cDNA, it has already been possible to gain some insight into the number of class II antigen loci, the distribution of variant amino acids among allelic forms of class II antigens, and the organization of archetypal genes for class II antigens. In this communication, we will briefly summarize some of our data on these points.
Summary. We have developed a semi-nested PCR assay which measures gene conversion in mouse MHC cl... more Summary. We have developed a semi-nested PCR assay which measures gene conversion in mouse MHC class genes. Detectable frequencies occur in DNA from sperm, but not in somatic DNA, such as liver or fibroblast DNA. The frequency of gene conversion events in MHC class II genes varies strongly from one MHC haplotype to another, with the highest detected frequencies as high as 1140 000 for an individual heterozygous for both donor and acceptor sequences. Deletions or insertions in one gene relative to the other seem to lower the efficiency of gene conversion considerably. However, not all the variation in gene conversion frequency can be readily ascribed to the MHC genes themselves, implying that variation in the non-MHC background genes also might be of importance. Stretches within MHC genes amenable to gene conversion are located in CpG clusters, whereas MHC genes not involved in gene conversion have background CpG levels. This feature extends to some, but not all, of the non-MHC genes that have been reported to undergo gene conversion in mammals. DNA damage, either chern ical or radiation-induced, increases the frequency of gene conversion of MHC class II genes in cultured cells of the fibroblastoid lineage. The effect of chemical DNA damage seems roughly dose-dependent, whereas irradiation has maximal effect at low doses.
Mutation Research: Fundamental And Molecular Mechanisms Of Mutagenesis, 1999
We have explored the effects of γ-irradiation and etoposide on the gene conversion frequency betw... more We have explored the effects of γ-irradiation and etoposide on the gene conversion frequency between the endogenous major histocompatibility complex class II genes Abk and Ebd in a mouse testis cell line of non-germline origin with a polymerase chain reaction assay. Both γ-rays and etoposide were shown to increase the gene conversion frequency with up to 15-fold compared to untreated
Class II genes of the bovine major histocompatibility complex have been investigated by Southern ... more Class II genes of the bovine major histocompatibility complex have been investigated by Southern blot analysis using human cDNA probes for DQ alpha, DQ beta, DR alpha and DR beta. In this report restriction fragment length polymorphisms of DR alpha and DR beta are described. The polymorphisms were interpreted genetically by analysing five paternal half-sib families of the Swedish Red and White Breed, comprising altogether 28 offspring. Using the restriction enzymes BamHI, EcoRI and PvuII, three DR alpha and three DR beta allelic fragment patterns were resolved. The DR alpha and DR beta genes thus appear to be much less polymorphic than the previously described DQ alpha and DQ beta genes. Also, the observed linkage disequilibrium between DR genes was less pronounced than that between DQ genes, whereas the association between DR and DQ haplotypes was very strong. The family data available indicated strongly that the DQ alpha, DQ beta, DR alpha and DR beta genes are all closely linked.
The I-E molecule of the major histocompatibility complex (MHC) can prevent the spontaneous develo... more The I-E molecule of the major histocompatibility complex (MHC) can prevent the spontaneous development of diabetes in nonobese diabetic (NOD) mice. The mechanism of this protection has been investigated by breeding wild-type and promoter-mutated E kappa alpha transgenes onto the NOD genetic background. Animals carrying the various mutated transgenes expressed I-E on different subsets of immunocompetent cells, and thus cells important for the I-E protective effect could be identified. Although the wild-type transgene prevented the infiltration of lymphocytes into pancreatic islets, none of the mutants did. However, all of the transgenes could mediate the intrathymic elimination of T cells bearing antigen receptors with variable regions that recognize I-E. Thus, the I-E molecule does not protect NOD mice from diabetes simply by inducing the deletion of self-reactive T cells.
The nonobese diabetic (NOD) mouse is a useful model for human autoimmune diabetes. The gene for t... more The nonobese diabetic (NOD) mouse is a useful model for human autoimmune diabetes. The gene for the anti-apoptotic protein Bcl-2 has previously been suggested as a probable susceptibility candidate for the NOD mouse disease. In this study, we investigated how overexpression of Bcl-2 in lymphocytes might affect insulitis in NOD mice. A bcl-2 transgene expressed constitutively under the SV40-promoter and the 5'Igh enhancer, Emu, was bred onto NOD background. Two bcl-2 transgenic NOD strains were produced and analysed, one with overexpression of Bcl-2 on only B cells and the other with overexpression of Bcl-2 on both B and T cells. Subsequent to verification of expression pattern and functionality of the transgene, insulitis intensity was investigated in different backcross generations of the two transgenic strains. Overexpression of Bcl-2 on both B and T cells leads to a statistically significant protection of the mice from insulitis compared with normal littermates. Overexpression of Bcl-2 on only B cells, on the other hand, does not have any statistically significant effect on insulitis. Possible mechanisms for the effect of Bcl-2 on insulitis in NOD mice are discussed.
The NOD mouse is an animal model for insulin-dependent diabetes with many similarities to the hum... more The NOD mouse is an animal model for insulin-dependent diabetes with many similarities to the human disease. NOD mice which are transgenic for the Ea gene, allowing expression of the E molecule, are protected from diabetes and rarely develop insulitis. We have constructed bone marrow chimeras between transgenic and non-transgenic NOD mice to study the correlation of E expression on bone marrow derived cells and thymic epithelium vs the production of IL-4 and IFN-gamma. We show that NOD-E-->NOD-E and NOD-E-->NOD chimeras have elevated levels of IL-4 compared to NOD-->NOD and NOD-->NOD-E chimeras in the thymus. However, in the periphery the protected NOD-E-->NOD-E show much higher IL-4 levels than any of the other chimeras. This drop in peripheral IL-4 production seen in NOD-E-->NOD, NOD-->NOD-E and NOD-->NOD chimeras correlates with the increased insulitis seen in these mice compared to NOD-E-->NOD-E. In contrast, there were no differences in IFN-gamma production between the chimeras. We suggest that the precommitted, regulatory T cells, selected in an E-expressing thymic environment, need continuous interaction with E-expressing primary antigen presenting cells in the periphery for optimal IL-4 production. Decrease in IL-4 production correlates with increased insulitis.
Probes isolated from DR and DQ beta cDNA and genomic clones were used in hybridizations to restri... more Probes isolated from DR and DQ beta cDNA and genomic clones were used in hybridizations to restriction enzyme-digested DNA from human homozygous typing cells (HTC) as well as other DR homozygous cells in order to estimate the number of beta genes in the DR/DQ class II region. Varying numbers of DR beta genes were found in HTC of different DR specificities, from possibly one in DR 8 cells to three in cells of DR 2 to 7. The DR beta genes of different specificities seem to be related to one another in a distinct fashion. In contrast, all HTC contain two DQ beta genes per chromosome. The restriction site polymorphism of DQ beta genes is considerably more extensive than that of DQ serology, although one of the genes seems to be nonpolymorphic. In addition to the two DP beta genes identified previously, a minimum of three to five DQ and DR beta genes exist in the human haploid genome.
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