New loci regulating rat myelin oligodendrocyte glycoprotein-induced experimental autoimmune encephalomyelitis

Myelin oligodendrocyte glycoprotein-induced experimental autoimmune encephalomyelitis (EAE) is an inflammatory disease in rats that closely mimics many clinical and histopathological aspects of multiple sclerosis. Non-MHC quantitative trait loci regulating myelin oligodendrocyte glycoprotein-induced EAE have previously been identified in the EAE-permissive strain, DA, on rat chromosomes 4, 10, 15, and 18. To find any additional gene loci in another well-known EAE-permissive strain and thereby to assess any genetic heterogeneity in the regulation of the disease, we have performed a genome-wide linkage analysis in a reciprocal (LEW.1AV1 x PVG.1AV1) male/female F(2) population (n = 185). We examined reciprocal crosses, but no parent-of-origin effect was detected. The parental rat strains share the RT1(av1) MHC haplotype; thus, non-MHC genes control differences in EAE susceptibility. We identified Eae16 on chromosome 8 and Eae17 on chromosome 13, significantly linked to EAE phenotypes. Two loci, on chromosomes 1 and 17, respectively showed suggestive linkage to clinical and histopathological EAE phenotypes. Eae16 and Eae17 differ from those found in previously studied strain combinations, thus demonstrating genetic heterogeneity of EAE. Furthermore, we detected a locus-specific parent-of-origin effect with suggestive linkage in Eae17. Further genetic and functional dissection of these loci may disclose critical disease-regulating molecular mechanisms.     

Novel quantitative trait loci controlling development of experimental autoimmune encephalomyelitis and proportion of lymphocyte subpopulations

The B10.RIII mouse strain (H-2(r)) develops chronic experimental autoimmune encephalomyelitis (EAE) upon immunization with the myelin basic protein 89-101 peptide. EAE was induced and studied in a backcross between B10.RIII and the EAE-resistant RIIIS/J strain (H-2(r)), and a complete genome scan with microsatellite markers was performed. Five loci were significantly linked to different traits and clinical subtypes of EAE on chromosomes 1, 5, 11, 15, and 16, three of the loci having sex specificity. The quantitative trait locus on chromosome 15 partly overlapped with the Eae2 locus, previously identified in crosses between the B10.RIII and RIIIS/J mouse strains. The loci on chromosomes 11 and 16 overlapped with Eae loci identified in other mouse crosses. By analyzing the backcross animals for lymphocyte phenotypes, the proportion of B and T cells in addition to the levels of CD4(+)CD8(-) and CD4(-)CD8(+) T cells and the CD4(+)/CD8(+) ratio in spleen were linked to different loci on chromosomes 1, 2, 3, 5, 6, 11, and 15. On chromosome 16, we found significant linkage to spleen cell proliferation. Several linkages overlapped with the quantitative trait loci for disease phenotypes. The identification of subphenotypes that are linked to the same loci as disease traits could be most useful in the search for candidate genes and biological pathways involved in the pathological process.     

Detection of loci for allergic asthma using SMXA recombinant inbred strains of mice

Asthma is regarded as a multifactorial inflammatory disorder arising as a result of inappropriate immune responses in genetically susceptible individuals to common environmental antigens. However, the precise molecular basis is unknown. To identify genes for susceptibility to three asthma-related traits, airway hyperresponsiveness (AHR), eosinophil infiltration, and allergen-specific serum IgE levels, we conducted a genetic analysis using SMXA recombinant inbred (RI) strains of mice. Quantitative trait locus analysis detected a significant locus for AHR on chromosome 17. For eosinophil infiltration, significant loci were detected on chromosomes 9 and 16. Although we could not detect any significant loci for allergen-specific serum IgE, analysis of consomic strains showed that chromosomes 17 and 19 carried genes that affected this trait. We detected genetic susceptibility loci that separately regulated the three asthma-related phenotypes. Our results suggested that different genetic mechanisms regulate these asthma-related phenotypes. Genetic analyses using murine RI and consomic strains enhance understanding of the molecular mechanisms of asthma in human.     

A wild-derived inbred mouse strain,MSM/Ms,provides insights into novel skin tumor susceptibility genes

Cancer is one of the most catastrophic human genetic diseases. Experimental animal cancer models are essential for gaining insights into the complex interactions of different cells and genes in tumor initiation, promotion, and progression. Mouse models have been extensively used to analyze the genetic basis of cancer susceptibility. They have led to the identification of multiple loci that confer, either alone or in specific combinations, an increased susceptibility to cancer, some of which have direct translatability to human cancer. Additionally, wild-derived inbred mouse strains are an advantageous reservoir of novel genetic polymorphisms of cancer susceptibility genes, because of the evolutionary divergence between wild and classical inbred strains. Here, we review mapped Stmm (skin tumor modifier of MSM) loci using a Japanese wild-derived inbred mouse strain, MSM/Ms, and describe recent advances in our knowledge of the genes responsible for Stmm loci in the 7,12-dimethylbenz(a)anthracene (DMBA)/12-O-tetradecanoylphorbol-13-acetate (TPA) two-stage skin carcinogenesis model.     

Systematic analysis of emotionality in consomic mouse strains established from C57BL/6J and wild-derived MSM/Ms

Consomic strains have recently attracted attention as an advantageous method to screen for genes related to developmental, physiological, and behavioral phenotypes. Recently, a new set of consomic strains was established from the Japanese wild-derived mouse strain MSM/Ms and C57BL/6JJcl. By analyzing the entire consomic panel, we were able to identify a number of chromosomes associated with anxiety-like behaviors in the open-field (OF) test, a light-dark box and an elevated plus maze. Detailed observation of the OF behavior allowed us to identify chromosomes associated with those ethological traits, such as stretch attend, rearing, and jumping. Repeated OF test trials have different meanings for animals, and we found that some chromosomes responded to only the first or second trial, while others were consistent across both trials. By examining both male and female mice, sex-dependent effects were found in several measurements. Principal component analysis of anxiety-like behaviors extracted five factors: 'general locomotor activity', 'thigmotaxis', 'risk assessment', 'open-arm exploration' and 'autonomic emotionality'. We mapped chromosomes associated with these five factors of emotionality.     

Gene-Expression Profiling of Experimental Autoimmune Encephalomyelitis

Experimental autoimmune encephalomyelitis (EAE) is a mouse model that serves as an experimental tool for studying the etiology, pathogenesis, as well as new therapeutic approaches of multiple sclerosis (MS). EAE is a polygenic chronic inflammatory demyelinating disease of the nervous system that involves the interaction between genetic and environmental factors. Previous studies have identified multiple quantitative trait loci (QTL) controlling different aspects of disease pathogenesis. However, progress in identifying new susceptibility genes outside the MHC locus has been slow. With the advent of new global methods for genetic analysis such as large-scale sequencing, gene expression profiling combined with classic linkage analysis and congenic and physical mapping progress is considerably accelerating. Here we review our preliminary work on the use of gene expression mapping to identify new putative genetic pathways contributing to the pathogenesis of EAE.     

Establishment of consomic strains derived from A/J and SM/J mice for genetic analysis of complex traits

Consomic strains, in which one chromosome is derived from a donor strain and the other chromosomes are derived from the recipient strain, provide a powerful tool for the dissection of complex genetic traits. In this study we established ten consomic strains (A-2^SM, A-6^SM, A-11^SM, A-12^SM, A-13^SM, A-15^SM, A-17^SM, A-18^SM, A-19^SM, A-Y^SM) using the SM/J strain as the donor and the A/J strain as the recipient; these are the parental strains of a set of SMXA recombinant inbred (RI) strains that we had developed previously. We analyzed body weights and blood lipid levels in the consomic and parental strains. The mean values for each trait showed a continuous range of variation in the consomic strains suggesting that they are controlled by multiple genes. We previously identified suggestive QTLs for body weight on chromosome 6 in SMXA RI strains and (SM/J?×?A/J)F₂ mice. The observation that the A-6^SM consomic strain had a significantly lower mean body weight than the A/J strain supports the presence of this QTL on chromosome 6. Similarly, the higher blood triglyceride level in the A-11^SM strain shows the existence of a previously mapped QTL on chromosome 11, and the A-12^SM strain provides evidence of a QTL for blood total cholesterol level on chromosome 12. These consomic strains, along with the previously developed set of SMXA RI strains from A/J and SM/J mice, offer an invaluable and powerful resource for the analysis of complex genetic traits in mice.     

Segregation of a QTL cluster for home-cage activity using a new mapping method based on regression analysis of congenic mouse strains

Recent genetic studies have shown that genetic loci with significant effects in whole-genome quantitative trait loci (QTL) analyses were lost or weakened in congenic strains. Characterisation of the genetic basis of this attenuated QTL effect is important to our understanding of the genetic mechanisms of complex traits. We previously found that a consomic strain, B6-Chr6C^MSM, which carries chromosome 6 of a wild-derived strain MSM/Ms on the genetic background of C57BL/6J, exhibited lower home-cage activity than C57BL/6J. In the present study, we conducted a composite interval QTL analysis using the F2 mice derived from a cross between C57BL/6J and B6-Chr6C^MSM. We found one QTL peak that spans 17.6 Mbp of chromosome 6. A subconsomic strain that covers the entire QTL region also showed lower home-cage activity at the same level as the consomic strain. We developed 15 congenic strains, each of which carries a shorter MSM/Ms-derived chromosomal segment from the subconsomic strain. Given that the results of home-cage activity tests on the congenic strains cannot be explained by a simple single-gene model, we applied regression analysis to segregate the multiple genetic loci. The results revealed three loci (loci 1–3) that have the effect of reducing home-cage activity and one locus (locus 4) that increases activity. We also found that the combination of loci 3 and 4 cancels out the effects of the congenic strains, which indicates the existence of a genetic mechanism related to the loss of QTLs.     

Genetic analysis of disease subtypes and sexual dimorphisms in mouse experimental allergic encephalomyelitis (EAE): relapsing/remitting and monophasic remitting/nonrelapsing EAE are immunogenetically distinct

Experimental allergic encephalomyelitis (EAE) is the principal animal model of multiple sclerosis (MS), the major inflammatory disease of the central nervous system. Murine EAE is generally either an acute monophasic or relapsing disease. Because the clinical spectrum of MS is more diverse, the limited range of disease subtypes observed in EAE has raised concern regarding its relevance as a model for MS. During the generation of a large F2 mapping population between the EAE-susceptible SJL/J and EAE-resistant B10.S/DvTe inbred lines, we identified four distinct subtypes of murine EAE resembling clinical subtypes seen in MS. We observed acute progressive, chronic/nonremitting, remitting/relapsing, and monophasic remitting/nonrelapsing EAE. An additional subtype, benign EAE, was identified after histologic examination revealed that some mice had inflammatory infiltrates of the central nervous system, but did not show clinical signs of EAE. Genome exclusion mapping was performed to identify the loci controlling susceptibility to each disease subtype. We report three novel EAE-modifying loci on chromosomes 16, 7, and 13 (eae11-13, respectively). Additionally, unique loci with gender-specific effects govern susceptibility to remitting/relapsing (eae12) and monophasic remitting/nonrelapsing (eae7 and 13) EAE.     

Two loci on chromosome 15 control experimentally induced arthritis through the differential regulation of IL-6 and lymphocyte proliferation

Using genetic linkage analysis of proteoglycan-induced arthritis (PGIA), a murine model for rheumatoid arthritis, we identified two loci, Pgia8 and Pgia9, on chromosome 15 (chr15) that appear to be implicated in disease susceptibility. Immunization of congenic strains carrying the entire chr15 and separately each of the two loci of DBA/2 arthritis-resistant origin in susceptible BALB/c background confirmed locations of two loci on chr15: the major Pgia9 and lesser Pgia8 locus. Distal part of chr15 (Pgia9) showed a major suppressive effect on PGIA susceptibility in females (40%, p < 0.001), whereas the effect of this locus in congenic males was still significant but weaker. Proximal part of chr15 (Pgia8) demonstrated mild and transient effect upon arthritis; this effect was PGIA-promoting in males and suppressive in females. Pgia8 and Pgia9 loci demonstrated an additive mode of inheritance, since when they were both incorporated in consomic chr15 strain, the total effect was a sum of the two loci. Using F(2) population of the intercross of wild-type and chr15 consomic strain, we confirmed and refined quantitative trait locus positions and identified a strong correlation between disease susceptibility and lymphocyte-producing cytokines of TNF-alpha and IL-6. Both Pgia8 and Pgia9 loci on chr15 appear to control IL-6 production in spleen cultures of arthritic mice, providing an important link to the mechanism of autoimmune inflammation.     

Establishment of consomic strains derived from A/J and SM/J mice for genetic analysis of complex traits

Consomic strains, in which one chromosome is derived from a donor strain and the other chromosomes are derived from the recipient strain, provide a powerful tool for the dissection of complex genetic traits. In this study we established ten consomic strains (A-2^SM, A-6^SM, A-11^SM, A-12^SM, A-13^SM, A-15^SM, A-17^SM, A-18^SM, A-19^SM, A-Y^SM) using the SM/J strain as the donor and the A/J strain as the recipient; these are the parental strains of a set of SMXA recombinant inbred (RI) strains that we had developed previously. We analyzed body weights and blood lipid levels in the consomic and parental strains. The mean values for each trait showed a continuous range of variation in the consomic strains suggesting that they are controlled by multiple genes. We previously identified suggestive QTLs for body weight on chromosome 6 in SMXA RI strains and (SM/J × A/J)F₂ mice. The observation that the A-6^SM consomic strain had a significantly lower mean body weight than the A/J strain supports the presence of this QTL on chromosome 6. Similarly, the higher blood triglyceride level in the A-11^SM strain shows the existence of a previously mapped QTL on chromosome 11, and the A-12^SM strain provides evidence of a QTL for blood total cholesterol level on chromosome 12. These consomic strains, along with the previously developed set of SMXA RI strains from A/J and SM/J mice, offer an invaluable and powerful resource for the analysis of complex genetic traits in mice.

     

Sex-specific differences in chromosome-dependent regulation of vascular reactivity in female consomic rat strains from a SSxBN cross

High-throughput studies in the Medical College of Wisconsin Program for Genomic Applications (Physgen) were designed to link chromosomes with physiological function in consomic strains derived from a cross between Dahl salt-sensitive SS/JrHsdMcwi (SS) and Brown Norway normotensive BN/NHsdMcwi (BN) rats. The specific goal of the vascular protocol was to characterize the responses of aortic rings from these strains to vasoconstrictor and vasodilator stimuli (phenylephrine, acetylcholine, sodium nitroprusside, and bath hypoxia) to identify chromosomes that either increase or decrease vascular reactivity to these vasoactive stimuli. Because previous studies demonstrated sex-specific quantitative trait loci (QTLs) related to regulation of cardiovascular phenotypes in an F2 cross between the parental strains, males and females of each consomic strain were included in all experiments. As there were significant sex-specific differences in aortic sensitivity to vasoconstrictor and vasodilator stimuli compared with the parental SS strain, we report the results of the females separately from the males. There were also sex-specific differences in aortic ring sensitivity to these vasoactive stimuli in consomic strains that were fed a high-salt diet (4% NaCl) for 3 wk to evaluate salt-induced changes in vascular reactivity. Differences in genetic architecture could contribute to sex-specific differences in the development and expression of cardiovascular diseases via differential regulation and expression of genes. Our findings are the first to link physiological traits with specific chromosomes in female SS rats and support the idea that sex is an important environmental variable that plays a role in the expression and regulation of genes.     

QTL analysis of measures of mouse home-cage activity using B6/MSM consomic strains

The activity of mice in their home cage is influenced greatly by the cycle of light and dark. In addition, home-cage activity shows remarkable time-dependent changes that result in a prominent temporal pattern. The wild-derived mouse strain MSM/Ms (MSM) exhibits higher total activity in the home cage than does C57BL/6 (B6), a commonly used laboratory strain. In addition, there is a clear strain difference in the temporal pattern of home-cage activity. This study aimed to clarify the genetic basis of strain differences in the temporal pattern of home-cage activity between MSM and B6. Through the comparison of temporal patterns of home-cage activity between B6 and MSM, the pattern can be classified into five temporal components: (1) resting phase, (2) anticipation phase, (3) 1st phase, (4) 2nd phase, and (5) 3rd phase. To identify quantitative trait loci (QTLs) involved in these temporal components, we used consomic strains established from crosses between B6 and MSM. Five consomic strains, for Chrs 2T (telomere), 3, 4, 13, and 14, showed significantly higher total activity than B6. In contrast, the consomic strains of Chrs 6C (centromere), 7T, 9, 11, and 15 were less active than B6. This indicates that multigenic factors regulate the total activity. Further analysis showed an impact of QTLs on the temporal components of home-cage activity. The present data showed that each temporal component was regulated by different combinations of multigenic factors, with some overlap. These temporal component-related QTLs are important to understand fully the genetic mechanisms that underlie home-cage activity.     

Generation and mapping of Mus spretus strain-specific markers for rapid genomic scanning

We describe here a set of genetic markers, based on IRS–PCR amplification difference, that are specifically designed for efficient, high throughput genetic mapping in [(M. domesticus× wild-derived) F₁×M. domesticus] interspecific backcrosses. 146 new genetic loci have been mapped, and strain distribution for these markers has been determined in 96 mouse strains. 103 (81%) of 127 tested markers are present only in one or more wild-derived strains, but absent in 76 other commonly used strains, demonstrating their utility in a variety of mouse pair combinations. Because of the ease of genotyping with this marker set, rapid genome scans for complex genetic trait loci involving crosses between wild-derived strains and other commonly used strains can now be carried out efficiently with large numbers of animals. Received: 2 October 1995 / Accepted: 12 December 1995     

Fine haplotype structure of a chromosome 17 region in the laboratory and wild mouse

Extensive linkage disequilibrium among classical laboratory strains represents an obstacle in the high-resolution haplotype mapping of mouse quantitative trait loci (QTL). To determine the potential of wild-derived mouse strains for fine QTL mapping, we constructed a haplotype map of a 250-kb region of the t-complex on chromosome 17 containing the Hybrid sterility 1 (Hst1) gene. We resequenced 33 loci from up to 80 chromosomes of five mouse (sub)species. Trans-species single-nucleotide polymorphisms (SNPs) were rare between Mus m. musculus (Mmmu) and Mus m. domesticus (Mmd). The haplotypes in Mmmu and Mmd differed and therefore strains from these subspecies should not be combined for haplotype-associated mapping. The haplotypes of t-chromosomes differed from all non-t Mmmu and Mmd haplotypes. Half of the SNPs and SN indels but only one of seven longer rearrangements found in classical laboratory strains were useful for haplotype mapping in the wild-derived M. m. domesticus. The largest Mmd haplotype block contained three genes of a highly conserved synteny. The lengths of the haplotype blocks deduced from 36 domesticus chromosomes were in tens of kilobases, suggesting that the wild-derived Mmd strains are suitable for fine interval-specific mapping.     

Marker-assisted congenic screening (MACS): A database tool for the efficient production and characterization of congenic lines

Over the past decades, genetic studies in rodent models of human multifactorial disorders have led to the detection of numerous chromosomal regions associated with disease phenotypes. Owing to the complex control of these phenotypes and the size of the disease loci, identifying the underlying genes requires further analyses in new original models, including chromosome substitution (consomic) and congenic lines, derived to evaluate the phenotypic effects of disease susceptibility loci and fine-map the disease genes. We have developed a relational database (MACS) specifically designed for the genetic marker-assisted production of large series of rodent consomic and congenic lines (speed congenics), the organization of their genetic and phenotypic characterizations, and the acquisition and archiving of both genetic and phenotypic data. This database, originally optimized for the production of rat congenics, can also be applied to mouse mapping projects. MACS represents an essential system for significantly improving efficiency and accuracy in investigations of multiple consomic and congenic lines simultaneously derived for different disease loci, and ultimately cloning genes underlying complex phenotypes.     

Unique genetic variation revealed by a microsatellite polymorphism survey in ten wild-derived inbred strains

Here we report on a genome polymorphism survey using 254 microsatellite markers in ten recently wild-derived inbred strains. Allele size analysis showed that the rate of polymorphism of these wild-derived mouse strains when compared with any of the common laboratory strains is on average 79.8%. We found 632 wild-derived alleles that were not present in the common laboratory strains, representing a 61% increase over the genetic variation observed in the laboratory strains. We also found that on average 14.5% of the microsatellite alleles of any given wild-derived inbred strain were unique. Our results indicate that the recently wild-derived mouse strains represent repositories of unique naturally occurring genetic variability and may prove invaluable for the study of complex phenotypes and in the construction of new mouse models of human disease.     

Parent-of-Origin Effects Implicate Epigenetic Regulation of Experimental Autoimmune Encephalomyelitis and Identify Imprinted Dlk1 as a Novel Risk Gene

Parent-of-origin effects comprise a range of genetic and epigenetic mechanisms of inheritance. Recently, detection of such effects implicated epigenetic mechanisms in the etiology of multiple sclerosis (MS), a chronic inflammatory disease of the central nervous system. We here sought to dissect the magnitude and the type of parent-of-origin effects in the pathogenesis of experimental neuroinflammation under controlled environmental conditions. We investigated inheritance of an MS-like disease in rat, experimental autoimmune encephalomyelitis (EAE), using a backcross strategy designed to identify the parental origin of disease-predisposing alleles. A striking 37–54% of all detected disease-predisposing loci depended on parental transmission. Additionally, the Y chromosome from the susceptible strain contributed to disease susceptibility. Accounting for parent-of-origin enabled more powerful and precise identification of novel risk factors and increased the disease variance explained by the identified factors by 2-4-fold. The majority of loci displayed an imprinting–like pattern whereby a gene expressed only from the maternal or paternal copy exerts an effect. In particular, a locus on chromosome 6 comprises a well-known cluster of imprinted genes including the paternally expressed Dlk1, an atypical Notch ligand. Disease-predisposing alleles at the locus conferred lower Dlk1 expression in rats and, together with data from transgenic overexpressing Dlk1 mice, demonstrate that reduced Dlk1 drives more severe disease and modulates adaptive immune reactions in EAE. Our findings suggest a significant epigenetic contribution to the etiology of EAE. Incorporating these effects enables more powerful and precise identification of novel risk factors with diagnostic and prognostic implications for complex disease.