Chromosome Substitution Strains Research Articles

Quantitative trait loci on chromosomes 9 and 19 modulate AII amacrine cell number in the mouse retina.

Sequence variants modulating gene function or expression affect various heritable traits, including the number of neurons within a population. The present study employed a forward-genetic approach to identify candidate causal genes and their sequence variants controlling the number of one type of retinal neuron, the AII amacrine cell. Data from twenty-six recombinant inbred (RI) strains of mice derived from the parental C57BL/6J (B6/J) and A/J laboratory strains were used to identify genomic loci regulating cell number. Large variation in cell number is present across the RI strains, from a low of ∼57,000 cells to a high of ∼87,000 cells. Quantitative trait locus (QTL) analysis revealed three prospective controlling genomic loci, on Chromosomes (Chrs) 9, 11, and 19, each contributing additive effects that together approach the range of variation observed. Composite interval mapping validated two of these loci, and chromosome substitution strains, in which the A/J genome for Chr 9 or 19 was introgressed on a B6/J genetic background, showed increased numbers of AII amacrine cells as predicted by those two QTL effects. Analysis of the respective genomic loci identified candidate controlling genes defined by their retinal expression, their established biological functions, and by the presence of sequence variants expected to modulate gene function or expression. Two candidate genes, Dtx4 on Chr 19, being a regulator of Notch signaling, and Dixdc1 on Chr 9, a modulator of the WNT-β-catenin signaling pathway, were explored in further detail. Postnatal overexpression of Dtx4 was found to reduce the frequency of amacrine cells, while Dixdc1 knockout retinas contained an excess of AII amacrine cells. Sequence variants in each gene were identified, being the likely sources of variation in gene expression, ultimately contributing to the final number of AII amacrine cells.

Linkage mapping reveals loci that underlie differences in Caenorhabditis elegans growth.

Growth rate and body size are complex traits that contribute to the fitness of organisms. The identification of loci that underlie differences in these traits provides insights into the genetic contributions to development. Leveraging Caenorhabditis elegans as a tractable metazoan model for quantitative genetics, we can identify genomic regions that underlie differences in growth. We measured postembryonic growth of the laboratory-adapted wild-type strain (N2) and a wild strain from Hawaii (CB4856) and found differences in body size. Using linkage mapping, we identified three distinct quantitative trait loci (QTL) on chromosomes IV, V, and X that are associated with variation in body growth. We further examined these growth-associated quantitative trait loci using chromosome substitution strains and near-isogenic lines and validated the chromosome X quantitative trait loci. In addition, we generated a list of candidate genes for the chromosome X quantitative trait loci. These genes could potentially contribute to differences in animal growth and should be evaluated in subsequent studies. Our work reveals the genetic architecture underlying animal growth variation and highlights the genetic complexity of growth in Caenorhabditis elegans natural populations.

Genetic Modifiers of Oral Nicotine Consumption in Chrna5 Null Mutant Mice.

The gene CHRNA5 is strongly associated with the level of nicotine consumption in humans and manipulation of the expression or function of Chrna5 similarly alters nicotine consumption in rodents. In both humans and rodents, reduced or complete loss of function of Chrna5 leads to increased nicotine consumption. However, the mechanism through which decreased function of Chrna5 increases nicotine intake is not well-understood. Toward a better understanding of how loss of function of Chrna5 increases nicotine consumption, we have initiated efforts to identify genetic modifiers of Chrna5 deletion-dependent oral nicotine consumption in mice. For this, we introgressed the Chrna5 knockout (KO) mutation onto a panel of C57BL/6J-Chr#A/J/NAJ chromosome substitution strains (CSS) and measured oral nicotine consumption in 18 CSS and C57BL/6 (B6) mice homozygous for the Chrna5 KO allele as well as their Chrna5 wild type littermates. As expected, nicotine consumption was significantly increased in Chrna5 KO mice relative to Chrna5 wildtype mice on a B6 background. Among the CSS homozygous for the Chrna5 KO allele, several exhibited altered nicotine consumption relative to B6 Chrna5 KO mice. Sex-independent modifiers were detected in CSS possessing A/J chromosomes 5 and 11 and a male-specific modifier was found on chromosome 15. In all cases nicotine consumption was reduced in the CSS Chrna5 KO mice relative to B6 Chrna5 KO mice and consumption in the CSS KO mice was indistinguishable from their wild type littermates. Nicotine consumption was also reduced in both Chrna5 KO and wildtype CSS mice possessing A/J chromosome 1 and increased in both KO and wild type chromosome 17 CSS relative to KO and wild type B6 mice. These results demonstrate the presence of several genetic modifiers of nicotine consumption in Chrna5 KO mice as well as identify loci that may affect nicotine consumption independent of Chrna5 genotype. Identification of the genes that underlie the altered nicotine consumption may provide novel insight into the mechanism through which Chrna5 deletion increases nicotine consumption and, more generally, a better appreciation of the neurobiology of nicotine intake.

PWD/Ph-Encoded Genetic Variants Modulate the Cellular Wnt/β-Catenin Response to Suppress Apc Min-Triggered Intestinal Tumor Formation.

Genetic predisposition affects the penetrance of tumor-initiating mutations, such as APC mutations that stabilize β-catenin and cause intestinal tumors in mice and humans. However, the mechanisms involved in genetically predisposed penetrance are not well understood. Here, we analyzed tumor multiplicity and gene expression in tumor-prone Apc Min/+ mice on highly variant C57BL/6J (B6) and PWD/Ph (PWD) genetic backgrounds. (B6 × PWD) F1 APC Min offspring mice were largely free of intestinal adenoma, and several chromosome substitution (consomic) strains carrying single PWD chromosomes on the B6 genetic background displayed reduced adenoma numbers. Multiple dosage-dependent modifier loci on PWD chromosome 5 each contributed to tumor suppression. Activation of β-catenin-driven and stem cell-specific gene expression in the presence of Apc Min or following APC loss remained moderate in intestines carrying PWD chromosome 5, suggesting that PWD variants restrict adenoma initiation by controlling stem cell homeostasis. Gene expression of modifier candidates and DNA methylation on chromosome 5 were predominantly cis controlled and largely reflected parental patterns, providing a genetic basis for inheritance of tumor susceptibility. Human SNP variants of several modifier candidates were depleted in colorectal cancer genomes, suggesting that similar mechanisms may also affect the penetrance of cancer driver mutations in humans. Overall, our analysis highlights the strong impact that multiple genetic variants acting in networks can exert on tumor development. SIGNIFICANCE: These findings in mice show that, in addition to accidental mutations, cancer risk is determined by networks of individual gene variants.

A Novel Mapping Strategy Utilizing Mouse Chromosome Substitution Strains Identifies Multiple Epistatic Interactions That Regulate Complex Traits.

The genetic contribution of additive vs. non-additive (epistatic) effects in the regulation of complex traits is unclear. While genome-wide association studies typically ignore gene-gene interactions, in part because of the lack of statistical power for detecting them, mouse chromosome substitution strains (CSSs) represent an alternate approach for detecting epistasis given their limited allelic variation. Therefore, we utilized CSSs to identify and map both additive and epistatic loci that regulate a range of hematologic- and metabolism-related traits, as well as hepatic gene expression. Quantitative trait loci (QTL) were identified using a CSS-based backcross strategy involving the segregation of variants on the A/J-derived substituted chromosomes 4 and 6 on an otherwise C57BL/6J genetic background. In the liver transcriptomes of offspring from this cross, we identified and mapped additive QTL regulating the hepatic expression of 768 genes, and epistatic QTL pairs for 519 genes. Similarly, we identified additive QTL for fat pad weight, platelets, and the percentage of granulocytes in blood, as well as epistatic QTL pairs controlling the percentage of lymphocytes in blood and red cell distribution width. The variance attributed to the epistatic QTL pairs was approximately equal to that of the additive QTL; however, the SNPs in the epistatic QTL pairs that accounted for the largest variances were undetected in our single locus association analyses. These findings highlight the need to account for epistasis in association studies, and more broadly demonstrate the importance of identifying genetic interactions to understand the complete genetic architecture of complex traits.

Chromosome-wide characterization of meiotic noncrossovers (gene conversions) in mouse hybrids.

During meiosis, the recombination-initiating DNA double-strand breaks (DSBs) are repaired by crossovers or noncrossovers (gene conversions). While crossovers are easily detectable, noncrossover identification is hampered by the small size of their converted tracts and the necessity of sequence polymorphism. We report identification and characterization of a mouse chromosome-wide set of noncrossovers by next-generation sequencing of 10 mouse intersubspecific chromosome substitution strains. Based on 94 identified noncrossovers, we determined the mean length of a conversion tract to be 32 bp. The spatial chromosome-wide distribution of noncrossovers and crossovers significantly differed, although both sets overlapped the known hotspots of PRDM9-directed histone methylation and DNA DSBs, thus supporting their origin in the standard DSB repair pathway. A significant deficit of noncrossovers descending from asymmetric DSBs proved their proposed adverse effect on meiotic recombination and pointed to sister chromatids as an alternative template for their repair. The finding has implications for the molecular mechanism of hybrid sterility in mice from crosses between closely related Mus musculus musculus and Mus musculus domesticus subspecies.

Identification of novel loci controlling inflammatory bowel disease susceptibility utilizing the genetic diversity of wild-derived mice.

Inflammatory bowel disease (IBD) is a complex disorder that imposes a growing health burden. Multiple genetic associations have been identified in IBD, but the mechanisms underlying many of these associations are poorly understood. Animal models are needed to bridge this gap, but conventional laboratory mouse strains lack the genetic diversity of human populations. To more accurately model human genetic diversity, we utilized a panel of chromosome (Chr) substitution strains, carrying chromosomes from the wild-derived and genetically divergent PWD/PhJ (PWD) strain on the commonly used C57BL/6J (B6) background, as well as their parental B6 and PWD strains. Two models of IBD were used, TNBS- and DSS-induced colitis. Compared with B6 mice, PWD mice were highly susceptible to TNBS-induced colitis, but resistant to DSS-induced colitis. Using consomic mice, we identified several PWD-derived loci that exhibited profound effects on IBD susceptibility. The most pronounced of these were loci on Chr1 and Chr2, which yielded high susceptibility in both IBD models, each acting at distinct phases of the disease. Leveraging transcriptomic data from B6 and PWD immune cells, together with a machine learning approach incorporating human IBD genetic associations, we identified lead candidate genes, including Itga4, Pip4k2a, Lcn10, Lgmn, and Gpr65.

Abstract P455: Epistatic Regulation of Hematological-related Traits and Gene Expression in Mice Reveals Additional Heritability Missed by Standard GWAS-type Analyses

The genetic contribution of additive versus non-additive (epistasis) effects in the regulation of hematologic and other complex traits is unclear. Although many variants have been associated with a range of complex traits via genome wide association studies (GWAS), these loci combined in additive models do not account for most of the trait heritability. GWAS-type analyses typically ignore gene-gene interactions, in part because of the difficulty in detecting them in complex multicellular organisms, especially humans. We have previously shown that mouse chromosome substitution strains (CSSs) are a powerful model for detecting epistasis, and that for certain complex traits the relative contribution of epistasis to heritability is as important as additivity. We have now applied the use of these CSSs to identify and map additive and epistatic loci that regulate a range of hematological-related traits and hepatic gene expression levels. A modified backcross was performed with CSS strains carrying the A/J-derived substituted chromosomes 4 and 6 on an otherwise C57BL/6J genetic background. By analyzing the transcriptomes of offspring from this cross, we identified and mapped additive quantitative trait loci (QTLs) that regulated the expression of 770 genes, and epistatic QTLs for 802 genes. Similarly we performed a complete blood analysis of offspring from the cross and identified additive QTLs for platelets and percentage of granulocyte in the blood as well as epistatic QTLs controlling the percentage of lymphocytes in the blood (rs13477644, rs13478739; LOD = 3.4) and red cell distribution width (rs13477864, rs13478802; LOD = 3.7). The variance attributable to the epistatic QTLs was approximately equal to that of the additive QTLs, highlighting the importance of identifying genetic interactions. Of note, even the SNPs associated with the most significant epistatic interactions were undetected in our single loci GWAS-like association analyses, demonstrating the need to specifically test for gene-gene interactions in studies of complex traits. In summary, our studies identified epistatic loci in mice that are important regulators of hematological-related traits and gene expression. Additionally, our studies call attention to the importance of extending single loci GWAS-type analyses to include analyses of gene-gene interactions to improve our ability to identify genetic variants that regulate complex traits.

Contribution of Chromosome 14 to Exercise Capacity and Training Responses in Mice.

Quantitative trait loci for exercise capacity and training-induced changes in exercise capacity were identified previously on mouse Chromosome 14. The aim of this study was to further investigate the role of Chromosome 14 in exercise capacity and responses to training in mice. Exercise phenotypes were measured in chromosome substitution strain mice carrying Chromosome 14 from the PWD/PhJ donor strain on the genetic background of a host C57BL/6J (B6) strain (B6.PWD14). Eight week old female and male mice from both strains completed a graded exercise test to exhaustion to assess intrinsic or baseline exercise capacity. A separate group of 12-week old female and male mice, randomly assigned to sedentary control (SED) or exercise training (EX) groups, completed a graded exercise test before and after a 4-week exercise training period. EX mice completed a 4-week training program consisting of treadmill running 5 days/week, 60 min/day at a final intensity of approximately 65% of maximum. For intrinsic exercise capacity, exercise time and work were significantly greater in female and male B6.PWD14 than sex-matched B6 mice. In the training study, female B6.PWD14 mice had higher pre-training exercise capacity than B6 mice. In contrast, there were no significant differences for pre-training exercise capacity between male B6 and B6.PWD14 mice. There were no significant strain differences for responses to training. These data demonstrate that PWD/PhJ alleles on Chromosome 14 significantly affect intrinsic exercise capacity. Furthermore, these results support continued efforts to identify candidate genes on Chromosome 14 underlying variation in exercise capacity.

Identification of novel loci controlling IBD susceptibility utilizing the genetic diversity of wild-derived mice

Abstract Inflammatory bowel disease (IBD) is a complex disease that imposes a significant and growing health burden in industrialized nations. Recent genetic studies have identified multiple genetic associations with IBD, but the mechanisms underlying these associations are poorly understood. Animal models are needed to bridge this gap in knowledge, but conventional laboratory mouse strains lack the genetic diversity of human populations. To more accurately model human genetic diversity and to identify new genetic loci that control susceptibility to IBD, we utilized a panel of chromosome (Chr) substitution (consomic) strains, carrying chromosomes from the wild-derived and genetically divergent PWD/PhJ (PWD) strain on the commonly used C57BL/6J (B6) background. Two models of IBD were used, TNBS- and DSS-induced colitis. Multiple disease severity parameters were evaluated, including weight loss, disease activity, fecal Lipocalin-2 production, and histopathology. Strikingly, compared with B6 mice, wild-derived PWD mice were highly susceptible to TNBS-induced colitis, but resistant to DSS-induced colitis. Using consomic mice, we found several PWD-derived loci that exhibited significant effects on IBD susceptibility. Most pronounced was a locus on Chr2, which resulted in very high susceptibility to both DSS and TNBS colitis, while several other PWD-derived loci were protective. Altogether, our experiments identified novel IBD susceptibility loci, harboring genes that control susceptibility to various aspects of IBD pathogenesis, and underscoring the utility of wild-derived mouse genetics. Future studies will identify these genes and their functions, to generate new insights into IBD etiology and pathophysiology.

Shared Genomic Regions Underlie Natural Variation in Diverse Toxin Responses.

Phenotypic complexity is caused by the contributions of environmental factors and multiple genetic loci, interacting or acting independently. Studies of yeast and Arabidopsis often find that the majority of natural variation across phenotypes is attributable to independent additive quantitative trait loci (QTL). Detected loci in these organisms explain most of the estimated heritable variation. By contrast, many heritable components underlying phenotypic variation in metazoan models remain undetected. Before the relative impacts of additive and interactive variance components on metazoan phenotypic variation can be dissected, high replication and precise phenotypic measurements are required to obtain sufficient statistical power to detect loci contributing to this missing heritability. Here, we used a panel of 296 recombinant inbred advanced intercross lines of Caenorhabditis elegans and a high-throughput fitness assay to detect loci underlying responses to 16 different toxins, including heavy metals, chemotherapeutic drugs, pesticides, and neuropharmaceuticals. Using linkage mapping, we identified 82 QTL that underlie variation in responses to these toxins, and predicted the relative contributions of additive loci and genetic interactions across various growth parameters. Additionally, we identified three genomic regions that impact responses to multiple classes of toxins. These QTL hotspots could represent common factors impacting toxin responses. We went further to generate near-isogenic lines and chromosome substitution strains, and then experimentally validated these QTL hotspots, implicating additive and interactive loci that underlie toxin-response variation.

High-Resolution Maps of Mouse Reference Populations.

Genetic reference panels are widely used to map complex, quantitative traits in model organisms. We have generated new high-resolution genetic maps of 259 mouse inbred strains from recombinant inbred strain panels (C57BL/6J × DBA/2J, ILS/IbgTejJ × ISS/IbgTejJ, and C57BL/6J × A/J) and chromosome substitution strain panels (C57BL/6J-Chr#<A/J>, C57BL/6J-Chr#<PWD/Ph>, and C57BL/6J-Chr#<MSM/Ms>). We genotyped all samples using the Affymetrix Mouse Diversity Array with an average intermarker spacing of 4.3 kb. The new genetic maps provide increased precision in the localization of recombination breakpoints compared to the previous maps. Although the strains were presumed to be fully inbred, we found residual heterozygosity in 40% of individual mice from five of the six panels. We also identified de novo deletions and duplications, in homozygous or heterozygous state, ranging in size from 21 kb to 8.4 Mb. Almost two-thirds (46 out of 76) of these deletions overlap exons of protein coding genes and may have phenotypic consequences. Twenty-nine putative gene conversions were identified in the chromosome substitution strains. We find that gene conversions are more likely to occur in regions where the homologous chromosomes are more similar. The raw genotyping data and genetic maps of these strain panels are available at http://churchill-lab.jax.org/website/MDA.

Widespread epistasis regulates glucose homeostasis and gene expression.

The relative contributions of additive versus non-additive interactions in the regulation of complex traits remains controversial. This may be in part because large-scale epistasis has traditionally been difficult to detect in complex, multi-cellular organisms. We hypothesized that it would be easier to detect interactions using mouse chromosome substitution strains that simultaneously incorporate allelic variation in many genes on a controlled genetic background. Analyzing metabolic traits and gene expression levels in the offspring of a series of crosses between mouse chromosome substitution strains demonstrated that inter-chromosomal epistasis was a dominant feature of these complex traits. Epistasis typically accounted for a larger proportion of the heritable effects than those due solely to additive effects. These epistatic interactions typically resulted in trait values returning to the levels of the parental CSS host strain. Due to the large epistatic effects, analyses that did not account for interactions consistently underestimated the true effect sizes due to allelic variation or failed to detect the loci controlling trait variation. These studies demonstrate that epistatic interactions are a common feature of complex traits and thus identifying these interactions is key to understanding their genetic regulation.

Candidate genes on murine chromosome 8 are associated with susceptibility to Staphylococcus aureus infection in mice and are involved with Staphylococcus aureus septicemia in humans.

We previously showed that chromosome 8 of A/J mice was associated with susceptibility to S. aureus infection. However, the specific genes responsible for this susceptibility are unknown. Chromosome substitution strain 8 (CSS8) mice, which have chromosome 8 from A/J but an otherwise C57BL/6J genome, were used to identify the genetic determinants of susceptibility to S. aureus on chromosome 8. Quantitative trait loci (QTL) mapping of S. aureus-infected N2 backcross mice (F1 [C8A] × C57BL/6J) identified a locus 83180780–88103009 (GRCm38/mm10) on A/J chromosome 8 that was linked to S. aureus susceptibility. All genes on the QTL (n~ 102) were further analyzed by three different strategies: 1) different expression in susceptible (A/J) and resistant (C57BL/6J) mice only in response to S. aureus, 2) consistently different expression in both uninfected and infected states between the two strains, and 3) damaging non-synonymous SNPs in either strain. Eleven candidate genes from the QTL region were significantly differently expressed in patients with S. aureus infection vs healthy human subjects. Four of these 11 genes also exhibited significantly different expression in S. aureus-challenged human neutrophils: Ier2, Crif1, Cd97 and Lyl1. CD97 ligand binding was evaluated within peritoneal neutrophils from A/J and C57BL/6J. CD97 from A/J had stronger CD55 but weaker integrin α5β1 ligand binding as compared with C57BL/6J. Because CD55/CD97 binding regulates immune cell activation and cytokine production, and integrin α5β1 is a membrane receptor for fibronectin, which is also bound by S. aureus, strain-specific differences could contribute to susceptibility to S. aureus. Down-regulation of Crif1 with siRNA was associated with increased host cell apoptosis among both naïve and S. aureus-infected bone marrow-derived macrophages. Specific genes in A/J chromosome 8, including Cd97 and Crif1, may play important roles in host defense against S. aureus.

Mapping of a FEB3 homologous febrile seizure locus on mouse chromosome 2 containing candidate genes Scn1a and Scn3a.

Febrile seizures (FS) are the most common seizure type in children. Recurrent FS are a risk factor for developing temporal lobe epilepsy later in life and are known to have a strong genetic component. Experimental FS (eFS) can be elicited in mice by warm-air induced hyperthermia. We used this model to screen the chromosome substitution strain (CSS) panel derived from C57BL/6J and A/J for FS susceptibility and identified C57BL/6J-Chr2A /NaJ (CSS2), as the strain with the strongest FS susceptibility phenotype. The aim of this study was to map FS susceptibility loci and select candidate genes on mouse chromosome 2. We generated an F2 population by intercrossing the hybrids (F1 ) that were derived from CSS2 and C57BL/6J mice. All CSS2-F2 individuals were genotyped and phenotyped for eFS susceptibility, and QTL analysis was performed. Candidate gene selection was based on bioinformatics analyses and differential brain expression between CSS2 and C57BL/6J strains determined by microarray analysis. Genetic mapping of the eFS susceptibility trait identified two significant loci: FS-QTL2a (LOD-score 3.6) and FS-QTL2b (LOD-score 6.2). FS-QTL2a contained 44 genes expressed in the brain at post natal day 14. Four of these (Arl6ip6, Cytip, Fmnl2 Ifih1) contained a non-synonymous SNP comparing CSS2 and C57BL/6J, six genes (March7, Nr4a2, Gpd2, Grb14, Scn1a, Scn3a) were differentially expressed between these strains. A region within FS-QTL2a is homologous to the human FEB3 locus. The fact that we identify mouse FS-QTL2a with high FEB3 homology is strong support for the validity of the eFS mouse model to study genetics of human FS.

Genome Sequencing of Chromosome 1 Substitution Lines Derived from Chinese Wild Mice Revealed a Unique Resource for Genetic Studies of Complex Traits.

Mouse resources such as Collaborative Cross, outbred stocks, Hybrid Mouse Diversity Panel, and chromosome substitution strains have been instrumental to many progresses in the studies of complex traits genetics. We have established a population of chromosome 1 (Chr 1) substitution lines (C1SLs) in which donor chromosomes were derived from Chinese wild mice. Genome sequencing of 18 lines of this population showed that Chr 1 had been replaced by the donor chromosome. About 4.5 million unique single nucleotide polymorphisms and indels were discovered on Chr 1, of which 1.3 million were novel. Compared with sequenced classical inbred strains, Chr 1 of each C1SL had fivefold more variants, and more loss of function and potentially regulatory variants. Further haplotype analysis showed that the donor chromosome accumulated more historical recombination events, with the largest haplotype block being only 100 kb, and about 57% of the blocks were <1 kb. Subspecies origin analysis showed that these chromosomes had a mosaic genome structure that dominantly originated from Mus musculus musculus and M. m. castaneus subspecies, except for the C57BL/6J-Chr1KM line from M. m. domesticus. In addition, phenotyping four of these lines on blood biochemistry suggested that there were substantial phenotypic variations among our lines, especially line C57BL/6J-Chr1HZ and donor strain C57BL/6J. Further gene ontology enrichment revealed that the differentially expressed genes among liver-expressed genes between C57BL/6J and C57BL/6J-Chr1HZ were enriched in lipid metabolism biological processes. All these characteristics enable C1SLs to be a unique resource for identifying and fine mapping quantitative trait loci on mouse Chr 1, and carrying out systems genetics studies of complex traits.

Consomic mouse strain selection based on effect size measurement, statistical significance testing and integrated behavioral z-scoring: focus on anxiety-related behavior and locomotion.

BackgroundSelecting chromosome substitution strains (CSSs, also called consomic strains/lines) used in the search for quantitative trait loci (QTLs) consistently requires the identification of the respective phenotypic trait of interest and is simply based on a significant difference between a consomic and host strain. However, statistical significance as represented by P values does not necessarily predicate practical importance. We therefore propose a method that pays attention to both the statistical significance and the actual size of the observed effect. The present paper extends on this approach and describes in more detail the use of effect size measures (Cohen’s d, partial eta squared - ηp2) together with the P value as statistical selection parameters for the chromosomal assignment of QTLs influencing anxiety-related behavior and locomotion in laboratory mice.ResultsThe effect size measures were based on integrated behavioral z-scoring and were calculated in three experiments: (A) a complete consomic male mouse panel with A/J as the donor strain and C57BL/6J as the host strain. This panel, including host and donor strains, was analyzed in the modified Hole Board (mHB). The consomic line with chromosome 19 from A/J (CSS-19A) was selected since it showed increased anxiety-related behavior, but similar locomotion compared to its host. (B) Following experiment A, female CSS-19A mice were compared with their C57BL/6J counterparts; however no significant differences and effect sizes close to zero were found. (C) A different consomic mouse strain (CSS-19PWD), with chromosome 19 from PWD/PhJ transferred on the genetic background of C57BL/6J, was compared with its host strain. Here, in contrast with CSS-19A, there was a decreased overall anxiety in CSS-19PWD compared to C57BL/6J males, but not locomotion.ConclusionsThis new method shows an improved way to identify CSSs for QTL analysis for anxiety-related behavior using a combination of statistical significance testing and effect sizes. In addition, an intercross between CSS-19A and CSS-19PWD may be of interest for future studies on the genetic background of anxiety-related behavior.Electronic supplementary materialThe online version of this article (doi:10.1186/s12863-016-0411-4) contains supplementary material, which is available to authorized users.

Genetic Links between Recombination and Speciation

Genetic Links between Recombination and Speciation

Hybrid Sterility Locus on Chromosome X Controls Meiotic Recombination Rate in Mouse.

Meiotic recombination safeguards proper segregation of homologous chromosomes into gametes, affects genetic variation within species, and contributes to meiotic chromosome recognition, pairing and synapsis. The Prdm9 gene has a dual role, it controls meiotic recombination by determining the genomic position of crossover hotspots and, in infertile hybrids of house mouse subspecies Mus m. musculus (Mmm) and Mus m. domesticus (Mmd), it further functions as the major hybrid sterility gene. In the latter role Prdm9 interacts with the hybrid sterility X 2 (Hstx2) genomic locus on Chromosome X (Chr X) by a still unknown mechanism. Here we investigated the meiotic recombination rate at the genome-wide level and its possible relation to hybrid sterility. Using immunofluorescence microscopy we quantified the foci of MLH1 DNA mismatch repair protein, the cytological counterparts of reciprocal crossovers, in a panel of inter-subspecific chromosome substitution strains. Two autosomes, Chr 7 and Chr 11, significantly modified the meiotic recombination rate, yet the strongest modifier, designated meiotic recombination 1, Meir1, emerged in the 4.7 Mb Hstx2 genomic locus on Chr X. The male-limited transgressive effect of Meir1 on recombination rate parallels the male-limited transgressive role of Hstx2 in hybrid male sterility. Thus, both genetic factors, the Prdm9 gene and the Hstx2/Meir1 genomic locus, indicate a link between meiotic recombination and hybrid sterility. A strong female-specific modifier of meiotic recombination rate with the effect opposite to Meir1 was localized on Chr X, distally to Meir1. Mapping Meir1 to a narrow candidate interval on Chr X is an important first step towards positional cloning of the respective gene(s) responsible for variation in the global recombination rate between closely related mouse subspecies.

Three interacting genomic loci incorporating two novel mutations underlie the evolution of diet-induced diabetes.

We investigated the pathophysiology of diet-induced diabetes in the Cohen diabetic rat (CDs/y) from its induction to its chronic phase, using a multi-layered integrated genomic approach. We identified by linkage analysis two diabetes-related quantitative trait loci on RNO4 and RNO13. We determined their functional contribution to diabetes by chromosomal substitution, using congenic and consomic strains. To identify within these loci genes of relevance to diabetes, we sequenced the genome of CDs/y and compared it to 25 other rat strains. Within the RNO4 locus, we detected a novel high impact deletion in the Ndufa4 gene that was unique to CDs/y. Within the RNO13 locus, we found multiple SNPs and INDELs that were unique to CDs/y but were unable to prioritize any of the genes. Genome wide screening identified a novel third locus not detected by linkage analysis that consisted of a novel high impact deletion on RNO11 that was unique to CDs/y and that involved the Sdf2l1 gene. Using co-segregation analysis, we investigated in silico the relative contribution to the diabetic phenotype and the interaction between the three genomic loci on RNO4, RNO11 and RNO13. We found that the RNO4 locus plays a major role during the induction of diabetes, whereas the genomic loci on RNO13 and RNO11, while interacting with the RNO4 locus, contribute more significantly to the diabetic phenotype during the chronic phase of the disease. The mechanisms whereby the mutations on RNO4 and 11 and the RNO13 locus contribute to the development of diabetes are under continuing investigation.