A cross-species approach using an in vivo evaluation platform in mice demonstrates that sequence variation in human RABEP2 modulates ischemic stroke outcomes.

Ischemic stroke, caused by vessel blockage, results in cerebral infarction, the death of brain tissue. Previously, quantitative trait locus (QTL) mapping of cerebral infarct volume and collateral vessel number identified a single, strong genetic locus regulating both phenotypes. Additional studies identified RAB GTPase-binding effector protein 2 (Rabep2) as the casual gene. However, there is yet no evidence that variation in the human ortholog of this gene plays any role in ischemic stroke outcomes. We established an in vivo evaluation platform in mice by using adeno-associated virus (AAV) gene replacement and verified that both mouse and human RABEP2 rescue the mouse Rabep2 knockout ischemic stroke volume and collateral vessel phenotypes. Importantly, this cross-species complementation enabled us to experimentally investigate the functional effects of coding sequence variation in human RABEP2. We chose four coding variants from the human population that are predicted by multiple in silico algorithms to be damaging to RABEP2 function. In vitro and in vivo analyses verify that all four led to decreased collateral vessel connections and increased infarct volume. Thus, there are naturally occurring loss-of-function alleles. This cross-species approach will expand the number of targets for therapeutics development for ischemic stroke.

Impacts of Gestational FireMaster 550 Exposure on the Neonatal Cortex Are Sex Specific and Largely Attributable to the Organophosphate Esters.

INTRODUCTION: Flame retardants (FRs) are common bodily and environmental pollutants, creating concern about their potential toxicity. We and others have found that the commercial mixture FireMaster® 550 (FM 550) or its individual brominated (BFR) and organophosphate ester (OPFR) components are potential developmental neurotoxicants. Using Wistar rats, we previously reported that developmental exposure to FM 550 or its component classes produced sex- and compound-specific effects on adult socioemotional behaviors. The underlying mechanisms driving the behavioral phenotypes are unknown. METHODS: To further mechanistic understanding, here we conducted transcriptomics in parallel with a novel lipidomics approach using cortical tissues from newborn siblings of the rats in the published behavioral study. Inclusion of lipid composition is significant because it is rarely examined in developmental neurotoxicity studies. Pups were gestationally exposed via oral dosing to the dam to FM 550 or the BFR or OPFR components at environmentally relevant doses. RESULTS: The neonatal cortex was highly sexually dimorphic in lipid and transcriptome composition, and males were more significantly impacted by FR exposure. Multiple adverse modes of action for the BFRs and OPFRs on neurodevelopment were identified, with the OPFRs being more disruptive than the BFRs via multiple mechanisms including dysregulation of mitochondrial function and disruption of cholinergic and glutamatergic systems. Disrupted mitochondrial function by environmental factors has been linked to a higher risk of autism spectrum disorders and neurodegenerative disorders. Impacted lipid classes included ceramides, sphingomyelins, and triacylglycerides. Robust ceramide upregulation in the OPFR females could suggest a heightened risk of brain metabolic disease. CONCLUSIONS: This study reveals multiple mechanisms by which the components of a common FR mixture are developmentally neurotoxic and that the OPFRs may be the compounds of greatest concern.

Population-based dose-response analysis of liver transcriptional response to trichloroethylene in mouse.

Studies of gene expression are common in toxicology and provide important clues to mechanistic understanding of adverse effects of chemicals. Most prior studies have been performed in a single strain or cell line; however, gene expression is heavily influenced by the genetic background, and these genotype-expression differences may be key drivers of inter-individual variation in response to chemical toxicity. In this study, we hypothesized that the genetically diverse Collaborative Cross mouse population can be used to gain insight and suggest mechanistic hypotheses for the dose- and genetic background-dependent effects of chemical exposure. This hypothesis was tested using a model liver toxicant trichloroethylene (TCE). Liver transcriptional responses to TCE exposure were evaluated 24 h after dosing. Transcriptomic dose-responses were examined for both TCE and its major oxidative metabolite trichloroacetic acid (TCA). As expected, peroxisome- and fatty acid metabolism-related pathways were among the most dose-responsive enriched pathways in all strains. However, nearly half of the TCE-induced liver transcriptional perturbation was strain-dependent, with abundant evidence of strain/dose interaction, including in the peroxisomal signaling-associated pathways. These effects were highly concordant between the administered TCE dose and liver levels of TCA. Dose-response analysis of gene expression at the pathway level yielded points of departure similar to those derived from the traditional toxicology studies for both non-cancer and cancer effects. Mapping of expression-genotype-dose relationships revealed some significant associations; however, the effects of TCE on gene expression in liver appear to be highly polygenic traits that are challenging to positionally map. This study highlights the usefulness of mouse population-based studies in assessing inter-individual variation in toxicological responses, but cautions that genetic mapping may be challenging because of the complexity in gene exposure-dose relationships.

Genome Wide Identification of SARS-CoV Susceptibility Loci Using the Collaborative Cross.

New systems genetics approaches are needed to rapidly identify host genes and genetic networks that regulate complex disease outcomes. Using genetically diverse animals from incipient lines of the Collaborative Cross mouse panel, we demonstrate a greatly expanded range of phenotypes relative to classical mouse models of SARS-CoV infection including lung pathology, weight loss and viral titer. Genetic mapping revealed several loci contributing to differential disease responses, including an 8.5Mb locus associated with vascular cuffing on chromosome 3 that contained 23 genes and 13 noncoding RNAs. Integrating phenotypic and genetic data narrowed this region to a single gene, Trim55, an E3 ubiquitin ligase with a role in muscle fiber maintenance. Lung pathology and transcriptomic data from mice genetically deficient in Trim55 were used to validate its role in SARS-CoV-induced vascular cuffing and inflammation. These data establish the Collaborative Cross platform as a powerful genetic resource for uncovering genetic contributions of complex traits in microbial disease severity, inflammation and virus replication in models of outbred populations.

A multi-megabase copy number gain causes maternal transmission ratio distortion on mouse chromosome 2.

Significant departures from expected Mendelian inheritance ratios (transmission ratio distortion, TRD) are frequently observed in both experimental crosses and natural populations. TRD on mouse Chromosome (Chr) 2 has been reported in multiple experimental crosses, including the Collaborative Cross (CC). Among the eight CC founder inbred strains, we found that Chr 2 TRD was exclusive to females that were heterozygous for the WSB/EiJ allele within a 9.3 Mb region (Chr 2 76.9 - 86.2 Mb). A copy number gain of a 127 kb-long DNA segment (designated as responder to drive, R2d) emerged as the strongest candidate for the causative allele. We mapped R2d sequences to two loci within the candidate interval. R2d1 is located near the proximal boundary, and contains a single copy of R2d in all strains tested. R2d2 maps to a 900 kb interval, and the number of R2d copies varies from zero in classical strains (including the mouse reference genome) to more than 30 in wild-derived strains. Using real-time PCR assays for the copy number, we identified a mutation (R2d2WSBdel1) that eliminates the majority of the R2d2WSB copies without apparent alterations of the surrounding WSB/EiJ haplotype. In a three-generation pedigree segregating for R2d2WSBdel1, the mutation is transmitted to the progeny and Mendelian segregation is restored in females heterozygous for R2d2WSBdel1, thus providing direct evidence that the copy number gain is causal for maternal TRD. We found that transmission ratios in R2d2WSB heterozygous females vary between Mendelian segregation and complete distortion depending on the genetic background, and that TRD is under genetic control of unlinked distorter loci. Although the R2d2WSB transmission ratio was inversely correlated with average litter size, several independent lines of evidence support the contention that female meiotic drive is the cause of the distortion. We discuss the implications and potential applications of this novel meiotic drive system.

Elucidation of the transcription network governing mammalian sex determination by exploiting strain-specific susceptibility to sex reversal.

Despite the identification of some key genes that regulate sex determination, most cases of disorders of sexual development remain unexplained. Evidence suggests that the sexual fate decision in the developing gonad depends on a complex network of interacting factors that converge on a critical threshold. To elucidate the transcriptional network underlying sex determination, we took the first expression quantitative trait loci (eQTL) approach in a developing organ. We identified reproducible differences in the transcriptome of the embryonic day 11.5 (E11.5) XY gonad between C57BL/6J (B6) and 129S1/SvImJ (129S1), indicating that the reported sensitivity of B6 to sex reversal is consistent with a higher expression of a female-like transcriptome in B6. Gene expression is highly variable in F2 XY gonads from B6 and 129S1 intercrosses, yet strong correlations emerged. We estimated the F2 coexpression network and predicted roles for genes of unknown function based on their connectivity and position within the network. A genetic analysis of the F2 population detected autosomal regions that control the expression of many sex-related genes, including Sry (sex-determining region of the Y chromosome) and Sox9 (Sry-box containing gene 9), the key regulators of male sex determination. Our results reveal the complex transcription architecture underlying sex determination, and provide a mechanism by which individuals may be sensitized for sex reversal.

Modeling host genetic regulation of influenza pathogenesis in the collaborative cross.

Genetic variation contributes to host responses and outcomes following infection by influenza A virus or other viral infections. Yet narrow windows of disease symptoms and confounding environmental factors have made it difficult to identify polymorphic genes that contribute to differential disease outcomes in human populations. Therefore, to control for these confounding environmental variables in a system that models the levels of genetic diversity found in outbred populations such as humans, we used incipient lines of the highly genetically diverse Collaborative Cross (CC) recombinant inbred (RI) panel (the pre-CC population) to study how genetic variation impacts influenza associated disease across a genetically diverse population. A wide range of variation in influenza disease related phenotypes including virus replication, virus-induced inflammation, and weight loss was observed. Many of the disease associated phenotypes were correlated, with viral replication and virus-induced inflammation being predictors of virus-induced weight loss. Despite these correlations, pre-CC mice with unique and novel disease phenotype combinations were observed. We also identified sets of transcripts (modules) that were correlated with aspects of disease. In order to identify how host genetic polymorphisms contribute to the observed variation in disease, we conducted quantitative trait loci (QTL) mapping. We identified several QTL contributing to specific aspects of the host response including virus-induced weight loss, titer, pulmonary edema, neutrophil recruitment to the airways, and transcriptional expression. Existing whole-genome sequence data was applied to identify high priority candidate genes within QTL regions. A key host response QTL was located at the site of the known anti-influenza Mx1 gene. We sequenced the coding regions of Mx1 in the eight CC founder strains, and identified a novel Mx1 allele that showed reduced ability to inhibit viral replication, while maintaining protection from weight loss.

Integrative genetic analysis of allergic inflammation in the murine lung.

Airway allergen exposure induces inflammation among individuals with atopy that is characterized by altered airway gene expression, elevated levels of T helper type 2 cytokines, mucus hypersecretion, and airflow obstruction. To identify the genetic determinants of the airway allergen response, we employed a systems genetics approach. We applied a house dust mite mouse model of allergic airway disease to 151 incipient lines of the Collaborative Cross, a new mouse genetic reference population, and measured serum IgE, airway eosinophilia, and gene expression in the lung. Allergen-induced serum IgE and airway eosinophilia were not correlated. We detected quantitative trait loci (QTL) for airway eosinophilia on chromosome (Chr) 11 (71.802-87.098 megabases [Mb]) and allergen-induced IgE on Chr 4 (13.950-31.660 Mb). More than 4,500 genes expressed in the lung had gene expression QTL (eQTL), the majority of which were located near the gene itself. However, we also detected approximately 1,700 trans-eQTL, and many of these trans-eQTL clustered into two regions on Chr 2. We show that one of these loci (at 147.6 Mb) is associated with the expression of more than 100 genes, and, using bioinformatics resources, fine-map this locus to a 53 kb-long interval. We also use the gene expression and eQTL data to identify a candidate gene, Tlcd2, for the eosinophil QTL. Our results demonstrate that hallmark allergic airway disease phenotypes are associated with distinct genetic loci on Chrs 4 and 11, and that gene expression in the allergically inflamed lung is controlled by both cis and trans regulatory factors.

Genetic analysis of complex traits in the emerging Collaborative Cross.

The Collaborative Cross (CC) is a mouse recombinant inbred strain panel that is being developed as a resource for mammalian systems genetics. Here we describe an experiment that uses partially inbred CC lines to evaluate the genetic properties and utility of this emerging resource. Genome-wide analysis of the incipient strains reveals high genetic diversity, balanced allele frequencies, and dense, evenly distributed recombination sites-all ideal qualities for a systems genetics resource. We map discrete, complex, and biomolecular traits and contrast two quantitative trait locus (QTL) mapping approaches. Analysis based on inferred haplotypes improves power, reduces false discovery, and provides information to identify and prioritize candidate genes that is unique to multifounder crosses like the CC. The number of expression QTLs discovered here exceeds all previous efforts at eQTL mapping in mice, and we map local eQTL at 1-Mb resolution. We demonstrate that the genetic diversity of the CC, which derives from random mixing of eight founder strains, results in high phenotypic diversity and enhances our ability to map causative loci underlying complex disease-related traits.

Genetically determined severity of anti-myeloperoxidase glomerulonephritis.

Myeloperoxidase (MPO) is a target antigen for antineutrophil cytoplasmic autoantibodies (ANCA). There is evidence that MPO-ANCA cause necrotizing and crescentic glomerulonephritis (NCGN) and vasculitis. NCGN severity varies among patients with ANCA disease, and genetic factors influence disease severity. The role of genetics in MPO-ANCA NCGN severity was investigated using 13 inbred mouse strains, F1 and F2 hybrids, bone marrow chimeras, and neutrophil function assays. Mouse strains include founders of the Collaborative Cross. Intravenous injection of anti-MPO IgG induced glomerular crescents in >60% of glomeruli in 129S6/SvEv and CAST/EiJ mice, but <1% in A/J, DBA/1J, DBA/2J, NOD/LtJ, and PWK/PhJ mice. C57BL6J, 129S1/SvImJ, LP/J, WSB/EiJ, NZO/HILtJ, and C3H mice had intermediate severity. High-density genotypes at 542,190 single nucleotide polymorphisms were used to identify candidate loci for disease severity by identifying genomic regions that are different between 129S6/SvEv and 129S1/SvImJ mice, which are genetically similar but phenotypically distinct. C57BL/6 × 129S6 F2 mice were genotyped at 76 SNPs to capture quantitative trait loci for disease severity. The absence of a dominant quantitative trait locus suggests that differences in severity are the result of multiple gene interactions. In vivo studies using bone marrow chimeric mice and in vitro studies of neutrophil activation by anti-MPO IgG indicated that severity of NCGN is mediated by genetically determined differences in the function of neutrophils.

Architecture of energy balance traits in emerging lines of the Collaborative Cross.

The potential utility of the Collaborative Cross (CC) mouse resource was evaluated to better understand complex traits related to energy balance. A primary focus was to examine if genetic diversity in emerging CC lines (pre-CC) would translate into equivalent phenotypic diversity. Second, we mapped quantitative trait loci (QTL) for 15 metabolism- and exercise-related phenotypes in this population. We evaluated metabolic and voluntary exercise traits in 176 pre-CC lines, revealing phenotypic variation often exceeding that seen across the eight founder strains from which the pre-CC was derived. Many phenotypic correlations existing within the founder strains were no longer significant in the pre-CC population, potentially representing reduced linkage disequilibrium (LD) of regions harboring multiple genes with effects on energy balance or disruption of genetic structure of extant inbred strains with substantial shared ancestry. QTL mapping revealed five significant and eight suggestive QTL for body weight (Chr 4, 7.54 Mb; CI 3.32-10.34 Mb; Bwq14), body composition, wheel running (Chr 16, 33.2 Mb; CI 32.5-38.3 Mb), body weight change in response to exercise (1: Chr 6, 77.7Mb; CI 72.2-83.4 Mb and 2: Chr 6, 42.8 Mb; CI 39.4-48.1 Mb), and food intake during exercise (Chr 12, 85.1 Mb; CI 82.9-89.0 Mb). Some QTL overlapped with previously mapped QTL for similar traits, whereas other QTL appear to represent novel loci. These results suggest that the CC will be a powerful, high-precision tool for examining the genetic architecture of complex traits such as those involved in regulation of energy balance.

From classical genetics to quantitative genetics to systems biology: modeling epistasis.

Gene expression data has been used in lieu of phenotype in both classical and quantitative genetic settings. These two disciplines have separate approaches to measuring and interpreting epistasis, which is the interaction between alleles at different loci. We propose a framework for estimating and interpreting epistasis from a classical experiment that combines the strengths of each approach. A regression analysis step accommodates the quantitative nature of expression measurements by estimating the effect of gene deletions plus any interaction. Effects are selected by significance such that a reduced model describes each expression trait. We show how the resulting models correspond to specific hierarchical relationships between two regulator genes and a target gene. These relationships are the basic units of genetic pathways and genomic system diagrams. Our approach can be extended to analyze data from a variety of experiments, multiple loci, and multiple environments.

A Neuroprotective Locus Modulates Ischemic Stroke Infarction Independent of Collateral Vessel Anatomy.

Although studies with inbred strains of mice have shown that infarct size is largely determined by the extent of collateral vessel connections between arteries in the brain that enable reperfusion of the ischemic territory, we have identified strain pairs that do not vary in this vascular phenotype, but which nonetheless exhibit large differences in infarct size. In this study we performed quantitative trait locus (QTL) mapping in mice from an intercross between two such strains, WSB/EiJ (WSB) and C57BL/6J (B6). This QTL mapping revealed only one neuroprotective locus on Chromosome 8 (Chr 8) that co-localizes with a neuroprotective locus we mapped previously from F2 progeny between C3H/HeJ (C3H) and B6. The allele-specific phenotypic effect on infarct volume at the genetic region identified by these two independent mappings was in the opposite direction of the parental strain phenotype; namely, the B6 allele conferred increased susceptibility to ischemic infarction. Through two reciprocal congenic mouse lines with either the C3H or B6 background at the Chr 8 locus, we verified the neuroprotective effects of this genetic region that modulates infarct volume without any effect on the collateral vasculature. Additionally, we surveyed non-synonymous coding SNPs and performed RNA-sequencing analysis to identify potential candidate genes within the genetic interval. Through these approaches, we suggest new genes for future mechanistic studies of infarction following ischemic stroke, which may represent novel gene/protein targets for therapeutic development.

Age and Genetic Background Modify Hybrid Male Sterility in House Mice.

Hybrid male sterility (HMS) contributes to reproductive isolation commonly observed among house mouse (Mus musculus) subspecies, both in the wild and in laboratory crosses. Incompatibilities involving specific Prdm9 alleles and certain Chromosome (Chr) X genotypes are known determinants of fertility and HMS, and previous work in the field has demonstrated that genetic background modifies these two major loci. We constructed hybrids that have identical genotypes at Prdm9 and identical X chromosomes, but differ widely across the rest of the genome. In each case, we crossed female PWK/PhJ mice representative of the M. m. musculus subspecies to males from a classical inbred strain representative of M. m. domesticus: 129S1/SvImJ, A/J, C57BL/6J, or DBA/2J. We detected three distinct trajectories of fertility among the hybrids using breeding experiments. The PWK129S1 males were always infertile. PWKDBA2 males were fertile, despite their genotypes at the major HMS loci. We also observed age-dependent changes in fertility parameters across multiple genetic backgrounds. The PWKB6 and PWKAJ males were always infertile before 12 weeks and after 35 weeks. However, some PWKB6 and PWKAJ males were transiently fertile between 12 and 35 weeks. This observation could resolve previous contradictory reports about the fertility of PWKB6. Taken together, these results point to multiple segregating HMS modifier alleles, some of which have age-related modes of action. The ultimate identification of these alleles and their age-related mechanisms will advance understanding both of the genetic architecture of HMS and of how reproductive barriers are maintained between house mouse subspecies.

Novel Neuroprotective Loci Modulating Ischemic Stroke Volume in Wild-Derived Inbred Mouse Strains.

To identify genes involved in cerebral infarction, we have employed a forward genetic approach in inbred mouse strains, using quantitative trait loci (QTL) mapping for cerebral infarct volume after middle cerebral artery occlusion. We had previously observed that infarct volume is inversely correlated with cerebral collateral vessel density in most strains. In this study, we expanded the pool of allelic variation among classical inbred mouse strains by utilizing the eight founder strains of the Collaborative Cross and found a wild-derived strain, WSB/EiJ, that breaks this general rule that collateral vessel density inversely correlates with infarct volume. WSB/EiJ and another wild-derived strain, CAST/EiJ, show the highest collateral vessel densities of any inbred strain, but infarct volume of WSB/EiJ mice is 8.7-fold larger than that of CAST/EiJ mice. QTL mapping between these strains identified four new neuroprotective loci modulating cerebral infarct volume while not affecting collateral vessel phenotypes. To identify causative variants in genes, we surveyed nonsynonymous coding SNPs between CAST/EiJ and WSB/EiJ and found 96 genes harboring coding SNPs predicted to be damaging and mapping within one of the four intervals. In addition, we performed RNA-sequencing for brain tissue of CAST/EiJ and WSB/EiJ mice and identified 79 candidate genes mapping in one of the four intervals showing strain-specific differences in expression. The identification of the genes underlying these neuroprotective loci will provide new understanding of genetic risk factors of ischemic stroke, which may provide novel targets for future therapeutic intervention of human ischemic stroke.

Permissiveness to form pluripotent stem cells may be an evolutionarily derived characteristic in Mus musculus.

Mus musculus is the only known species from which embryonic stem cells (ESC) can be isolated under conditions requiring only leukemia inhibitory factor (LIF). Other species are non-permissive in LIF media, and form developmentally primed epiblast stem cells (EpiSC) similar to cells derived from post-implantation, egg cylinders. To evaluate whether non-permissiveness extends to induced pluripotent stem cells (iPSC), we derived iPSC from the eight founder strains of the mouse Collaborative Cross. Two strains, NOD/ShiLtJ and the WSB/EiJ, were non-permissive, consistent with the previous classification of NOD/ShiLtJ as non-permissive to ESC derivation. We determined non-permissiveness is recessive, and that non-permissive genomes do not compliment. We overcame iPSC non-permissiveness by using GSK3B and MEK inhibitors with serum, a technique we termed 2iS reprogramming. Although used for ESC derivation, GSK3B and MEK inhibitors have not been used during iPSC reprogramming because they inhibit survival of progenitor differentiated cells. iPSC derived in 2iS are more transcriptionally similar to ESC than EpiSC, indicating that 2iS reprogramming acts to overcome genetic background constraints. Finally, of species tested for ESC or iPSC derivation, only some M. musculus strains are permissive under LIF culture conditions suggesting that this is an evolutionarily derived characteristic in the M. musculus lineage.

eQTL Viewer: visualizing how sequence variation affects genome-wide transcription.

BACKGROUND: Expression Quantitative Trait Locus (eQTL) mapping methods have been used to identify the genetic basis of gene expression variations. To map eQTL, thousands of expression profiles are related with sequence polymorphisms across the genome through their correlated variations. These eQTL distribute in many chromosomal regions, each of which can include many genes. The large number of mapping results produced makes it difficult to consider simultaneously the relationships between multiple genomic regions and multiple expressional profiles. There is a need for informative bioinformatics tools to assist the visualization and interpretation of these mapping results. RESULTS: We have developed a web-based tool, called eQTL Viewer, to visualize the relationships between the expression trait genes and the candidate genes in the eQTL regions using Scalable Vector Graphics. The plot generated by eQTL Viewer has the capacity to display mapping results with high resolutions at a variety of scales, and superimpose biological annotations onto the mapping results dynamically. CONCLUSION: Our tool provides an efficient and intuitive way for biologists to explore transcriptional regulation patterns, and to generate hypotheses on the genetic basis of transcriptional regulations.

Editor's Highlight: Collaborative Cross Mouse Population Enables Refinements to Characterization of the Variability in Toxicokinetics of Trichloroethylene and Provides Genetic Evidence for the Role of PPAR Pathway in Its Oxidative Metabolism.

Background: Trichloroethylene (TCE) is a known carcinogen in humans and rodents. Previous studies of inter-strain variability in TCE metabolism were conducted in multi-strain panels of classical inbred mice with limited genetic diversity to identify gene-environment interactions associated with chemical exposure. Objectives: To evaluate inter-strain variability in TCE metabolism and identify genetic determinants that are associated with TCE metabolism and effects using Collaborative Cross (CC), a large panel of genetically diverse strains of mice. Methods: We administered a single oral dose of 0, 24, 80, 240, or 800 mg/kg of TCE to mice from 50 CC strains, and collected organs 24 h post-dosing. Levels of trichloroacetic acid (TCA), a major oxidative metabolite of TCE were measured in multiple tissues. Protein expression and activity levels of TCE-metabolizing enzymes were evaluated in the liver. Liver transcript levels of known genes perturbed by TCE exposure were also quantified. Genetic association mapping was performed on the acquired phenotypes. Results: TCA levels varied in a dose- and strain-dependent manner in liver, kidney, and serum. The variability in TCA levels among strains did not correlate with expression or activity of a number of enzymes known to be involved in TCE oxidation. Peroxisome proliferator-activated receptor alpha (PPARα)-responsive genes were found to be associated with strain-specific differences in TCE metabolism. Conclusions: This study shows that CC mouse population is a valuable tool to quantitatively evaluate inter-individual variability in chemical metabolism and to identify genes and pathways that may underpin population differences.

Male Infertility Is Responsible for Nearly Half of the Extinction Observed in the Mouse Collaborative Cross.

The goal of the Collaborative Cross (CC) project was to generate and distribute over 1000 independent mouse recombinant inbred strains derived from eight inbred founders. With inbreeding nearly complete, we estimated the extinction rate among CC lines at a remarkable 95%, which is substantially higher than in the derivation of other mouse recombinant inbred populations. Here, we report genome-wide allele frequencies in 347 extinct CC lines. Contrary to expectations, autosomes had equal allelic contributions from the eight founders, but chromosome X had significantly lower allelic contributions from the two inbred founders with underrepresented subspecific origins (PWK/PhJ and CAST/EiJ). By comparing extinct CC lines to living CC strains, we conclude that a complex genetic architecture is driving extinction, and selection pressures are different on the autosomes and chromosome X Male infertility played a large role in extinction as 47% of extinct lines had males that were infertile. Males from extinct lines had high variability in reproductive organ size, low sperm counts, low sperm motility, and a high rate of vacuolization of seminiferous tubules. We performed QTL mapping and identified nine genomic regions associated with male fertility and reproductive phenotypes. Many of the allelic effects in the QTL were driven by the two founders with underrepresented subspecific origins, including a QTL on chromosome X for infertility that was driven by the PWK/PhJ haplotype. We also performed the first example of cross validation using complementary CC resources to verify the effect of sperm curvilinear velocity from the PWK/PhJ haplotype on chromosome 2 in an independent population across multiple generations. While selection typically constrains the examination of reproductive traits toward the more fertile alleles, the CC extinct lines provided a unique opportunity to study the genetic architecture of fertility in a widely genetically variable population. We hypothesize that incompatibilities between alleles with different subspecific origins is a key driver of infertility. These results help clarify the factors that drove strain extinction in the CC, reveal the genetic regions associated with poor fertility in the CC, and serve as a resource to further study mammalian infertility.

Genetic Background, Maternal Age, and Interaction Effects Mediate Rates of Crossing Over in Drosophila melanogaster Females.

Meiotic recombination is a genetic process that is critical for proper chromosome segregation in many organisms. Despite being fundamental for organismal fitness, rates of crossing over vary greatly between taxa. Both genetic and environmental factors contribute to phenotypic variation in crossover frequency, as do genotype-environment interactions. Here, we test the hypothesis that maternal age influences rates of crossing over in a genotypic-specific manner. Using classical genetic techniques, we estimated rates of crossing over for individual Drosophila melanogaster females from five strains over their lifetime from a single mating event. We find that both age and genetic background significantly contribute to observed variation in recombination frequency, as do genotype-age interactions. We further find differences in the effect of age on recombination frequency in the two genomic regions surveyed. Our results highlight the complexity of recombination rate variation and reveal a new role of genotype by maternal age interactions in mediating recombination rate.