Genetic Differences between 129S Substrains Affect Antiretroviral Immune Responses.

Inbred mouse lines vary in their ability to mount protective antiretroviral immune responses, and even closely related strains can exhibit opposing phenotypes upon retroviral infection. Here, we found that 129S mice inherit a previously unknown mechanism for the production of anti-murine leukemia virus (MLV) antibodies and control of infection. The resistant phenotype in 129S1 mice is controlled by two dominant loci that are independent from known MLV resistance genes. We also show that production of anti-MLV antibodies in 129S7 mice, but not 129S1 mice, is independent of interferon gamma signaling. Thus, our data indicate that 129S mice inherit an unknown mechanism for control of MLV infection and demonstrate that there is genetic variability in 129S substrains that affects their ability to mount antiviral immune responses. IMPORTANCE Understanding the genetic basis for production of protective antiviral immune responses is crucial for the development of novel vaccines and adjuvants. Additionally, characterizing the genetic and phenotypic variability in inbred mice has implications for the selection of strains for targeted mutagenesis, choice of controls, and for broader understanding of the requirements for protective immunity.

Utilizing preclinical models of genetic diversity to improve translation of phytochemical activities from rodents to humans and inform personalized nutrition.

Mouse models are an essential tool in different areas of research, including nutrition and phytochemical research. Traditional inbred mouse models have allowed the discovery of therapeutical targets and mechanisms of action and expanded our knowledge of health and disease. However, these models lack the genetic variability typically found in human populations, which hinders the translatability of the results found in mice to humans. The development of genetically diverse mouse models, such as the collaborative cross (CC) or the diversity outbred (DO) models, has been a useful tool to overcome this obstacle in many fields, such as cancer, immunology and toxicology. However, these tools have not yet been widely adopted in the field of phytochemical research. As demonstrated in other disciplines, use of CC and DO models has the potential to provide invaluable insights for translation of phytochemicals from rodents to humans, which are desperately needed given the challenges and numerous failed clinical trials in this field. These models may prove informative for personalized use of phytochemicals in humans, including: predicting interindividual variability in phytochemical bioavailability and efficacy, identifying genetic loci or genes governing response to phytochemicals, identifying phytochemical mechanisms of action and therapeutic targets, and understanding the impact of genetic variability on individual response to phytochemicals. Such insights would prove invaluable for personalized implementation of phytochemicals in humans. This review will focus on the current work performed with genetically diverse mouse populations, and the research opportunities and advantages that these models can offer to phytochemical research.

McH-lpr/lpr-RA1 mice: A novel spontaneous mouse model of autoimmune sialadenitis.

Many studies of the autoimmune disease Sjögren's syndrome have been performed using spontaneous mouse models. In the present study, we describe the characteristics of McH/lpr-RA1 mice and propose their use as a novel murine model of autoimmune sialadenitis. The McH/lpr-RA1 mouse is a recombinant congenic strain derived from generation F54 or more of MRL-Faslpr x (MRL- Faslpr x C3H- Faslpr) F1. We show for the first time that this mouse spontaneously develops autoimmune sialadenitis and vasculitis in submandibular gland tissues. Sialadenitis was accompanied by extensive inflammatory cell infiltration and tissue destruction. Immunohistochemical studies revealed that the salivary gland lesions strongly expressed four sialadenitis-related molecules: SSA and SSB (autoantigens of Sjögren's syndrome), gp91phox (an accelerator of reactive oxygen species production) and single strand DNA (a marker of apoptotic cells). In contrast, expression of aquaporin-5 (AQP5), which stimulates salivary secretion was weak or negligible. Statistical correlation analyses indicated that the apoptosis of salivary gland cells provoked by oxidative stress contributed to the severe sialadenitis and reduced expression of AQP5. Our study has demonstrated that McH/lpr-RA1 mice spontaneously develop the pathognomonic features of autoimmune sialadenitis and thus could be used as a new animal model of Sjögren's syndrome.

53BP1 Repair Kinetics for Prediction of In Vivo Radiation Susceptibility in 15 Mouse Strains.

We present a novel mathematical formalism to predict the kinetics of DNA damage repair after exposure to both low- and high-LET radiation (X rays; 350 MeV/n 40Ar; 600 MeV/n 56Fe). Our method is based on monitoring DNA damage repair protein 53BP1 that forms radiation-induced foci (RIF) at locations of DNA double-strand breaks (DSB) in the nucleus and comparing its expression in primary skin fibroblasts isolated from 15 mice strains. We previously reported strong evidence for clustering of nearby DSB into single repair units as opposed to the classic "contact-first" model where DSB are considered immobile. Here we apply this clustering model to evaluate the number of remaining RIF over time. We also show that the newly introduced kinetic metrics can be used as surrogate biomarkers for in vivo radiation toxicity, with potential applications in radiotherapy and human space exploration. In particular, we observed an association between the characteristic time constant of RIF repair measured in vitro and survival levels of immune cells collected from irradiated mice. Moreover, the speed of DNA damage repair correlated not only with radiation-induced cellular survival in vivo, but also with spontaneous cancer incidence data collected from the Mouse Tumor Biology database, suggesting a relationship between the efficiency of DSB repair after irradiation and cancer risk.

Differential manifestation of ocular phenotypes in TALEN-mediated p19arf knockout FVB/N and C57BL/6J mouse lines.

BACKGROUND: p19arf, primarily known as a tumor suppressor, has also been reported to play an essential role in normal development of mouse eyes. Consistently, lack of p19arf has been associated with ocular defects, but the mixed background of the knockout (KO) mouse strain used raised a concern on the accuracy of the phenotypes observed in association with the targeted gene due to genetic heterogeneity. OBJECT: We carried out a study to investigate into the effect of genetic background on the manifestation of p19arf KO associated phenotypes. METHODS: We characterized the phenotypes of novel p19arf KO mouse lines generated in FVB/N and C57BL/6J using a transcription activator-like effector nuclease (TALEN) system in comparison to the reported phenotypes of three other p19arf-deficient mouse lines generated using homologous recombination. RESULTS: Ninety-five percent of FVB/N-p19arf KO mice showed ocular opacity from week 4 after birth which worsened rapidly until week 6, while such abnormality was absent in C57BL/6J-p19arf KO mice up to the age of 26 weeks. Histopathological analysis revealed retrolental masses and dysplasia in the retinal layer in FVB/N-p19arf KO mice from week 4. Besides these, both strains developed normally from birth to week 26 without increased tumorigenesis except for a subcutaneous tumor found in a C57BL/6J-p19arf KO mouse. CONCLUSION: Our findings demonstrated surprisingly variable manifestation of p19arf-linked phenotypes between FVB/N and C57BL/6J mice, and furthermore between our mouse lines and the established lines, indicating a critical impact of genetic background on functional study of genes using gene targeting strategies in mice.

Mitochondrial AIF loss causes metabolic reprogramming, caspase-independent cell death blockade, embryonic lethality, and perinatal hydrocephalus.

OBJECTIVES: Apoptosis-Inducing Factor (AIF) is a protein involved in mitochondrial electron transport chain assembly/stability and programmed cell death. The relevant role of this protein is underlined because mutations altering mitochondrial AIF properties result in acute pediatric mitochondriopathies and tumor metastasis. By generating an original AIF-deficient mouse strain, this study attempted to analyze, in a single paradigm, the cellular and developmental metabolic consequences of AIF loss and the subsequent oxidative phosphorylation (OXPHOS) dysfunction. METHODS: We developed a novel AIF-deficient mouse strain and assessed, using molecular and cell biology approaches, the cellular, embryonic, and adult mice phenotypic alterations. Additionally, we conducted ex vivo assays with primary and immortalized AIF knockout mouse embryonic fibroblasts (MEFs) to establish the cell death characteristics and the metabolic adaptive responses provoked by the mitochondrial electron transport chain (ETC) breakdown. RESULTS: AIF deficiency destabilized mitochondrial ETC and provoked supercomplex disorganization, mitochondrial transmembrane potential loss, and high generation of mitochondrial reactive oxygen species (ROS). AIF-/Y MEFs counterbalanced these OXPHOS alterations by mitochondrial network reorganization and a metabolic reprogramming toward anaerobic glycolysis illustrated by the AMPK phosphorylation at Thr172, the overexpression of the glucose transporter GLUT-4, the subsequent enhancement of glucose uptake, and the anaerobic lactate generation. A late phenotype was characterized by the activation of P53/P21-mediated senescence. Notably, approximately 2% of AIF-/Y MEFs diminished both mitochondrial mass and ROS levels and spontaneously proliferated. These cycling AIF-/Y MEFs were resistant to caspase-independent cell death inducers. The AIF-deficient mouse strain was embryonic lethal between E11.5 and E13.5 with energy loss, proliferation arrest, and increased apoptotic levels. Contrary to AIF-/Y MEFs, the AIF KO embryos were unable to reprogram their metabolism toward anaerobic glycolysis. Heterozygous AIF+/- females displayed progressive bone marrow, thymus, and spleen cellular loss. In addition, approximately 10% of AIF+/- females developed perinatal hydrocephaly characterized by brain development impairment, meningeal fibrosis, and medullar hemorrhages; those mice died 5 weeks after birth. AIF+/- with hydrocephaly exhibited loss of ciliated epithelium in the ependymal layer. This phenotype was triggered by the ROS excess. Accordingly, it was possible to diminish the occurrence of hydrocephalus AIF+/- females by supplying dams and newborns with an antioxidant in drinking water. CONCLUSIONS: In a single knockout model and at 3 different levels (cell, embryo, and adult mice) we demonstrated that by controlling the mitochondrial OXPHOS/metabolism, AIF is a key factor regulating cell differentiation and fate. Additionally, by providing new insights into the pathological consequences of mitochondrial OXPHOS dysfunction, our new findings pave the way for novel pharmacological strategies.

RNA-seq analysis of chondrocyte transcriptome reveals genetic heterogeneity in LG/J and SM/J murine strains.

OBJECTIVE: To investigate the transcriptomic differences in chondrocytes obtained from LG/J (large, healer) and SM/J (small, non-healer) murine strains in an attempt to discern the molecular pathways implicated in cartilage regeneration and susceptibility to osteoarthritis (OA). DESIGN: We performed RNA-sequencing on chondrocytes derived from LG/J (n = 16) and SM/J (n = 16) mice. We validated the expression of candidate genes and compared single nucleotide polymorphisms (SNPs) between the two mouse strains. We also examined gene expression of positional candidates for ear pinna regeneration and long bone length quantitative trait loci (QTLs) that display differences in cartilaginous expression. RESULTS: We observed a distinct genetic heterogeneity between cells derived from LG/J and SM/J mouse strains. We found that gene ontologies representing cell development, cartilage condensation, and regulation of cell differentiation were enriched in LG/J chondrocytes. In contrast, gene ontologies enriched in the SM/J chondrocytes were mainly related to inflammation and degeneration. Moreover, SNP analysis revealed that multiple validated genes vary in sequence between LG/J and SM/J in coding and highly conserved noncoding regions. Finally, we showed that most QTLs have 20-30% of their positional candidates displaying differential expression between the two mouse strains. CONCLUSIONS: While the enrichment of pathways related to cell differentiation, cartilage development and cartilage condensation infers superior healing potential of LG/J strain, the enrichment of pathways related to cytokine production, immune cell activation and inflammation entails greater susceptibility of SM/J strain to OA. These data provide novel insights into chondrocyte transcriptome and aid in identification of the quantitative trait genes and molecular differences underlying the phenotypic differences associated with individual QTLs.

A New Polygenic Model for Nonfamilial Colorectal Cancer Inheritance Based on the Genetic Architecture of the Azoxymethane-Induced Mouse Model.

The azoxymethane model of colorectal cancer (CRC) was used to gain insights into the genetic heterogeneity of nonfamilial CRC. We observed significant differences in susceptibility parameters across 40 mouse inbred strains, with 6 new and 18 of 24 previously identified mouse CRC modifier alleles detected using genome-wide association analysis. Tumor incidence varied in F1 as well as intercrosses and backcrosses between resistant and susceptible strains. Analysis of inheritance patterns indicates that resistance to CRC development is inherited as a dominant characteristic genome-wide, and that susceptibility appears to occur in individuals lacking a large-effect, or sufficient numbers of small-effect, polygenic resistance alleles. Our results suggest a new polygenic model for inheritance of nonfamilial CRC, and that genetic studies in humans aimed at identifying individuals with elevated susceptibility should be pursued through the lens of absence of dominant resistance alleles rather than for the presence of susceptibility alleles.

Strains matter: Success of murine in vitro spermatogenesis is dependent on genetic background.

The current strategy to preserve fertility of male prepubertal cancer patients consists of cryopreservation of a testicular tissue biopsy containing spermatogonial stem cells (SSCs). While in humans, fertility restoration strategies from prepubertal testicular tissues are still under investigation and have not yet resulted in complete germ cell differentiation, in mice various studies have described production of sperm and offspring through testicular organ culture and transplantation of in vitro propagated SSCs. Organ culture has shown to be successful in generating mature spermatozoa when using testicular fragments from various mouse strains, including CD1 and C57BL/6 J. Conversely, in vitro proliferation of SSCs from C57BL/6 J mice is highly inefficient when compared to other strains such as DBA2 or hybrid mice of C57BL/6 J and DBA2 with 75% C57BL/6 J background (B6D2F2). In this study, we investigated in vitro spermatogenesis by organ culture using testicular tissue from C57BL/6 J and B6D2F2 mice. Whereas spermatogenesis was initiated and completed in C57BL/6 J fragments, it could not be effectively supported in B6D2F2 testicular tissue. While maturation of Sertoli cells and Leydig cells functionality appeared to be identical between the two strains, in B6D2F2 tissue spermatogenesis did not proceed past the spermatocyte step, followed by a rapid decline of the number of all germ cells in the fragments. This suggests that the spermatogenic potential in vitro is dependent on specialized sites in the genome and therefore the organ culture conditions suboptimal for some strains of mice.

Distribution of endogenous gammaretroviruses and variants of the Fv1 restriction gene in individual mouse strains and strain subgroups.

Inbred laboratory mouse strains carry endogenous retroviruses (ERVs) classed as ecotropic, xenotropic or polytropic mouse leukemia viruses (E-, X- or P-MLVs). Some of these MLV ERVs produce infectious virus and/or contribute to the generation of intersubgroup recombinants. Analyses of selected mouse strains have linked the appearance of MLVs and virus-induced disease to the strain complement of MLV E-ERVs and to host genes that restrict MLVs, particularly Fv1. Here we screened inbred strain DNAs and genome assemblies to describe the distribution patterns of 45 MLV ERVs and Fv1 alleles in 58 classical inbred strains grouped in two ways: by common ancestry to describe ERV inheritance patterns, and by incidence of MLV-associated lymphomagenesis. Each strain carries a unique set of ERVs, and individual ERVs are present in 5-96% of the strains, often showing lineage-specific distributions. Two ERVs are alternatively present as full-length proviruses or solo long terminal repeats. High disease incidence strains carry the permissive Fv1n allele, tested strains have highly expressed E-ERVs and most have the Bxv1 X-ERV; these three features are not present together in any low-moderate disease strain. The P-ERVs previously implicated in P-MLV generation are not preferentially found in high leukemia strains, but the three Fv1 alleles that restrict inbred strain E-MLVs are found only in low-moderate leukemia strains. This dataset helps define the genetic basis of strain differences in spontaneous lymphomagenesis, describes the distribution of MLV ERVs in strains with shared ancestry, and should help annotate sequenced strain genomes for these insertionally polymorphic and functionally important proviruses.

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.

Stress of Strains: Inbred Mice in Liver Research.

Inbred mice are the most popular animals used for in vivo liver research. These mice are genetically defined, readily available, less expensive to maintain than larger animals, and enjoy a broad array of commercial reagents for scientific characterization. C57BL/6 mice are the most commonly used strain. However, other strains discussed, including BALB/c, C3H, A/J, and FVB/N, may be better suited to a particular disease model or line of investigation. Understanding the phenotypes of different inbred mouse strains facilitates informed decision making during experimental design. Model systems influenced by strain-dependent phenotype include tissue regeneration, drug-induced liver injury (DILI; e.g., acetaminophen), fibrosis (e.g., carbon tetrachloride, CCl4), Fas-induced apoptosis, cholestasis, alcohol-induced liver disease and cirrhosis, nonalcoholic fatty liver disease and steatohepatitis (NAFLD/NASH), and hepatocellular carcinoma (HCC). Thoughtful consideration of the strengths and weaknesses of each inbred strain in a given model system will lead to more robust data and a clearer understanding of translational relevance to human liver disease.

Comparative utility of NRG and NRGS mice for the study of normal hematopoiesis, leukemogenesis, and therapeutic response.

Cell-line-derived xenografts (CDXs) or patient-derived xenografts (PDXs) in immune-deficient mice have revolutionized our understanding of normal and malignant human hematopoiesis. Transgenic approaches further improved in vivo hematological research, allowing the development of human-cytokine-producing mice, which show superior human cell engraftment. The most popular mouse strains used in research, the NOG (NOD.Cg-Prkdcscid Il2rγtm1Sug/Jic) and the NSG (NOD/SCID-IL2Rγ-/-, NOD.Cg-PrkdcscidIl2rγtm1Wjl/SzJ) mouse, and their human-cytokine-producing (interleukin-3, granulocyte-macrophage colony-stimulating factor, and stem cell factor) counterparts (huNOG and NSGS), rely partly on a mutation in the DNA repair protein PRKDC, causing a severe combined immune deficiency (SCID) phenotype and rendering the mice less tolerant to DNA-damaging therapeutics, thereby limiting their usefulness in the investigation of novel acute myeloid leukemia (AML) therapeutics. NRG (NOD/RAG1/2-/-IL2Rγ-/-) mice show equivalent immune ablation through a defective recombination activation gene (RAG), leaving DNA damage repair intact, and human-cytokine-producing NRGS (NRG-SGM3) mice were generated, improving myeloid engraftment. Our findings indicate that unconditioned NRG and NRGS mice can harbor established AML CDXs and can tolerate aggressive induction chemotherapy at higher doses than NSG mice without overt toxicity. However, unconditioned NRGS mice developed less clinically relevant disease, with CDXs forming solid tumors throughout the body, whereas unconditioned NRG mice were incapable of efficiently supporting PDX or human hematopoietic stem cell engraftment. These findings emphasize the contextually dependent utility of each of these powerful new strains in the study of normal and malignant human hematopoiesis. Therefore, the choice of mouse strain cannot be random, but must be based on the experimental outcomes and questions to be addressed.

Brain damage resulting from postnatal hypoxic-ischemic brain injury is reduced in C57BL/6J mice as compared to C57BL/6N mice.

Perinatal hypoxia is a critical complication during delivery and is mostly studied in animal models of postnatal hypoxic-ischemic brain injury. We here studied the effects of postnatal hypoxic-ischemic brain injury in two different sub-strains of C57BL/6 mice, i.e. C57BL/6J and C57BL/6N mice. These two sub-strains show different metabolic properties, for instance an impaired glucose tolerance in C57BL/6J mice. Genetically, this was linked to differences in their nicotinamide nucleotide transhydrogenase (Nnt) genes: In C57BL/6J mice, exons 7-11 of the Nnt gene are deleted, resulting in the absence of functional Nnt protein. The mitochondrial Nnt-protein is one of several enzymes that catalyses the generation of NADPH, which in turn is important for the elimination of reactive oxygen species (ROS). As ROS is thought to contribute to the pathophysiology of hypoxia-ischemia, the lack of Nnt might indirectly increase ROS levels and therefore result in increased brain damage. We therefore hypothesize that lesion score and lesion size will increase in C57BL/6J mice as compared to C57BL/6N mice. Surprisingly, the results showed exactly the opposite: C57BL/6J mice showed a decrease in lesion score and size, associated with a reduced number of apoptotic cells and activated microglia. In contrast, the number of cells with ROS-induced DNA modifications (detected by 8OHdG) was higher in C57BL/6J than C57BL/6N mice. In conclusion, C57BL/6J mice showed reduced ischemic consequences after postnatal hypoxic-ischemic brain injury compared to C57BL/6N mice, with the exception of the amount of ROS-induced DNA-damage. These differences might relate to the lack of Nnt, but also to a modified metabolic setting (cardiovascular parameters, oxygen and glucose metabolism, immune function) in C57BL/6J mice.

Deep genome sequencing and variation analysis of 13 inbred mouse strains defines candidate phenotypic alleles, private variation and homozygous truncating mutations.

BACKGROUND: The Mouse Genomes Project is an ongoing collaborative effort to sequence the genomes of the common laboratory mouse strains. In 2011, the initial analysis of sequence variation across 17 strains found 56.7 M unique single nucleotide polymorphisms (SNPs) and 8.8 M indels. We carry out deep sequencing of 13 additional inbred strains (BUB/BnJ, C57BL/10J, C57BR/cdJ, C58/J, DBA/1J, I/LnJ, KK/HiJ, MOLF/EiJ, NZB/B1NJ, NZW/LacJ, RF/J, SEA/GnJ and ST/bJ), cataloguing molecular variation within and across the strains. These strains include important models for immune response, leukaemia, age-related hearing loss and rheumatoid arthritis. We now have several examples of fully sequenced closely related strains that are divergent for several disease phenotypes. RESULTS: Approximately 27.4 M unique SNPs and 5 M indels are identified across these strains compared to the C57BL/6 J reference genome (GRCm38). The amount of variation found in the inbred laboratory mouse genome has increased to 71 M SNPs and 12 M indels. We investigate the genetic basis of highly penetrant cancer susceptibility in RF/J finding private novel missense mutations in DNA damage repair and highly cancer associated genes. We use two highly related strains (DBA/1J and DBA/2J) to investigate the genetic basis of collagen-induced arthritis susceptibility. CONCLUSIONS: This paper significantly expands the catalogue of fully sequenced laboratory mouse strains and now contains several examples of highly genetically similar strains with divergent phenotypes. We show how studying private missense mutations can lead to insights into the genetic mechanism for a highly penetrant phenotype.

Efficient analysis of mouse genome sequences reveal many nonsense variants.

Genetic polymorphisms in coding genes play an important role when using mouse inbred strains as research models. They have been shown to influence research results, explain phenotypical differences between inbred strains, and increase the amount of interesting gene variants present in the many available inbred lines. SPRET/Ei is an inbred strain derived from Mus spretus that has ∼1% sequence difference with the C57BL/6J reference genome. We obtained a listing of all SNPs and insertions/deletions (indels) present in SPRET/Ei from the Mouse Genomes Project (Wellcome Trust Sanger Institute) and processed these data to obtain an overview of all transcripts having nonsynonymous coding sequence variants. We identified 8,883 unique variants affecting 10,096 different transcripts from 6,328 protein-coding genes, which is about 28% of all coding genes. Because only a subset of these variants results in drastic changes in proteins, we focused on variations that are nonsense mutations that ultimately resulted in a gain of a stop codon. These genes were identified by in silico changing the C57BL/6J coding sequences to the SPRET/Ei sequences, converting them to amino acid (AA) sequences, and comparing the AA sequences. All variants and transcripts affected were also stored in a database, which can be browsed using a SPRET/Ei M. spretus variants web tool (www.spretus.org), including a manual. We validated the tool by demonstrating the loss of function of three proteins predicted to be severely truncated, namely Fas, IRAK2, and IFNγR1.

Approaches to Investigating Complex Genetic Traits in a Large-Scale Inbred Mouse Aging Study.

Inbred mice are a unique model system for studying aging because of the genetic homogeneity within inbred strains, the short life span of mice relative to humans, and the rich array of analytic tools that are available. A large-scale aging study was conducted on 28 inbred strains representing great genetic diversity to determine, via histopathology, the type and diversity of spontaneous diseases that aging mice develop. A total of 20 885 different diagnoses were made, with an average of 12 diagnoses per mouse in the study. Eighteen inbred strains have had their genomes sequenced, and many others have been partially sequenced to provide large repositories of data on genetic variation among the strains. This vast amount of genomic information can be utilized in genome-wide association studies to find candidate genes that are involved in the pathogenesis of spontaneous diseases. As an illustration, this article presents a genome-wide association study of the genetic associations of age-related intestinal amyloidosis, which implicated 3 candidate genes: translocating chain-associated membrane protein 1 (Tram1); splicing factor 3b, subunit 5 (Sf3b5); and syntaxin 11 (Stx11). Representative photomicrographs are available on the Mouse Tumor Biology Database and Pathbase to serve as a reference when evaluating inbred mice used in other genetic or experimental studies to rule out strain background lesions. Many of the age-related mouse diseases are similar, if not identical, to human diseases; therefore, the genetic discoveries have direct translational benefit.

Using whole-genome sequences of the LG/J and SM/J inbred mouse strains to prioritize quantitative trait genes and nucleotides.

BACKGROUND: The laboratory mouse is the most commonly used model for studying variation in complex traits relevant to human disease. Here we present the whole-genome sequences of two inbred strains, LG/J and SM/J, which are frequently used to study variation in complex traits as diverse as aging, bone-growth, adiposity, maternal behavior, and methamphetamine sensitivity. RESULTS: We identified small nucleotide variants (SNVs) and structural variants (SVs) in the LG/J and SM/J strains relative to the reference genome and discovered novel variants in these two strains by comparing their sequences to other mouse genomes. We find that 39% of the LG/J and SM/J genomes are identical-by-descent (IBD). We characterized amino-acid changing mutations using three algorithms: LRT, PolyPhen-2 and SIFT. We also identified polymorphisms between LG/J and SM/J that fall in regulatory regions and highly informative transcription factor binding sites (TFBS). We intersected these functional predictions with quantitative trait loci (QTL) mapped in advanced intercrosses of these two strains. We find that QTL are both over-represented in non-IBD regions and highly enriched for variants predicted to have a functional impact. Variants in QTL associated with metabolic (231 QTL identified in an F16 generation) and developmental (41 QTL identified in an F34 generation) traits were interrogated and we highlight candidate quantitative trait genes (QTG) and nucleotides (QTN) in a QTL on chr13 associated with variation in basal glucose levels and in a QTL on chr6 associated with variation in tibia length. CONCLUSIONS: We show how integrating genomic sequence with QTL reduces the QTL search space and helps researchers prioritize candidate genes and nucleotides for experimental follow-up. Additionally, given the LG/J and SM/J phylogenetic context among inbred strains, these data contribute important information to the genomic landscape of the laboratory mouse.

CD201 and CD27 identify hematopoietic stem and progenitor cells across multiple murine strains independently of Kit and Sca-1.

Identification and isolation of hematopoietic stem cells (HSCs) in mice is most commonly based on the expression of surface molecules Kit and Sca-1 and the absence of markers of mature lineages. However, Sca-1 is absent or weakly expressed in hematopoietic progenitors in many strains, including nonobese diabetic (NOD), BALB/c, C3H, and CBA mice. In addition, both Kit and Sca-1 levels are modulated following bone marrow injury. In these cases, other markers and dye exclusion methods have been employed to identify HSCs, yet there is no antibody-based stain that enables identification of HSCs and early progenitors when Kit and Sca-1 are inadequate. CD201 is a marker that is highly restricted to HSCs and progenitors, and CD27 is expressed at moderate-to-high levels on HSCs. We show here that combining CD201 and CD27 enables highly efficient isolation of long-term HSCs in NOD mice as well as in other strains, including SJL, FVB, AKR, BALB/c, C3H, and CBA. We also find that HSCs appear to maintain expression of CD201 and CD27 after hematopoietic injury when Kit expression is downregulated. These results suggest a widely applicable yet simple alternative for HSC isolation in settings where Kit and Sca-1 expression are insufficient.

Maternal-foetal genomic conflict and speciation: no evidence for hybrid placental dysplasia in crosses between two house mouse subspecies.

Interspecific hybridization between closely related mammalian species, including various species of the genus Mus, is commonly associated with abnormal growth of the placenta and hybrid foetuses, a phenomenon known as hybrid placental dysplasia (HPD). The role of HPD in speciation is anticipated but still poorly understood. Here, we studied placental and foetal growth in F1 crosses between four inbred mouse strains derived from two house mouse subspecies, Mus musculus musculus and Mus musculus domesticus. These subspecies are in the early stage of speciation and still hybridize in nature. In accordance with the maternal-foetal genomic conflict hypothesis, we found different parental influences on placental and foetal development, with placental weight most affected by the father's body weight and foetal weight by the mother's body weight. After removing the effects of parents' body weight, we did not find any significant differences in foetal or placental weights between intra-subspecific and inter-subspecific F1 crosses. Nevertheless, we found that the variability in placental weight in inter-subspecific crosses is linked to the X chromosome, similarly as for HPD in interspecific mouse crosses. Our results suggest that maternal-foetal genomic conflict occurs in the house mouse system, but has not yet diverged sufficiently to cause abnormalities in placental and foetal growth in inter-subspecific crosses. HPD is thus unlikely to contribute to speciation in the house mouse system. However, we cannot rule out that it might have contributed to other speciation events in the genus Mus, where differences in the levels of polyandry exist between the species.