Chromosome Substitution Strains Research Articles

Genetic Factors on Mouse Chromosome 18 Affecting Susceptibility to Testicular Germ Cell Tumors and Permissiveness to Embryonic Stem Cell Derivation

Despite strong heritability, little is known about the genetic control of susceptibility to testicular germ cell tumors (TGCT) in humans or mice. Although the mouse model of spontaneous TGCTs has been extensively studied, conventional linkage analysis has failed to locate the factors that control teratocarcinogenesis in the susceptible 129 family of inbred strains. As an alternative approach, we used both chromosome substitution strains (CSS) to identify individual chromosomes that harbor susceptibility genes and a panel of congenic strains derived from a selected CSS to determine the number and location of susceptibility variants on the substituted chromosome. We showed that 129-Chr 18(MOLF) males are resistant to spontaneous TGCTs and that at least four genetic variants control susceptibility in males with this substituted chromosome. In addition, early embryonic cells from this strain fail to establish embryonic stem cell lines as efficiently as those from the parental 129/Sv strain. For the first time, 129-derived genetic variants that control TGCT susceptibility and fundamental aspects of embryonic stem cell biology have been localized in a genetic context in which the genes can be identified and functionally characterized.

Chromosomal mapping of excessive physical activity in mice in response to a restricted feeding schedule

Excessive physical activity plays an important role in the progression of anorexia nervosa (AN) by accelerating weight loss during dietary restriction. To search for mechanisms underlying this trait, a panel of mouse chromosome substitution strains derived from C57BL/6J and A/J strains was exposed to a scheduled feeding paradigm and to voluntary running wheel (RW) access. Here, we showed that A/J chromosomes 4, 12 and 13 contribute to the development of a disrupted RW activity in response to daily restricted feeding. This pattern is characterized by intense RW activity during the habitual rest phase and leads to accelerated body weight loss. Regions on mouse chromosomes 4, 12 and 13 display homology with regions on human chromosomes linked with anxiety and obsessionality in AN cohorts. Therefore, our data open new roads for interspecies genetic studies of AN and for unraveling novel mechanisms and potential effective treatment strategies for these neurobehavioral traits.

Genomic survey of prepulse inhibition in mouse chromosome substitution strains

Prepulse inhibition (PPI) is a measure of sensorimotor gating, a pre-attentional inhibitory brain mechanism that filters extraneous stimuli. Prepulse inhibition is correlated with measures of cognition and executive functioning, and is considered an endophenotype of schizophrenia and other psychiatric illnesses in which patients show PPI impairments. As a first step toward identifying genes that regulate PPI, we performed a quantitative trait locus (QTL) screen of PPI phenotypes in a panel of mouse chromosome substitution strains (CSSs). We identified five CSSs with altered PPI compared with the host C57BL/6J strain: CSS-4 exhibited decreased PPI, whereas CSS-10, -11, -16 and -Y exhibited higher PPI compared with C57BL/6J. These data indicate that A/J chromosomes 4, 10, 11, 16 and Y harbor at least one QTL region that modulates PPI in these CSSs. Quantitative trait loci for the acoustic startle response were identified on seven chromosomes. Like PPI, habituation of the startle response is also disrupted in schizophrenia, and in the present study CSS-7 and -8 exhibited deficits in startle habituation. Linkage analysis of an F(2) intercross identified a highly significant QTL for PPI on chromosome 11 between positions 101.5 and 114.4 Mb (peak LOD = 4.54). Future studies will map the specific genes contributing to these QTLs using congenic strains and other genomic approaches. Identification of genes that modulate PPI will provide insight into the neural mechanisms underlying sensorimotor gating, as well as the psychopathology of disorders characterized by gating deficits.

A major QTL on chromosome 11 influences psychostimulant and opioid sensitivity in mice

The identification of genes influencing sensitivity to stimulants and opioids is important for determining their mechanism of action and may provide fundamental insights into the genetics of drug abuse. We used a panel of C57BL/6J (B6; recipient)x A/J (donor) chromosome substitution strains (CSSs) to identify quantitative trait loci (QTL) for both open field activity and sensitivity to the locomotor stimulant response to methamphetamine (MA). Mice were injected with saline (days 1 and 2) and MA (day 3; 2 mg/kg i.p.). We analyzed the total distance traveled in the open field for 30 min following each injection. CSS-8, -11 and -16 showed reduced MA-induced locomotor activity relative to B6, whereas CSS-10 and -12 showed increased MA-induced locomotor activity. Further analysis focused on CSS-11 because it was robustly different from B6 following MA injection, but did not differ in activity following saline injection and because it also showed reduced locomotor activity in response to the mu-opioid receptor agonist fentanyl (0.2 mg/kg i.p.). Thus, CSS-11 captures QTLs for the response to both psychostimulants and opioids. Using a B6 x CSS-11 F(2) intercross, we identified a dominant QTL for the MA response on chromosome 11. We used haplotype association mapping of cis expression QTLs and bioinformatic resources to parse among genes within the 95% confidence interval of the chromosome 11 QTL. Identification of the genes underlying QTLs for response to psychostimulants and opioids may provide insights about genetic factors that modulate sensitivity to drugs of abuse.

A Locus Mapping to Mouse Chromosome 7 Determines Infarct Volume in a Mouse Model of Ischemic Stroke

In a mouse model of focal cerebral ischemia, infarct volume is highly variable and strain dependent, but the natural genetic determinants responsible for this difference remain unknown. To identify genetic determinants regulating ischemic neuronal damage and to dissect apart the role of individual genes and physiological mechanisms in infarction in mice, we performed quantitative trait locus analysis of surgically induced cerebral infarct volume. After permanent occlusion of the distal middle cerebral artery, infarct volume was determined for 16 inbred strains of mice, chromosome substitution strains, and for 2 intercross cohorts, F2 (B6xBALB/c) and F2 (B6xSWR/J). Genome-wide linkage analysis was performed for infarct volume as a quantitative trait. Infarct volume varied up to 30-fold between strains, with heritability estimated at 0.88. Overall, 3 quantitative trait locus were identified that modulate infarct volume, with a major locus (Civq1) on chromosome 7 accounting for >50% of the variation, with a combined LOD score of 21.7. Interval-specific single nucleotide polymorphism haplotype analysis for Civq1 results in 12 candidate genes. The extent of ischemic tissue damage after distal middle cerebral artery occlusion in inbred strains of mice is modulated by genetic variation mapping to at least 3 different loci. A single locus on chromosome 7 determines the majority of the observed variation in the trait. This locus seems to be identical to LSq1, a locus conferring limb salvage and reperfusion in a mouse model of hindlimb ischemia. The identification of the genes underlying these loci may uncover novel genetic and physiological pathways that modulate cerebral infarction and provide new targets for therapeutic intervention in ischemic stroke, and possibly other ischemic diseases.

A verification of previously identified QTLs for cocaine-induced activation using a panel of B6.A chromosome substitution strains (CSS) and A/J x C57Bl/6J F2 mice

The objective of this study was to confirm provisional quantitative trait loci (QTL) for cocaine-induced locomotor activation, on chromosomes 1, 5, 6, 9, 12, 15, 16, 17, and 18, previously identified in the AXB/BXA recombinant inbred (RI) and AcB/BcA recombinant congenic (RC) strains of mice derived from A/J (A) and C57BL/6J (B6) progenitors. This was accomplished through a genetic analysis of cocaine-induced activity in an AxB6 F2 cross and a phenotypic survey across a panel of B6.A chromosome substitution strains (CSS) mice. Mice were tested for cocaine-induced activity, following administration of saline and cocaine (20 mg/kg), utilizing an open-field procedure. Among AxB6 F2 mice, differences in cocaine-induced activity were associated with loci on chromosome 1 (D1Mit305), 5 (D5Mit409), 16 (D16Mit131), and 18 (D18Mit189). A survey of the CSS panel confirmed cocaine QTLs on chromosomes 5 and 15, previously identified in RI or RC strains. Overall, the regions on chromosomes 5 and 18 represent verification of QTL previously identified in both the RC and RI strains. Additionally, the B6 allele for these QTL was consistently associated with greater relative cocaine activation. Collectively, chromosome 5 and 18 QTL have now been replicated in multiple independent crosses derived from the A/J and C57BL/6J progenitors. The use of an in silico analysis highlighted potential candidate genes on chromosomes 5 and 18. The present results complement the targeted gene approach currently prevalent in the study of cocaine and provide a broader empirically based focus for subsequent candidate gene studies.

Variations in ventral root axon morphology and locomotor behavior components across different inbred strains of mice

Locomotion is a complex behavior affected by many different brain- and spinal cord systems, as well as by variations in the peripheral nervous system. Recently, we found increased gene expression for EphA4, a gene intricately involved in motor neuron development, between high-active parental strain C57BL/6J and the low-active chromosome substitution strain 1 (CSS1). CSS1 mice carry chromosome 1 from A/J mice in a C57BL/6J genetic background, allowing localization of quantitative trait loci (QTL) on chromosome 1. To find out whether differences in motor neuron anatomy, possibly related to the changes in EphA4 expression, are involved in the motor activity differences observed in these strains, motor performance in various behavioral paradigms and anatomical differences in the ventral roots were investigated. To correlate the behavioral profiles to the spinal motor neuron morphology, not only CSS1 and its parental strains C57BL/6J (host) and A/J (donor) were examined, but also a set of other mouse inbred strains (AKR/J, 129×1/SvJ and DBA/2J). Significant differences were found between inbred strains on home cage motor activity levels, the beam balance test, grip test performance, and on alternating versus synchronous hind limb movement (hind limb hopping). Also, considerable differences were found in spinal motor neuron morphology, with A/J and CSS1 showing smaller, possibly less developed, motor neuron axons compared to all other inbred strains. For CSS1 and C57BL/6J, only genetically different for chromosome 1, a correlation was found between motor activity levels, synchronous hind limb movement and neuro-anatomical differences in spinal motor neurons. Inclusion of the other inbred strains, however, did not show this direct correlation. These data verifies the complex nature of the mammalian motor system that may be further dissected using genetic mapping populations derived from these inbred strains.

Epistatic interactions govern chemically‐induced lung tumor susceptibility and Kras mutation site in murine C57BL/6J‐ChrA/J chromosome substitution strains

Cancer susceptibility results from interactions between sensitivity and resistance alleles. We employed murine chromosome substitution strains to study how resistance alleles affected sensitive alleles during chemically-induced lung carcinogenesis. The C57BL/6J-Chr#(A/J) strains, constructed by selectively breeding sensitive A/J and resistant C57BL/6J (B6) mice, each contain one pair of A/J chromosomes within an otherwise B6 genome. Pas1, the major locus responsible for this differential strain response to urethane carcinogenesis, resides on Chr 6, but C57BL/6J-Chr6(A/J) mice (hereafter CSS-6) developed few tumors following a single urethane injection, which demonstrates epistatic interactions with other B6 alleles. CSS6 mice developed dozens of lung tumors after chronic urethane exposure, however, indicating that these epistatic interactions could be overcome by repeated carcinogen administration. Unlike A/J, but similar to B6 mice, CSS6 mice were resistant to lung carcinogenesis induced by 3-methylcholanthrene (MCA). Tumor multiplicity increased if BHT administration followed urethane exposure, showing that a Chr 6 gene(s) regulates sensitivity to chemically-induced tumor promotion. Unlike A/J tumors (predominantly codon 61 A-->T transversions), Kras mutations in tumors induced by urethane in CSS-6 mice were similar to B6 tumors (codon 61 A-->G transitions). DNA repair genes not located on Chr 6 may determine the nature of Kras mutations. CSS-6 mice are a valuable resource for testing the ability of candidate genes to modulate lung carcinogenesis.

A grandparent-influenced locus for alcohol preference on mouse chromosome 2

Loci on mouse chromosome 2 have previously been associated with ethanol consumption. Here, we used a limited access choice paradigm in which mice consume large quantities of ethanol (2-3 g/kg/2 h) with a high preference (>80%). In addition, mouse chromosome substitution strains were used to further evaluate the contribution of chromosome 2 to ethanol consumption. First, we compared the two parental inbred mouse strains, C57BL/6J and A/J, in the limited access choice paradigm for ethanol intake and ethanol preference, as well as for ethanol metabolism and taste sensitivity. Then, the effect of chromosome 2 substitution on these measures was determined. Compared with C57BL/6J mice, A/J and C57BL/6J-Chr 2/NaJ (CSS-2) mice showed profoundly reduced ethanol intake and preference. The strains were not different with regard to ethanol metabolism or taste sensitivity. Limited access ethanol consumption in F2 progeny derived from reciprocal C57BL/6J xCSS-2 and CSS-2 xC57BL/6J intercrosses and subsequent quantitative trait loci mapping identified two loci: one locus on chromosome 2 for ethanol intake and a separate locus on distal chromosome 2 for ethanol preference. This latter locus was dependent on the grandparental origin. Using a limited access choice paradigm, we found that mouse chromosome 2 carries an allelic variant of a locus for ethanol intake and a distinct locus selective for ethanol preference. The heritability of alcoholism has been suggested to be parent-specific, perhaps resulting from genetic imprinting. Our findings suggest that grandparent-influenced vulnerability for ethanol consumption is conferred by genes on chromosome 2, providing important new leads to enhance our understanding of the heritability of alcoholism.

Interspecies Trait Genetics Reveals Association of Adcy8 with Mouse Avoidance Behavior and a Human Mood Disorder

Identifying susceptibility genes for endophenotypes by studying analogous behaviors across species is an important strategy for understanding the pathophysiology underlying psychiatric disorders. This approach provides novel biological pathways plus validated animal models critical for selective drug development. One such endophenotype is avoidance behavior. In the present study, novel automated registration methods for longitudinal behavioral assessment in home cages are used to screen a panel of recently generated mouse chromosome substitution strains that are very powerful in quantitative trait loci (QTL) detection of complex traits. In this way, we identified chromosomes regulating avoidance behavior (increased sheltering preference) independent of motor activity levels (horizontal distance moved). Genetic information from the mouse QTL-interval was integrated with that from the homologous human linkage region for a mood disorder. We genetically mapped a QTL for avoidance behavior on mouse chromosome 15, homologous with a human genome region (8q24) linked to bipolar disorder. Integrating the syntenic mouse QTL-interval with genotypes of 1868 BPD cases versus 14,311 control subjects revealed two associated genes (ADCY8 and KCNQ3). Adenylyl cyclase 8 (Adcy8) was differentially expressed in specific brain regions of mouse strains that differ in avoidance behavior levels. Finally, we showed that chronic infusion of the human mood stabilizer carbamazepine (that acts via adenylyl cyclase activity) significantly reduced mouse avoidance behavior, providing a further link between human mood disorders and this mouse home cage behavior. Our data suggest that Adcy8 might encode a translational behavioral endophenotype of bipolar disorder.

Genetic analysis of daily physical activity using a mouse chromosome substitution strain

There is considerable evidence for a genetic basis underlying individual differences in spontaneous physical activity in humans and animals. Previous publications indicate that the physical activity level and pattern vary among inbred strains of mice and identified a genomic region on chromosome 13 as quantitative trait loci (QTL) for physical activity. To confirm and further characterize the role of chromosome 13 in regulating daily physical activity level and pattern, we conducted a comprehensive phenotypic study in the chromosome 13 substitution strain (CSS-13) in which the individual chromosome 13 from the A/J strain was substituted into an otherwise complete C57BL/6J (B6) genome. The B6 and A/J parental strains exhibited pronounced differences in daily physical activity, sleep-wake structure, circadian period and body weight. Here we report that a single A/J chromosome 13 in the context of a B6 genetic background conferred a profound reduction in both total cage activity and wheel-running activity under a 14:10-h light-dark cycle, as well as in constant darkness, compared with B6 controls. Additionally, CSS-13 mice differed from B6 controls in the diurnal distribution of activity and the day-to-day variability in activity onset. We further performed a linkage analysis and mapped a significant QTL on chromosome 13 regulating the daily wheel running activity level in mice. Taken together, our findings indicate a QTL on chromosome 13 with dramatic and specific effects on daily voluntary physical activity, but not on circadian period, sleep, or other aspects of activity that are different between B6 and A/J strains.

Dissecting the genetic basis of kidney tubule response to hyperoxaluria using chromosome substitution strains

Whether genetics may play a role in the pathophysiologic response of kidney tubules to oxalate exposure remains unexplored despite that as many as 15% of the U.S. population annually will experience a kidney stone composed of calcium oxalate. To explore this issue, we utilized a panel of chromosome substitution strains in which one chromosome at a time was transferred from the Brown Norway (BN) rat onto the Dahl salt-sensitive (SS) genetic background. Hyperoxaluria was induced by adding hydroxyproline (HP) to the drinking water. A dose-response (0-2% HP) study found that both SS and BN exhibited the same level of oxalate excretion as HP concentration increased, but only the BN exhibited changes in urothelial pathology and demonstrated crystal deposition at sites of urothelial injury as a function of dose (at 1.5-2.0%). The consomic panel was treated with 2.0% HP and evaluated for hyperoxaluria, renal injury, and crystal deposition. Tubular injury (% Area) and crystal deposition (% Area) were similar between the resistant SS and SS-4, -6, -7, -8, -9, -11, -16, and -20(BN) consomic rats. However, tubular injury was significantly increased in SS-2(BN) compared with the SS parental (9.8 +/- 1.56 and 4.2 +/- 1.09%, respectively). Crystal deposition was observed in SS-2(BN) and SS-18(BN) (4.7 +/- 0.70 and 3.5 +/- 1.3%, respectively) to the same extent as seen in the susceptible BN (3.2 +/- 0.44%). The fact that crystal deposition was observed in SS-18(BN) without extensive overall tubule injury, compared with the more severe widespread tubular injury seen in SS-2(BN), suggests that the underlying mechanism of each locus is different. In conclusion, these studies establish that BN rats demonstrate oxalate-associated pathology and they retain calcium oxalate crystals coincident with urothelial injury but SS rats do not. These observations establish that BN rat chromosome 2 and 18 harbor genes that contribute to these processes.

The Role of the Mouse Y Chromosome on Susceptibility to Testicular Germ Cell Tumors

Testicular germ cell tumors (TGCT) are sex limited, occurring only in males with a Y chromosome. Recently, the gr/gr deletion on the human Y chromosome was associated with increased risk of TGCTs. In addition, the presence of Y chromosome sequences is associated with TGCTs in cases of gonadal dysgenesis. TGCTs in strain 129 males recapitulate many aspects of testicular cancer in human infants and can be used to evaluate the role of the Y chromosome in TGCT risk. We used chromosome substitution strains and a sex-reversing mutant to test the role of the Y chromosome on TGCT susceptibility. Our results show that a Y-linked gene that does not differ among the tested strains is essential for tumorigenesis.

Chr 19(A/J) modifies tumor resistance in a sex- and parent-of-origin-specific manner.

Neurofibromatosis type 1 (NF1) is one of the most common human genetic diseases affecting the nervous system and predisposes individuals to cancer, including peripheral nerve sheath tumors (PNSTs) and astrocytomas. Modifiers in the genetic background affect the severity of the disease and we have previously mapped two modifier loci, Nstr1 and Nstr2, that influence resistance to PNSTs in the Nf1-/+;Trp53-/+cis mouse model of NF1. We report here the analysis of Nstr1 in isolation from other epistatic loci using a chromosome substitution strain, and further show that a modifier locus (or loci) on chromosome 19 influences resistance to both PNSTs and astrocytomas. This modifier locus interacts with sex, resulting in sex-specific modification of tumors. Allele variability on chromosome 19 affects both the timing and the penetrance of the growth of different tumor types associated with NF1, specifically PNSTs and astrocytoma. These results indicate that modifiers of cancer susceptibility interact and affect tumorigenesis under different genetic conditions and demonstrate the power of chromosome substitution strains to study genetic modifiers.

Phenotyping mouse chromosome substitution strains reveal multiple QTLs for febrile seizure susceptibility

Febrile seizures (FS) are the most common seizure type in children and recurrent FS are a risk factor for developing temporal lobe epilepsy. Although the mechanisms underlying FS are largely unknown, recent family, twin and animal studies indicate that genetics are important in FS susceptibility. Here, a forward genetic strategy was used employing mouse chromosome substitution strains (CSS) to identify novel FS susceptibility quantitative trait loci (QTLs). FS were induced by exposure to warm air at postnatal day 14. Video electroencephalogram monitoring identified tonic-clonic convulsion onset, defined as febrile seizure latency (FSL), as a reliable phenotypic parameter to determine FS susceptibility. FSL was determined in both sexes of the host strain (C57BL/6J), the donor strain (A/J) and CSS. C57BL/6J mice were more susceptible to FS than A/J mice. Phenotypic screening of the CSS panel identified six strains(CSS1, -2, -6 -10, -13 and -X) carrying QTLs for FS susceptibility. CSS1, -10 and -13 were less susceptible (protective QTLs), whereas CSS2, -6 and -X were more susceptible (susceptibility QTLs) to FS than the C57BL/6J strain. Our data show that mouse FS susceptibility is determined by complex genetics, which is distinct from that for chemically induced seizures. This is the first dataset using CSS to screen for a seizure trait in mouse pups. It provides evidence for common FS susceptibility QTLs that serve as starting points to fine map FS susceptibility QTLs and to identify FS susceptibility genes. This will increase our understanding of human FS, working toward the identification of new therapeutic targets.

High‐resolution genetic mapping of mammalian motor activity levels in mice

The generation of motor activity levels is under tight neural control to execute essential behaviors, such as movement toward food or for social interaction. To identify novel neurobiological mechanisms underlying motor activity levels, we studied a panel of chromosome substitution (CS) strains derived from mice with high (C57BL/6J strain) or low motor activity levels (A/J strain) using automated home cage behavioral registration. In this study, we genetically mapped the expression of baseline motor activity levels (horizontal distance moved) to mouse chromosome 1. Further genetic mapping of this trait revealed an 8.3-Mb quantitative trait locus (QTL) interval. This locus is distinct from the QTL interval for open-field anxiety-related motor behavior on this chromosome. By data mining, an existing phenotypic and genotypic data set of 2445 genetically heterogeneous mice (http://gscan.well.ox.ac.uk/), we confirmed linkage to the peak marker at 79 970 253 bp and refined the QTL to a 312-kb interval containing a single gene (A830043J08Rik). Sequence analysis showed a nucleotide deletion in the 3' untranslated region of the Riken gene. Genome-wide microarray gene expression profiling in brains of discordant F(2) individuals from CS strain 1 showed a significant upregulation of Epha4 in low-active F(2) individuals. Inclusion of a genetic marker for Epha4 confirmed that this gene is located outside of the QTL interval. Both Epha4 and A830043J08Rik are expressed in brain motor circuits, and similar to Epha4 mutants, we found linkage between reduced motor neurons number and A/J chromosome 1. Our findings provide a novel QTL and a potential downstream target underlying motor circuitry development and the expression of physical activity levels.

Genetic factors for resistance to diet-induced obesity and associated metabolic traits on mouse chromosome 17.

Obesity is associated with increased susceptibility to dyslipidemia, insulin resistance, and hypertension, a combination of traits that comprise the traditional definition of the metabolic syndrome. Recent evidence suggests that obesity is also associated with the development of nonalcoholic fatty liver disease (NAFLD). Despite the high prevalence of obesity and its related conditions, their etiologies and pathophysiology remains unknown. Both genetic and environmental factors contribute to the development of obesity and NAFLD. Previous genetic analysis of high-fat, diet-induced obesity in C57BL/6J (B6) and A/J male mice using a panel of B6-Chr(A/J)/NaJ chromosome substitution strains (CSSs) demonstrated that 17 CSSs conferred resistance to high-fat, diet-induced obesity. One of these CSS strains, CSS-17, which is homosomic for A/J-derived chromosome 17, was analyzed further and found to be resistant to diet-induced steatosis. In the current study we generated seven congenic strains derived from CCS-17, fed them either a high-fat, simple-carbohydrate (HFSC) or low-fat, simple-carbohydrate (LFSC) diet for 16 weeks and then analyzed body weight and related traits. From this study we identified several quantitative trait loci (QTLs). On a HFSC diet, Obrq13 protects against diet-induced obesity, steatosis, and elevated fasting insulin and glucose levels. On the LFSC diet, Obrq13 confers lower hepatic triglycerides, suggesting that this QTL regulates liver triglycerides regardless of diet. Obrq15 protects against diet-induced obesity and steatosis on the HFSC diet, and Obrq14 confers increased final body weight and results in steatosis and insulin resistance on the HFSC diet. In addition, on the LFSC diet, Obrq 16 confers decreased hepatic triglycerides and Obrq17 confers lower plasma triglycerides on the LFSC diet. These congenic strains provide mouse models to identify genes and metabolic pathways that are involved in the development of NAFLD and aspects of diet-induced metabolic syndrome.

Genetic architecture of complex traits: Large phenotypic effects and pervasive epistasis

The genetic architecture of complex traits underlying physiology and disease in most organisms remains elusive. We still know little about the number of genes that underlie these traits, the magnitude of their effects, or the extent to which they interact. Chromosome substitution strains (CSSs) enable statistically powerful studies based on testing engineered inbred strains that have single, unique, and nonoverlapping genetic differences, thereby providing measures of phenotypic effects that are attributable to individual chromosomes. Here, we report a study of phenotypic effects and gene interactions for 90 blood, bone, and metabolic traits in a mouse CSS panel and 54 traits in a rat CSS panel. Two key observations emerge about the genetic architecture of these traits. First, the traits tend to be highly polygenic: across the genome, many individual chromosome substitutions each had significant phenotypic effects and, within each of the chromosomes studied, multiple distinct loci were found. Second, strong epistasis was found among the individual chromosomes. Specifically, individual chromosome substitutions often conferred surprisingly large effects (often a substantial fraction of the entire phenotypic difference between the parental strains), with the result that the sum of these individual effects often dramatically exceeded the difference between the parental strains. We suggest that strong, pervasive epistasis may reflect the presence of several phenotypically-buffered physiological states. These results have implications for identification of complex trait genes, developmental and physiological studies of phenotypic variation, and opportunities to engineer phenotypic outcomes in complex biological systems.

Genetic analysis of the psychostimulant effects of nicotine in chromosome substitution strains and F2 crosses derived from A/J and C57BL/6J progenitors

Previous research utilizing the AcB/BcA recombinant congenic strains (RCS) of mice mapped provisional quantitative trait loci (QTLs) for the psychostimulant effects of nicotine to multiple regions on chromosomes 7, 11, 12, 14, 16, and 17. The current study was designed to confirm these QTLs in an A/J (A) x C57Bl/6J (B6) F2 cross and a panel of B6.A chromosome substitution strains (CSS). The panel of B6.A CSS consists of 21 strains, each carrying a different A/J chromosome on a B6 background. The A x B6 F2, CSS, A, and B6 mice were tested for sensitivity to the effects of nicotine on locomotor activity using a computerized open-field apparatus. In A x B6 F2 mice two QTLs were identified which confirm those previously observed in the AcB/BcA RCS. Significant differences in the expression of nicotine-induced activity were associated with loci on chromosome 11 (D11Mit62) and chromosome 16 (D16Mit131) in the A x B6 F2. At the chromosome 11 QTL, an A allele was associated with lower nicotine-induced activity scores relative to the B6. In contrast, the A allele was associated with greater relative nicotine activity values for the chromosome 16 QTL. A survey of the CSS panel confirmed the presence of QTLs for nicotine activation on chromosomes 2, 14, 16, and 17 previously identified in the AcB/BcA RCS. In the informative CSS strains, A alleles were consistently associated with greater nicotine-induced activity scores compared to the B6. The results of the present study are the first to validate QTLs for sensitivity to the effects of nicotine across multiple strains of mice. QTLs on chromosomes 2, 11, 14, 16, and 17 were confirmed in CSS and/or F2 mice. Significantly, the identification of a QTL on chromosome 16 has now been replicated in three crosses derived from the A and B6 progenitors.

Evidence for Epigenetic Interactions for Loci on Mouse Chromosome 1 Regulating Open Field Activity

The expression of motor activity levels in response to novel situations is under complex genetic and environmental control. Several genetic loci have been implicated in the regulation of this behavioral phenotype, but their relationship to epigenetic and epistatic interactions is relatively unknown. Here, we report on a quantitative trait locus (QTL) on mouse chromosome 1 for novelty-induced motor activity in the open field, using chromosome substitution strains derived from a high active host strain (C57BL/6J) and a low active donor strain (A/J). The QTL for open field (horizontal distance moved) peaked at the location of Kcnj9, however, QTL detection was initially masked by an interplay of both grandparent genetic origin and genetic co-factors influencing behavior on chromosome 1. Our findings indicate that epigenetic interactions can play an important role in the identification of behavioral QTLs and must be taken into consideration when applying behavioral genetic strategies.