Identification of loci controlling restriction of parasite growth in experimental Taenia crassiceps cysticercosis

Human neurocysticercosis (NC) caused by Taenia solium is a parasitic disease of the central nervous system that is endemic in many developing countries. In this study, a genetic approach using the murine intraperitoneal cysticercosis caused by the related cestode Taenia crassiceps was employed to identify host factors that regulate the establishment and proliferation of the parasite. A/J mice are permissive to T. crassiceps infection while C57BL/6J mice (B6) are comparatively restrictive, with a 10-fold difference in numbers of peritoneal cysticerci recovered 30 days after infection. The genetic basis of this inter-strain difference was explored using 34 AcB/BcA recombinant congenic strains derived from A/J and B6 progenitors, that were phenotyped for T. crassiceps replication. In agreement with their genetic background, most AcB strains (A/J-derived) were found to be permissive to infection while most BcA strains (B6-derived) were restrictive with the exception of a few discordant strains, together suggesting a possible simple genetic control. Initial haplotype association mapping using >1200 informative SNPs pointed to linkages on chromosomes 2 (proximal) and 6 as controlling parasite replication in the AcB/BcA panel. Additional linkage analysis by genome scan in informative [AcB55xDBA/2]F1 and F2 mice (derived from the discordant AcB55 strain), confirmed the effect of chromosome 2 on parasite replication, and further delineated a major locus (LOD = 4.76, p<0.01; peak marker D2Mit295, 29.7 Mb) that we designate Tccr1 (T. crassiceps cysticercosis restrictive locus 1). Resistance alleles at Tccr1 are derived from AcB55 and are inherited in a dominant fashion. Scrutiny of the minimal genetic interval reveals overlap of Tccr1 with other host resistance loci mapped to this region, most notably the defective Hc/C5 allele which segregates both in the AcB/BcA set and in the AcB55xDBA/2 cross. These results strongly suggest that the complement component 5 (C5) plays a critical role in early protective inflammatory response to infection with T. crassiceps.     

Recombinant congenic strains derived from A/J and C57BL/6J: a tool for genetic dissection of complex traits

Complex genetic traits can be dissected in mice, using well-defined sets of recombinant inbred strains, congenic strains, and recombinant congenic strains (RCS). We report the creation of a series of 37 independent RCS derived from the commonly used inbred strains of laboratory mouse A/J (A) and C57BL/6J (B6). These RCS were derived by systematic inbreeding of independent pairs of animals from a (F1 x A) x A and a (F1 x B) x B double backcross (N3), to create AcB and BcA strains, respectively. Fifteen AcB strains and 22 BcA strains at between 18 and 30 generations of inbreeding have been generated, are healthy, and show stable breeding performance. These strains have been genotyped for a total of 625 informative microsatellite DNA markers covering the entire genome, with an average spacing of 2.6 cM. Haplotype analyses indicate that on average, AcB and BcA strains contain 13.25% of the donor genome, a value close to the 12.5% expected from the breeding scheme used in their creation. In the AcB set, approximately 79% of the B6 genome has been transferred in independent strains, while in the BcA set approximately 84% of the A genome is represented on the B6 background. This represents an excellent coverage of congenic segments from both parental genomes in the two sets of strains, which can now be used to map simple and complex traits in a genome-wide fashion. As an example of the power of AcB/BcA strains as a mapping tool, the 37 strains were typed for susceptibility to infection with Legionella pneumophila, a monogenic trait controlled by the Lgn1 locus on Chromosome 13. Analysis of the strain distribution pattern of L. pneumophila susceptibility allowed direct mapping of Lgn1 to a 3-cM interval. The AcB/BcA set should prove a useful tool with which to investigate the complex genetic basis of known interstrain differences between A and B6 for many important diseases.     

Genetic analysis of alcohol intake in recombinant inbred and congenic strains derived from A/J and C57BL/6J progenitors

The objective of the present study was to map quantitative trait loci (QTL) for alcohol intake using A × B/B × A recombinant inbred (RI) and AcB/BcA recombinant congenic (RC) strains of mice that were independently derived from the A/J and C57BL/6J progenitors. Mice were screened for levels of alcohol consumption with four days of forced exposure to alcohol, followed by three weeks of free choice between water and a 10% alcohol solution. Alcohol consumption data previously collected for 27 A × B/B × A RI strains were reanalyzed using a larger marker set and composite interval mapping. The reanalysis found markers on Chromosome 2 (D2Mit74, 107 cM) (males and females) and on Chromosome 11 (Pmv22, 8 cM) (females only) that exceeded the threshold for significant loci, and found suggestive loci (in males) on Chromosomes 10 (D10 Mit126, 21 cM), 12 (D12Mit37, 1 cM), 15 (Pdgfb, 46.8 cM), and 16 (D16Mit125, 29 cM). An additional suggestive locus was identified in female RI mice on Chromosome 11 (D11Mit120, 47.5 cM). Composite interval mapping (CIM) analysis indicated that there was a significant association between loci at Pdgfb and D2Mit74 in both males and females. Analysis of the AcB/BcA RC strains identified 11 QTL on Chromosomes 2, 3, 5,6, 7, 8, 9, 10, 12, 13, and 15. QTL on Chromosomes 7, 10, 12, and 15 were identified in both the A × B/B × A RI and AcB/BcA RC strains of mice. Additional QTLs identified on Chromosomes 2, 3, 7, 11, and 15 overlap with those previously identified in the literature using strains of mice with a C57BL/6J progenitor.     

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) × 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 × 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 × 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 × 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.     

Genetic control of high density lipoprotein-cholesterol in AcB/BcA recombinant congenic strains of mice

Epidemiological studies show that high HDL-cholesterol (HDLc) decreases the risk of cardiovascular disease. To map genes controlling lipid metabolism, particularly HDLc levels, we screened the plasma lipids of 36 AcB/BcA RC mouse strains subjected to either a normal or a high-fat/cholesterol diet. Strains BcA68 and AcB65 showed deviant HDLc plasma levels compared with the parental A/J and C57BL/6J strains; they were thus selected to generate informative F2 crosses. Linkage analyses in the AcB65 strain identified a locus on chromosome 4 (Hdlq78) responsible for high post-high fat diet HDLc levels. This locus has been previously associated at genome-wide significance to two regions in the human genome. A second linkage analysis in strain BcA68 identified linkage in the vicinity of a gene cluster known to control HDLc levels. Sequence analysis of these candidates identified a de novo, loss-of-function mutation in the ApoA1 gene of BcA68 that prematurely truncates the ApoA1 protein. The possibility of dissecting the specific effects of this new ApoA1 deficiency in the context of isogenic controls makes the BcA68 mouse a valuable new tool.     

Mapping hyper-susceptibility to colitis-associated colorectal cancer in FVB/NJ mice

Inbred strains of mice differ in susceptibility to colitis-associated colorectal cancer (CA-CRC). We tested 10 inbred strains of mice for their response to azoxymethane/dextran sulfate sodium-induced CA-CRC and identified a bimodal inter-strain distribution pattern when tumor multiplicity was used as a phenotypic marker of susceptibility. The FVB/NJ strain was particularly susceptible showing a higher tumor burden than any other susceptible strains (12.5-week post-treatment initiation). FVB/NJ hyper-susceptibility was detected as early as 8-week post-treatment initiation with FVB/NJ mice developing 5.5-fold more tumors than susceptible A/J or resistant B6 control mice. Linkage analysis by whole genome scan in informative (FVB/NJ×C3H/HeJ)F2 mice identified a novel susceptibility locus designated as C olon c ancer s usceptibility 6 (Ccs6) on proximal mouse chromosome 6. When gender was used as a covariate, a LOD score of 5.4 was computed with the peak marker being positioned at rs13478727, 43.8 Mbp. Mice homozygous for FVB/NJ alleles at this locus had increased tumor multiplicity compared to homozygous C3H/HeJ mice. Positional candidates in this region of chromosome 6 were analyzed with respect to a possible role in carcinogenesis and a role in inflammatory response using a new epigenetic gene scoring tool (Myeloid Inflammation Score).     

Genetic basis for the psychostimulant effects of nicotine: a quantitative trait locus analysis in AcB/BcA recombinant congenic mice

Genetic differences in sensitivity to nicotine have been reported in both animals and humans. The present study utilized a novel methodology to map genes involved in regulating both the psychostimulant and depressant effects of nicotine in the AcB/BcA recombinant congenic strains (RCS) of mice. Locomotor activity was measured in a computerized open-field apparatus following subcutaneous administration of saline (days 1 and 2) or nicotine on day 3. The phenotypic measures obtained from this experimental design included total basal locomotor activity, as well as total nicotine activity, nicotine difference scores, nicotine percent change and nicotine regression residual scores. The results indicated that the C57BL/6J (B6) were insensitive to nicotine over the entire dose-response curve (0.1, 0.2, 0.4 and 0.8 mg/kg). However, the 0.8-mg/kg dose of nicotine produced a significant decrease in the locomotor activity in the A/J strain and a wide and continuous range of both locomotor excitation and depression among the AcB/BcA RCS. Single-locus association analysis in the AcB RCS identified quantitative trait loci (QTL) for the psychostimulant effects of nicotine on chromosomes 11, 12, 13, 14 and 17 and one QTL for nicotine-induced depression on chromosome 11. In the BcA RCS, nicotine-induced locomotor activation was associated with seven putative regions on chromosomes 2, 7, 8, 13, 14, 16 and 17. There were no overlapping QTL and no genetic correlations between saline- and nicotine-related phenotypes in the AcB/BcA RCS. A number of putative candidate genes were in proximity to regions identified with nicotine sensitivity, including the alpha2 subunit of the nicotinic acetylcholine receptor and the dopamine D3 receptor.     

Mapping of a Chromosome 12 Region Associated with Airway Hyperresponsiveness in a Recombinant Congenic Mouse Strain and Selection of Potential Candidate Genes by Expression and Sequence Variation Analyses

In a previous study we determined that BcA86 mice, a strain belonging to a panel of AcB/BcA recombinant congenic strains, have an airway responsiveness phenotype resembling mice from the airway hyperresponsive A/J strain. The majority of the BcA86 genome is however from the hyporesponsive C57BL/6J strain. The aim of this study was to identify candidate regions and genes associated with airway hyperresponsiveness (AHR) by quantitative trait locus (QTL) analysis using the BcA86 strain. Airway responsiveness of 205 F2 mice generated from backcrossing BcA86 strain to C57BL/6J strain was measured and used for QTL analysis to identify genomic regions in linkage with AHR. Consomic mice for the QTL containing chromosomes were phenotyped to study the contribution of each chromosome to lung responsiveness. Candidate genes within the QTL were selected based on expression differences in mRNA from whole lungs, and the presence of coding non-synonymous mutations that were predicted to have a functional effect by amino acid substitution prediction tools. One QTL for AHR was identified on Chromosome 12 with its 95% confidence interval ranging from 54.6 to 82.6 Mbp and a maximum LOD score of 5.11 (p = 3.68×10⁻³). We confirmed that the genotype of mouse Chromosome 12 is an important determinant of lung responsiveness using a Chromosome 12 substitution strain. Mice with an A/J Chromosome 12 on a C57BL/6J background have an AHR phenotype similar to hyperresponsive strains A/J and BcA86. Within the QTL, genes with deleterious coding variants, such as Foxa1, and genes with expression differences, such as Mettl21d and Snapc1, were selected as possible candidates for the AHR phenotype. Overall, through QTL analysis of a recombinant congenic strain, microarray analysis and coding variant analysis we identified Chromosome 12 and three potential candidate genes to be in linkage with airway responsiveness.     

Assignment of a locus for mouse lung tumor susceptibility to proximal Chromosome 19

Previous studies have hypothesized that at least three genetic loci contribute to differences in pulmonary adenoma susceptibility between mouse strains A/J and C57BL/6J. One gene that may confer susceptibility to lung tumorigenesis is the Kras protooncogene. To identify other relevant loci involved in this polygenic trait, we determined tumor multiplicity in 56 randomly chosen N-ethyl-N-nitrosourea-treated (A/J×C57BL/6J) N1×C57BL/6 backcross (AB6N2) progeny and correlated it with genotypes at 77 microsatellite markers spanning the genome. A correlation of lung tumor multiplicity phenotypes with genotypes of microsatellite markers on distal Chromosome (Chr) 6 in the Kras region (Pas1) was confirmed, and a new region on Chr 19 (designated Pas3) was identified that also contributes to susceptibility. Linkage analysis on Chr 19 with 270 AB6N2 mice localized the region flanked by D19Mit42 and D19Mit19 that is most closely associated with lung tumor susceptibility. The Pas3 locus may be an enhancer of the susceptibility locus on Chr 6.     

Mapping of clinical and expression quantitative trait loci in a sex-dependent effect of host susceptibility to mouse-adapted influenza H3N2/HK/1/68

Seasonal influenza outbreaks and recurrent influenza pandemics present major challenges to public health. By studying immunological responses to influenza in different host species, it may be possible to discover common mechanisms of susceptibility in response to various influenza strains. This could lead to novel therapeutic targets with wide clinical application. Using a mouse-adapted strain of influenza (A/HK/1/68-MA20 [H3N2]), we produced a mouse model of severe influenza that reproduces the hallmark high viral load and overexpression of cytokines associated with susceptibility to severe influenza in humans. We mapped genetic determinants of the host response using a panel of 29 closely related mouse strains (AcB/BcA panel of recombinant congenic strains) created from influenza-susceptible A/J and influenza-resistant C57BL/6J (B6) mice. Combined clinical quantitative trait loci (QTL) and lung expression QTL mapping identified candidate genes for two sex-specific QTL on chromosomes 2 and 17. The former includes the previously described Hc gene, a deficit of which is associated with the susceptibility phenotype in females. The latter includes the phospholipase gene Pla2g7 and Tnfrsf21, a member of the TNFR superfamily. Confirmation of the gene underlying the chromosome 17 QTL may reveal new strategies for influenza treatment.     

Genome-wide association and fine mapping of genetic loci predisposing to colon carcinogenesis in mice

To identify the genetic determinants of colon tumorigenesis, 268 male mice from 33 inbred strains derived from different genealogies were treated with azoxymethane (AOM; 10 mg/kg) once a week for six weeks to induce colon tumors. Tumors were localized exclusively within the distal colon in each of the strains examined. Inbred mouse strains exhibit a large variability in genetic susceptibility to AOM-induced colon tumorigenesis. The mean colon tumor multiplicity ranged from 0 to 38.6 (mean = 6.5 ± 8.6) and tumor volume ranged from 0 to 706.5 mm(3) (mean = 87.4 ± 181.9) at 24 weeks after the first dose of AOM. AOM-induced colon tumor phenotypes are highly heritable in inbred mice, and 68.8% and 71.3% of total phenotypic variation in colon tumor multiplicity and tumor volume, respectively, are attributable to strain-dependent genetic background. Using 97,854 single-nucleotide polymorphisms, we carried out a genome-wide association study (GWAS) of AOM-induced colon tumorigenesis and identified a novel susceptibility locus on chromosome 15 (rs32359607, P = 6.31 × 10(-6)). Subsequent fine mapping confirmed five (Scc3, Scc2, Scc12, Scc8, and Ccs1) of 16 linkage regions previously found to be associated with colon tumor susceptibility. These five loci were refined to less than 1 Mb genomic regions of interest. Major candidates in these loci are Sema5a, Fmn2, Grem2, Fap, Gsg1l, Xpo6, Rabep2, Eif3c, Unc5d, and Gpr65. In particular, the refined Scc3 locus shows high concordance with the human GWAS locus that underlies hereditary mixed polyposis syndrome. These findings increase our understanding of the complex genetics of colon tumorigenesis, and provide important insights into the pathways of colorectal cancer development and might ultimately lead to more effective individually targeted cancer prevention strategies.     

Susceptibility to a mouse acquired immunodeficiency syndrome is influenced by theH-2

The development of a mouse acquired immunodeficiency syndrome (MAIDS) induced following LP-BM5 MuLV infection depends on host genetic factors. Susceptible mice, such as C57BL/6J mice, develop a profound impairment of lymphoproliferative response to mitogens and hyperplasia of lymphoid organs and succumb to infection within 6 months. These changes do not occur in resistant mice, such as A/J mice. Resistance to MAIDS is a dominant trait since (C57BL/6JxA/J)F₁ hybrid mice did not develop any immune dysfunctions following infection. Genetic regulation of the trait of resistance/susceptibility to MAIDS was determined in AXB/BXA recombinant inbred (RI) mouse strains (derived from resistant A/J and susceptible C57BL/6J progenitors). Two different criteria were used to determine their resistance or susceptibility to developing MAIDS: the gross pathologic evaluation of lymphoid organs at 13–15 weeks of infection, and survival. RI mouse strains segregated into two non-overlapping groups. The first group did not develop any significant pathology, and these mouse strains were considered as resistant to MAIDS. The second group showed the virus-induced pathological changes as well as an immunological dysfunction as seen in C57BL/6J progenitor mice, and these strains were thus considered as susceptible to MAIDS. This bimodal strain distribution pattern of resistance/susceptibility to MAIDS among the RI strains suggests that this phenotype is controlled by a single gene. Linkage analysis with other allelic markers showed a strong association between resistance/susceptibility to MAIDS and theH-2 complex. Possession of theH-2 ^b haplotype derived from C57BL/6J mice was associated with susceptibility to MAIDS, while theH-2 ^a haplotype conferred resistance to the disease. This finding was confirmed by demonstrating thatH-2 ^a congenics on the susceptible C57BL/10 background were as resistant to MAIDS as A/J mice which donated theH-2 ^a locus. Gene(s) within theH-2 complex thus represent the major regulatory mechanism of resistance/susceptibility to MAIDS.     

Comparative biochemistry of murine arylsulfatase B

Arylsulfatase B was purified 4500-fold from liver and kidney of C57BL/6J mice. Hepatic and renal arysulfatase B are apparently determined by a single structural locus; however, posttranslational modification introduces inter- and intratissue microheterogeneity. Partially purified enzyme from C57BL/6J, A/J, C3H/HeJ, and SWR/J mice has similar catalytic properties. The 4500-fold-purified arylsulfatase B from SWR/J and C3H/HeJ mice was more thermostable than that from C57BL/6J and A/J mice, strongly suggesting that the thermostability difference reflects an alteration of the primary structure of the enzyme. Thermal stability of arylsulfatase B was pH dependent and markedly influenced by buffer anion. Variation of thermostability did not appear accountable for the observed activity variation among these strains; however, this possibility cannot be rigorously excluded by presently available data. Thirty-five murine strains were found to possess the As-1 ^a allele (thermostable enzyme), while As-1 ^b was largely restricted to A and C57 strains.This research was supported by PHS Biomedical Sciences Research Support Grant RR-07030.     

Radiation-induced lung response of AcB/BcA recombinant congenic mice

Lemay, A-M. and Haston, C. K. Radiation-Induced Lung Response of AcB/BcA Recombinant Congenic Mice. Radiat. Res. 170, 299-306 (2008).The genetic factors that influence the development of radiotherapy-induced lung disease are largely unknown. Herein we identified a strain difference in lung response to radiation wherein A/J mice developed alveolitis with increased levels of pulmonary mast cells and cells in bronchoalveolar lavage while the phenotype in C57BL/6J mice was fibrosis with fewer inflammatory cells. To identify genomic loci that may influence these phenotypes, we assessed recombinant congenic (RC) mice derived from the A/J and C57BL/6J strains for their propensity to develop alveolitis or fibrosis after 18 Gy whole-thorax irradiation. Mouse survival, lung histopathology and bronchoalveolar lavage cell types were recorded. Informative strains for each of mast cell influx, bronchoalveolar cell numbers, alveolitis and fibrosis were identified. In mice with the A/J strain background, the severity of alveolitis correlated with increased mast cell numbers while in C57BL/6J background strain mice fibrosis was correlated with the percentage of neutrophils in lavage. The data for RC mice support the association of specific inflammatory cells with the development of radiation-induced lung disease and provide informative strains with which to dissect the genetic basis of these complex traits.     

H2 control of natural T regulatory cell frequency in the lymph node correlates with susceptibility to day 3 thymectomy-induced autoimmune disease

Day 3 thymectomy (D3Tx) results in a loss of peripheral tolerance mediated by natural regulatory T cells (nTregs) and development of autoimmune ovarian dysgenesis (AOD) and autoimmune dacryoadenitis (ADA) in A/J and (C57BL/6J × A/J) F(1) hybrids (B6A), but not in C57BL/6J (B6) mice. Previously, using quantitative trait locus (QTL) linkage analysis, we showed that D3Tx-AOD is controlled by five unlinked QTL (Aod1-Aod5) and H2. In this study, using D3Tx B6-Chr(A/J)/NaJ chromosome (Chr) substitution strains, we confirm that QTL on Chr16 (Aod1a/Aod1b), Chr3 (Aod2), Chr1 (Aod3), Chr2 (Aod4), Chr7 (Aod5), and Chr17 (H2) control D3Tx-AOD susceptibility. In addition, we also present data mapping QTL controlling D3Tx-ADA to Chr17 (Ada1/H2), Chr1 (Ada2), and Chr3 (Ada3). Importantly, B6-ChrX(A/J) mice were as resistant to D3Tx-AOD and D3Tx-ADA as B6 mice, thereby excluding Foxp3 as a susceptibility gene in these models. Moreover, we report quantitative differences in the frequency of nTregs in the lymph nodes (LNs), but not spleen or thymus, of AOD/ADA-resistant B6 and AOD/ADA-susceptible A/J, B6A, and B6-Chr17(A/J) mice. Similar results correlating with experimental allergic encephalomyelitis and orchitis susceptibility were seen with B10.S and SJL/J mice. Using H2-congenic mice, we show that the observed difference in frequency of LN nTregs is controlled by Ada1/H2. These data support the existence of an LN-specific, H2-controlled mechanism regulating the prevalence of nTregs in autoimmune disease susceptibility.     

Assignment of the Ly-6--Ril-1--Sis--H-30--Pol-5/Xmmv-72--Ins-3--Krt-1--Int-1--Gdc-1 region to mouse chromosome 15

Previous work has demonstrated linkage between Ly-6, H-30, and a locus, Ril-1, that affects susceptibility to radiation-induced leukemia. Results of preliminary linkage analyses suggested further that the cluster might be linked to Ly-11 on the proximal portion of mouse chromosome 2. Using molecular probes to examine somatic cell lines and recombinant inbred and congenic strains of mice, we have re-evaluated these linkage relationships. A cloned genomic DNA fragment derived from a retroviral site has been used to define a novel locus, Pol-5, that is tightly linked to both H-30 and Ril-1 as shown by analysis of the B6.C-H-30 ^c congenic mouse strain. Following the segregation of the Pol-5 mouse-specific DNA fragment in a series of somatic cell hybrids carrying various combinations of mouse chromosomes on a rat or Chinese hamster background mapped Pol-5 to mouse chromosome 15. During the course of these studies, restriction fragment length polymorphisms were defined associated with several loci, including Pol-5, Ly-6, Sis, Ins-3, Krt-1, Int-1, and Gdc-1. Three of these loci, Sis, Int-1, and Gdc-1, have been previously mapped to chromosome 15 by others using somatic cell hybrids or isoenzyme analyses. Following the inheritance of these eight loci in recombinant inbred strains of mice allowed the definition of a linkage group on the chromosome with the order Ly-6-Ril-1--Sis--H-30--Pol-5--Ins-3--Krt-1--Int-1--Gdc-1. Analyses of alleles inherited as passengers in B6.C-H-30 ^c, C3H.B-Ly-6 ^b, and C57BL/6By-Eh/+ congenic mouse strains and in situ hybridization experiments support the above gene order and indicate further that the cluster is located on distal chromosome 15, with Ly-6 and Sis near Eh.Abbreviations A agouti - Abl cellular homolog of the Abelson leukemia virus oncogene - Ada adenosine deaminase - Ak-1 adenylate kinase-1 - AXB A/J × C57BL/6J recombinant inbred strain - B2m beta-2 microglobulin - BXA C57BL/6J × A/J recombinant inbred strain - BXD C57BL/6J × DBA/2J recombinant inbred strain - BXH C57BL/6J × C3H/HeJ recombinant inbred strain - CXB BALB/cBy × C57BL/6By recombinant inbred strain - DNA deoxyribonucleic acid - Eh hairy ears - Fpgs folypolyglutamyl synthetase - FXI fractionated x-irradiation - Gdc-1 glycerol phosphate dehydrogenase-1 - Il2r IL-2 receptor - Ins-3 a novel insulinlike gene - Int-1 mammary tumor integration site-1 - Itp inosine triphosphatase - Krt-1 the locus designated here includes a cluster of at least three keratin genes - LTR long terminal repeat - Ly lymphocyte - Lv-6 lymphocyte antigen-6 - Ly-11 lymphocyte antigen-11 - MIH minor histocompatibility - Myc cellular homolog of the Abelson leukemia virus oncogene; pa, pallid; - Pol-5 locus encoding retroviral polymerase-5 - RFLP restriction fragment length polymorphism - RI recombinant inbred mouse strains - Ril-1 radiation-induced leukemia susceptibility-1 locus - SDP strain distribution pattern - Sis cellular homolog of the simian sarcoma virus oncogene - SFFV spleen focus-forming virus - Tpi-1 triosephosphate isomerase-1 - Ve velvet     

BALB/c antiarsonate idiotypes: Gene complementation necessary for expression

The expression of two idiotype (id) families (5AF6 and 3C6) associated with the BALB/c p-azophenylarsonate-specific antibody response was examined in 11 mouse strains. Eight strains produced some of one or the other of these two id families with the mean percent expression in the anti-Ar responses of id(+) strains ranging from 8 to 43% for the 5AF6 and from 2 to 10% for the 3C6 idiotype. Four strains of mice (C58, AKR, PL, and RF) thought to have Lyt-3.1-linked V_L repertoire differences from other mouse strains (Lyt-3.2) were tested for their capacity to contribute to 5AF6 and 3C6 id expression. The RE strain was capable of producing 5AF6 id and small amounts of 3C6 id. Tests of Lyt-3.1 congenic strains C.AKR (AKR Lyt-3.1 on a BALB/c background) and C.C58 (C58 Lyt-3.1 on a BALB/c background) showed that C.AKR could produce 5AF6 id while C.C58 could not. 3C6 id expression was present but depressed in C.C58 mice compared with the high 3C6 id expression in C.AKR. Breeding studies mating C.C58 (bearing the required Igh-C^a-linked V _H genes) to other 5AF6(–) strains showed that gene complementation could result in 5AF6 expression in F₁ offspring. 5AF6(–) strains capable of complementation included CBA/J, C57BL/6J, AKR/J, and PL/J. C58/J (from which C.C58 were derived) was the only tested strain that failed to complement for 5AF6 id expression. Additional matings between C58/J[5AF6(–)] and CBA/J[5AF6 (–)] showed F1 offspring could produce 5AF6 id, indicating that C58/J can contribute functional V _H genes necessary for 5AF6 id expression. Depressed expression of 5AF6 and 3C6 id was noted in mice where the C58/J-derived Lyt-3.1 genotype was present. The possibility that the depression of 5AF6 and 3C6 id expression derived from C58/J mice was due to regulatory influences rather than a lack of the V _L structural genes is discussed.Abbreviations used in this paper Ar p-azophenylarsonate - DNP dinitrophenol - H heavy - HIS hyperimmune suppressed - id idiotype - Ig immunoglobulin - IEF isoelectric focusing - k kappa - L light - PC phosphorylcholine - S.D. standard deviation     

Impaired interleukin-3 (IL-3) response of the A/J mouse is caused by a branch point deletion in the IL-3 receptor alpha subunit gene.

Interleukin-3 (IL-3) alone does not support hematopoietic colony formation of bone marrow cells from the A/J mouse. To elucidate the molecular lesion in A/J mice, we examined expression of the alpha and beta subunits of the IL-3 receptor (IL-3R). While IL-3R beta was normally expressed, IL-3R alpha was not detectable on the surface of A/J-derived cells by antibody staining. Genetic linkage analysis using recombinant inbred mouse strains between A/J and IL-3-responsive C57BL/6 indicated that the IL-3R alpha gene locus was responsible for the impaired IL-3 response in A/J mice. Molecular cloning and characterization of A/J-derived IL-3R alpha cDNA revealed that it lacked the sequence corresponding to exon 8, which encodes 10 amino acid residues in the extracellular domain. The aberrant splicing was due to a 5 base pair deletion at the branch point in intron 7 and was reproduced in heterologous cells by transfecting with an IL-3R alpha minigene carrying the deleterious intron. The A/J-specific abnormal form of IL-3R alpha was localized inside the cells, but not on the cell surface, providing the molecular basis for the impaired IL-3 response in the A/J mouse.