Regulation of polyunsaturated fat induced postprandial hypercholesterolemia by a novel genePHC-2

Inbred mouse strains vary in susceptibility or resistance to dietary induced atherosclerosis. To investigate the effect of polyunsaturated fat feeding on postprandial serum cholesterol levels, in C57BL/67 (B6) and BALB/cJ inbred mice, we fed by stomach gavage previously fasted mice, a mixture containing 30% sunflower oil, 5% cholesterol, 2% sodium cholate and 0.5% choline chloride. The most significant difference in serum cholesterol levels between B6 and BALB/cJ mouse strains was observed at 2 h postfeeding. Susceptible B6 strain mice had a 41% postprandial increment in serum cholesterol. The resistant BALB/cJ strain had an insignificant 16% rise in serum cholesterol, at 2 h. We next examined eight other inbred mouse strains, to identify the gene(s) that regulate the observed 2 h postprandial hypercholesterolemia response, in the susceptible B6 mouse strain. Only the C57BR/cdJ and C57L/J strains developed postprandial hypercholesterolemia, at 2 h. The C57BR/cdJ strain had a 20% increase and the C57L/J strain a 62% increase in postprandial serum cholesterol levels. From this result, we found that the postprandial hypercholesterolemic response to an acute polyunsaturated fat-cholesterol feed, cosegregated with the a allele at the Gpd-1 and Ahd-1 loci, on mouse chromosome 4. In this study, non-responsiveness cosegregated with the b allele at the Gpd-1 and Ahd-1 loci. Thus polyunsaturated fat-cholesterol induced postprandial hypercholesterolemia appeared to be genetically determined by a gene located between the Gpd-1 and Ahd-1 loci, in mice. The putative gene regulating polyunsaturated fat-cholesterol induced post-absorptive hypercholesterolemia was designated Phc-2. Further studies with (BALB/cJ×C57BL/6J) Recombinant Inbred strains mapped the Phc-2 gene to a 2 cM region, within the Gpd-1 and Ahd-1 chromosomal segment, between the Pgd and Gpd-1 loci, on mouse chromosome 4.Abbreviations B6 C57BL/6J - HDL High Density Lipoprotein - LDL Low Density Lipoprotein - Phe Postprandial Hypercholesterolemia - PUFA-C Polyunsaturated Fat with Cholesterol - VLDL Very Low Density Lipoprotein     

Genetic variation of an intestinal leucine arylaminopeptidase (Lap-1) in the mouse and its location on chromosome 9

Electrophoretic variation for an intestinal enzyme that cleaves L-leucyl-beta-naphthylamide has been discovered among inbred mouse strains. Several strains including related strains C57BL/6J, C57BL/10J, C57BR/cdJ, C57L/J, and C58/J demonstrate an electrophoretic band of this enzyme that is absent in other strains and stocks thus far observed. The enzyme is tentatively being called leucine arylaminopeptidase (LAP) and the variant genetic locus Lap-1. The presence of the band is determined by an allele designated Lap-1a. Homozygotes for the alternate allele, Lap-1b, are without the band and heterozygotes are, under our electrophoretic conditions, indistinguishable from Lap-1a homozygotes. Data from recombinant inbred lines and a B6D2F1 X DBA/2J backcross established linkage of Lap-1 to dilute (d) and supernatant malic enzyme (Mod-1) on chromosome 9 in the following order: Lap-1-d-Mod-1. The Lap-1 to d map distance was estimated to be 21.3 +/- 4.6 cM from backcross data and 8.1 +/- 4.8 cM from recombinant inbred lines.     

Inhibition of 3beta-hydroxysteroid dehydrogenase-isomerase in mouse adrenal cells: a direct effect of testosterone

Gonadal steroids modulate adrenal gland size and function in a variety of species, and our previous studies demonstrate that circulating androgens suppress 3beta-hydroxysteroid dehydrogenase-isomerase (3betaHSD) activity in the adrenal cortex of male mice. The present study tests the hypothesis that androgens have a direct, receptor-mediated inhibitory effect on adrenal 3betaHSD. Treatment of cultured adrenal cells from C57BL/6J and C3H/HeJ mice with 0.02-2.0 microM testosterone for 7 days significantly reduces 3betaHSD activity in cells from both strains. However, treatment for 3 days reduces 3betaHSD activity in the adrenal cells from C3H/HeJ, but not C57BL/6J mice. The decreases in 3betaHSD activity in response to testosterone treatment is reflected in decreases in the amount of 3betaHSD immunoreactive protein, such that extended treatment decreases 3betaHSD immunoreactive protein in adrenal cells from both strains, but short-term treatment only decreases 3betaHSD immunoreactive protein in adrenal cells from C3H/HeJ mice. Thus, there appears to be a temporal difference between strains in the effect of the testosterone on 3betaHSD activity and immunoreactive protein. Treatment of the adrenal cells with androgen agonists and an antagonist indicate that the effect of testosterone is androgen receptor mediated. The effect of testosterone appears to be specific for 3betaHSD, since none of the treatments alter P450(scc) in cells from either strain. Testosterone treatment also causes a decrease in the amount of 3betaHSD mRNA. However, in contrast to the effect on activity and immunoreactive protein, there is no strain-related temporal difference because testosterone decreases 3betaHSD mRNA within 24h in adrenal cells from both strains. These results indicate that testosterone can act directly on the adrenal gland to decrease 3betaHSD activity, immunoreactive protein, and mRNA content in mouse adrenal glands, and thus contribute to the sex difference in adrenal function observed in many species.     

A major gene affecting age-related hearing loss is common to at least ten inbred strains of mice

Inbred strains of mice offer promising models for understanding the genetic basis of human presbycusis or age-related hearing loss (AHL). We previously mapped a major gene affecting AHL in C57BL/6J mice. Here, we show that the same Chromosome 10 gene (Ahl) is a major contributor to AHL in nine other inbred mouse strains-129P1/ReJ, A/J, BALB/cByJ, BUB/BnJ, C57BR/cdJ, DBA/2J, NOD/LtJ, SKH2/J, and STOCK760. F1 hybrids between each of these inbred strains and the normal-hearing inbred strain CAST/Ei retain good hearing, indicating that inheritance of AHL is recessive. To follow segregation of hearing loss, F1 hybrids were backcrossed to the parental strains with AHL. Auditory-evoked brain-stem response thresholds were used to assess hearing in more than 1500 N2 mice and analyzed as quantitative traits for linkage associations with Chromosome 10 markers. Highly significant linkage was found in all nine strain backcrosses, with the highest probability (LOD > 70) near the marker D10Mit112. This map position for Ahl is near the waltzer mutation (v) and the modifier of deaf waddler locus (mdfw), suggesting the possibility of allelism. Results from an intercross of C57BL/6J and NOD/LtJ mice indicate that the 6- to 10-month difference in AHL onset between these two strains is not due to allelic heterogeneity of the Ahl gene.     

Radiation-induced pulmonary endothelial dysfunction and hydroxyproline accumulation in four strains of mice

C57BL mice exposed to 14 Gy of whole-thorax irradiation develop significant histologic lung fibrosis within 52 weeks, whereas CBA and C3H mice do not exhibit substantial fibrosis during this time. The purpose of the present study was to determine whether this strain-dependent difference in radiation histopathology is associated with genetic differences in pulmonary endothelial metabolic activity or in endothelial radioresponsiveness. C57BL/6J, C57BL/10J, CBA/J, and C3H/HeJ mice were sacrificed 12 weeks after exposure to 0 or 14 Gy of 300-kV X rays to the whole thorax. Lung angiotensin converting enzyme (ACE) activity and plasminogen activator (PLA) activity were measured as indices of pulmonary endothelial function; and lung hydroxyproline (HP) content served as an index of pulmonary fibrosis. Lung ACE and PLA activities in sham-irradiated C57BL/6J and CB57BL/10J mice were only half as high as those in sham-irradiated CBA/J and C3H/HeJ mice. Exposure to 14 Gy of X rays produced a slight but nonsignificant reduction in lung ACE and PLA activity in the C57BL strains, and a significant reduction in the CBA/J and C3H/HeJ mice. Even after 14 Gy, however, lung ACE and PLA activities in CBA/J and C3H/HeJ mice were higher than those in sham-irradiated C57BL/6J and C57BL/10J mice. Lung HP content in all four strains increased significantly after irradiation, but this increase was accompanied by an increase in lung wet weight. As a result, HP concentration (per milligram wet weight) remained constant or increased slightly in both C57BL strains and actually decreased in the CBA/J and C3H/HeJ mice. These data demonstrate significant genetic differences in both intrinsic pulmonary endothelial enzyme activity and endothelial radioresponsiveness among the four strains of mice. Specifically, strains prone to radiation-induced pulmonary fibrosis (C57BL/6J, C57BL/10J) exhibit only half as much lung ACE and PLA activity as do strains resistant to fibrosis (CBA and C3H).     

Q- and C-Band Chromosome Markers in Inbred Strains of MUS MUSCULUS

Differences in the number of chromosomes with secondary constrictions and in the size of the C-band region on certain chromosomes have been observed among the following inbred strains of Mus musculus: C57BL/10J, C57BR/cdJ, DBA/1J, CBA/J, BALB/cJ, and AKR. These differences are useful as indicators of the location of rRNA genes and as normal chromosome markers. The size of each C-band region appears to remain constant over many generations. Only one probable change in the size of a C-band region was found.     

Optimal N-ethyl-N-nitrosourea (ENU) doses for inbred mouse strains

ENU is a powerful germline mutagen in the mouse, providing the opportunity to analyze the functions of large numbers of genes in the mammalian genome. In many mutagenesis experiments, it would be beneficial to exploit the advantages of inbred mouse strains. To perform an effective ENU mutagenesis screen using inbred mice, a dosage regimen is required to determine the optimal dose of ENU for that inbred strain, a time-consuming preliminary process. We have carried out dosage regimens for mutagenizing doses of ENU in ten inbred strains of mouse: 129X1/SvJ, 129S6/SvEv, A/J, BALB/cJ, BTBR/N, C3He/J, C3HeB/FeJ, C57BL/6J, C57BR/cdJ, and CBA/CaJ, and determined an optimal dose for each strain, defined by length of sterile period and number of males to survive treatment. Three strains: A/J, BALB/cJ and C57BL/6J, are able to tolerate high doses, up to 300 mg/kg body weight, and are highly recommended for mutagenesis studies.     

Effects of dietary fat on cholesterol movement between tissues in CBA/J and C57BR/cdJ mice.

Differences in dietary fats cause differences in cholesterol metabolism in mice. CBA/J mice are resistant to diet-induced hypercholesterolemia and atherosclerosis; they adjust hepatic hydroxymethyl-glutaryl-CoA reductase activity (HMGR) to maintain homeostasis; C57BR/cdJ mice are susceptible, but young animals are thought to maintain homeostasis by changing fecal excretion of sterols. Compartmental modelling of movement of [4-14C]cholesterol was used to analyze movement of cholesterol between serum and liver, heart, and carcass in mice fed 40 en% fat, polyunsaturated to saturated fatty acid ratio (P/S) = 0.24 (US74) or 30 en% fat, P/S = 1 (MOD). Dietary effects were quite pronounced, while strain effects were more subdued. The C57/cdJ animals appear to regulate the overall cholesterol balance by reducing synthesis, as do the CBA/J animals, even though synthesis is not reduced to the same degree as in the CBA/J animals. Both diet and strain influence the whole-animal turnover rate, with slower turnover occurring for C57BR/cdJ animals and animals fed the US74 diet.     

Linkage of isozyme loci in the mouse: Phosphoglucomutase-2 (Pgm-2), mitochondrial NADP malate dehydrogenase (Mod-2), and dipeptidase-1 (Dip-1)

The linkages of the isozyme genes Mod-2, Pgm-2, and Dip-1 have been determined in tests with established linkage group markers among inbred strains of mice. Unique alleles for both Mod-2 and Pgm-2 have been observed in the strain of SM/J. Linkage was determined from backcross progeny of the matings C57BL/6J×(SM/J×C57BL/6J)F₁, (SM/J×SWR/J)F₁×SM/J, and (SM/J×SWR/J)F₁×SJL/J. The gene Mod-2 is on linkage group 1. In a three-point cross of the loci Gpi-1, c, and Mod-2, the c locus was determined to be the middle gene. No double crossovers were observed. Our combined data show the following linkages: Gpi-1 to c, 28.3±3.2%; Gpi-1 to Mod-2, 33.3±3.0%; and c to Mod-2, 4.1±2.8%. The proposed gene order for four markers on LG I is Gpi-1-p-c-Mod-2. The gene Pgm-2 was linked to Gpd-1 (27.0±4.2%) on LGVIII. Two backcrosses segregating for Pgm-2 and b, (SM/J×DBA/2J) F₁×DBA/2J and (SM/J×DBA/2J)F₁×C57BR/cdJ, showed 9.1±4.3% recombination. The proposed gene order on LG VIII is b-Pgm-2-Gpd-1. The genes Pgm-1 and Pgm-2 are not linked (53.4±4.4%). Linkage of the isozyme genes Dip-1 and Id-1 on LG XIII was observed in backcross progeny of the crosses (SJL/J×C57BL/6J)F₁×SJL/J and C57BL/6J×(SM/J×C57BL/6J)F₁. The combined recombination was 23.8±2.8%. Two cases are established where genes whose enzyme products share substrate affinities (Pgm-1 and Pgm-2; Mod-1 and Mod-2) are not linked. Our data generally support the conclusion that functionally or metabolically related isozyme genes are not contiguous on mouse linkage groups.This investigation was supported in part by Public Health Service General Research Support Grant GM-09966 and in part by Public Health Service Training Grant 5T01 HD-00032-07 from the National Institute of Child Health and Human Development, and by Atomic Energy Commission contract AT(30-1)-3671.     

Association between Tph2 gene polymorphism, brain tryptophan hydroxylase activity and aggressiveness in mouse strains

The brain neurotransmitter serotonin is involved in the regulation of aggressive behavior. The main factor determining the brain serotonin level is the activity of the rate-limiting enzyme in the biosynthesis of the neurotransmitter--tryptophan hydroxylase isoform (TPH) 2 encoded by the Tph2 gene. Recently the C1473G single-nucleotide polymorphism in the Tph2 gene was reported. Here we study the C1473G polymorphism in 10 inbred mouse strains (C57BL/6J, AKR/J, DD/He, C3H/HeJ, YT/Y, BALB/cJLac, CC57BR/Mv and A/He) and demonstrate the association of the polymorphism with brain TPH activity and intermale aggressiveness. TPH activity in the midbrain of mice homozygous for the 1473C allele was higher than that in mice carrying 1473G alleles. A close association of the 1473C allele with increased number of attacks towards another male was found. The results support a link between the C1473G polymorphism in Tph2 gene, tryptophan hydroxylase activity and intensity of intermale aggression.     

Differential DNA-repair activity in prespermiogenic cells of various mouse strains

The present report demonstrates differential DNA-repair activity among 14 strains of immature (20 ± 2 days old) male mice (inbred strains: C57BL/6J, RF/J, Nude homo/nu, RIII/2J, Pl/J, AKR/J, Nude hetro/nude, C3H/HeJ, SWR/J, SM/J, ST/J, LP/J, BALB/cJ and random-bred strain: CD-1). The prespermiogenic cells were isolated and enriched by collagenase-trypsin digestion of seminiferous tubules and subsequent 3% albumin-gradient centrifugation. Enriched prespermiogenic cells demonstrated a viabiilty greater than 95% by trypan blue exclusion criteria. For in vitro unscheduled DNA synthesis (UDS) determination, prespermiogenic cells (10⁶ cells/ml) were incubated with methyl methanesulfonate (0.4 mM) in the presence of 20 mM hydroxyurea (HU). At 20 mM HU concentration, 90% of S-phase DNA activity in prespermiogenic cells was inhibited and thus, the net UDS activity following MMS exposure was readily determined. MMS-induced UDS activity in the CD-1 mouse strain was both linear up to 4 h of incubation and dose-dependent at 4 h incubation. The apparent K_m for MMS-induced UDS activity in prespermiogenic cells was approx. 1.8 × 10^?4 M. Of the 14 mice strains tested, C57BL/6J and RF/J exhibited the highest DNA-repair activity, while BALB/cJ, LP/J, and ST/J showed the lowest. A maximal difference in UDS activity fo 3.5-fold was observed between C57BL/6J and BALB/cJ. Furthermore, a 2.5-fold difference was also noted between RF/J and LP/J mouse strains. Thus, wide variations in DNA-repair activity among 14 mouse strans were clearly demonstrated. Whether genetically select mouse strains with the lowest DNA-repair activity should have greater sensitivity toward environmental mutagens needs to be tested.     

Genetic control of immune responses to the 18-kDa protein of Mycobacterium leprae. Different TH1 subsets may be involved in proliferative and delayed-type hypersensitivity responses.

In vitro and in vivo responses to the 18-kDa protein of Mycobacterium leprae have been analysed in different strains of mice. Lymphocytes from BALB/cJ (H-2d), BALB.B (H-2b), B10.BR (H-2k), and B10.M (H-2f) mice primed with 18-kDa protein yielded high T cell proliferative responses, while those from C57BL/10J (H-2b) mice yielded lower responses. Both H-2 and non-H-2 genes contributed to the magnitude of responsiveness. F1 mice from high and low responder strains showed high responsiveness to the 18-kDa protein. Supernatants from lymph node cell cultures prepared from 18-kDa protein-immunised BALB/cJ, B10.BR, and C57BL/10J mice contained IL-2 but no IL-4, indicating that activated T cells from both high and low responder mice were of a TH1 phenotype. Cell cultures from low responder C57BL/10J mice produced less IL-2 than those from high responders. The low responsiveness to the 18-kDa protein in proliferative assays might be due to a low frequency of antigen-specific T cells in the C57BL/10J mouse strain. BALB/cJ, C57BL/10J, and F1 (BALB/cJ x B10.BR) mouse strains were tested for in vivo DTH reactions to the 18-kDa protein. All strains, including C57BL/10J, were high DTH responders. Although DTH effector cells and 18-kDa protein-specific proliferative T cells belong to the TH1 subset, our data comparing high and low responder status indicate that distinct TH1 subpopulations are stimulated in response to the 18-kDa protein of M. leprae.     

Allotypes of mouse complement component C6 in inbred strains and some wild populations

This collaborative work was undertaken to resolve discrepancies in reports of the number of forms of complement component C6 present in the circulation of mice from various inbred strains. Plasma C6 was analyzed by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate and by isoelectric focusing (IEF), and C6 band patterns were developed by electroblotting and immunoprobing. Results of C6 allotyping of mice from 36 strains confirmed that while 20 strains (prototype strain BALB/c) possessed only one relative mass (M _r) form which typed C6A1 on IEF, the other 16 strains all possessed more than one C6 M _r form. Moreover, IEF analysis demonstrated additional polymorphic differences; among these 16 strains, 11 typed C6A l B l like the prototype strain CBA, the AKR and RF/J strains typed C6A2B2, and the Japanese MOM strain as well as the C57BR/cdJ and C57L/J strains possessed two forms with IEF mobilities intermediate between C6A1B1 and C6A2B2. These will now be referred to as C6A3B3. Thus, a total of four different mouse C6 haplotypes have been identified.Testing C6 allotypes in a limited number of wild mice revealed that haplotypes found in inbred strains of Western or Eastern origin tend to reflect haplotypes of the wild mice from Europe or Japan, respectively.     

Mouse senile amyloidosis. ASSAM amyloidosis in mice presents universally as a systemic age-associated amyloidosis.

Amyloid deposition in 11 inbred strains of mice (A/J, SJL/J, DDD, C57BL/6J, B10.BR, C57BL/10, B10A/SgSn, C3H/HeMs, B10A(5R), DBA/2 and C57BL/6Cr5/c) was studied using the peroxidase antiperoxidase (PAP) method and antisera against ASSAM and murine protein AA. Among the 170 mice examined, in 77 (45.3%) from the nine strains other than C3H/HeMs and DBA/2, there was evidence of spontaneous amyloid deposits in routine histological sections. Immunohistochemical studies using 54 mice with amyloid deposition, demonstrated ASSAM deposition in 45 mice (83.3%) in all nine strains, although the incidence and intensity of the deposition differed somewhat between strains. SJL/J and A/J had ASSAM deposits from the age of 8 months and the incidence increased with advancing age. In the other seven strains, ASSAM was first deposited at an older age than in the SJL/J and A/J strains. In A J, C57BL/6J, C57BL/10, B10.BR, B10A(5R) and C57BL/6Cr5/c, protein AA often coexisted with ASSAM. The distribution pattern of the ASSAM deposits was similar to that observed among the SAM strains. Thus, ASSAM is an ubiquitously distributed senile amyloid protein in the mouse. Determination of the molecular type of apoA-II, a serum precursor of ASSAM, among all 11 strains using the polymerase chain reaction (PCR) revealed the SAM-P/1 type apoA-II variant in SJL/J and A/J strains with a high susceptibility to ASSAM deposition. We concluded from this study that amino acid substitution in precursor apoA-II may be responsible for the early onset and severe amyloid deposition in the mouse.     

Genetic variation in erythrocyte NAD levels in the mouse and its effect on glyceraldehyde phosphate dehydrogenase activity and stability

During a screening of inbred strains for enzyme variation in glycolysis, differences were found in glyceraldehyde phosphate dehydrogenase (GAPDH) activity between C57BL/6J and SM/J mice. Segregation analysis did not reveal unequivocal monogenic inheritance. Dialysis of hemolysates caused decay of enzyme activity, especially in C57BL animals, which could be prevented by the presence of NAD. This led to the finding that erythrocyte NAD levels were threefold higher in SM than C57BL animals and, although additively inherited, did not appear to be monogenic. This is comparable with but independent of the differences and effect of erythrocyte NADP levels on glucose-6-phosphate dehydrogenase activity reported by R. P. Erickson [(1974) Biochem. Genet. 11:33] and emphasizes the range of mechanisms that can be involved in the genetic control of enzyme activity in mammalian systems.We thank the MRC for financial support.     

Androgen induction of beta-glucuronidase translational yield in submaxillary gland of B6.N mice

Androgens induce the synthesis of murine beta-glucuronidase (GUS) 10-fold in the submaxillary gland of B6.PAC-Gusn mice without a concomitant increase in GUS mRNA levels. Since the rate of GUS synthesis per mRNA molecule, or translational yield, is a function of both the efficiency with which the message is translated and the fraction of newly made polypeptides that are incorporated into GUS tetramers, we conclude that androgen induces at least one of these components in the submaxillary gland of B6.N mice. Genetic variation in the submaxillary gland induction response was tested for using six congenic mice strains, each carrying a different haplotype of the Gus gene complex on a C57BL/6J genetic background. The results indicate that the DNA sequences determining androgen responsiveness of the Gus gene in the submaxillary gland are linked to the Gus gene complex and that the DNA sequences determining the submaxillary gland response are distinct from those determining the androgen induction of GUS mRNA in kidney.     

Genetic Differences in Androgen Receptors and in Autoregulation of Testicular Human Chorionic Gonadotropin Binding Sites in the Mouse

Abstract

The autoregulation of testicular human chorionic gonadotropin (hCG) binding sites was studied in two strains of mice known to differ in their endocrine and reproductive characteristics (C57BL/10J and DBA/2J), and in their F₁ progeny (B10D2F₁). Basal hCG binding levels were higher in C57BL/10J than in DBA/2J mice, while B10D2F₁ mice had intermediate levels. Twenty-four h after injection, hCG produced dose-related changes in hCG binding in C57BL/10J and B10D2F₁ mice not observed in DBA/2J mice. However, 72 h after treatment with hCG there was a decrease in hCG binding in all the strains studied. These results suggest the participation of genetic factors in determining basal levels, dose-related changes and temporal response of testicular hCG binding sites to hCG administration. Androgen receptor levels were measured in the same strains of mice. DBA/2J mice had higher receptor levels in the kidney and coagulating gland, and lower levels in the hypothalamus and seminal vesicle when compred to C57BL/10J mice. B10D2F₁ mice had androgen receptor levels similar to those measured in C57BL/10J mice in all tissues studied, with the exception of the coagulating gland, where levels were similar to those observed in DBA/ZJ mice. These observations may indicate the existence of several loci coding for androgen receptors, with only one being expressed per tissue     

Dimethylnitrosamine metabolism: II. In vitro activation of dimethylnitrosamine by hepatic and renal tissues from crosses among BALB/c, DBA and C57BL mice

Crosses among BALB/c, C57BL and DBA mice were performed to investigate the genetic mechanisms involved in metabolism of DMN by renal and hepatic tissues. Liver S-9 fractions from parental strain DBA had the greatest potential to activate DMN and liver fractions from parental strain BALB/c had the lowest. No age or sex-related differences were observed within strain. Crossing of either C57BL or DBA to BALB/c mice resulted in F1 hybrids with liver microsomal enzymes that gave results similar to the BALB/c parental strain. There were no sex or age differences within crossbred strains in the potential of liver to activate DMN. In contrast male DBA and C57BL parental mice renal S-9 fractions did not differ significantly from each other but did differ significantly from male BALB/c renal fractions and from female and immature animals of all strains. Crossing of either DBA or C57BL mice with BALB/c mice resulted in male F1 hybrids whose renal S-9 fractions did not differ significantly from males of the parental BALB/c strain. In all instances, male renal S-9 fractions had a significantly greater potential to activate DMN than female or immature animals. F1 DBA X C57BL hybrids had renal S-9 fractions that did not differ significantly from the parental strains. These data suggest that the gene(s) for low DMN metabolism of BALB/c mice are apparently dominant over the genes from both DBA and C57BL. The exact genetic or physiological mechanism needs further elucidation.     

Molecular analysis of genetic differences among inbred mouse strains controlling tissue expression pattern of alcohol dehydrogenase 4

The ADH gene family in vertebrates is composed of at least seven distinct classes based upon sequence comparisons and enzyme properties. The Adh4 gene product may play an important role in differentiation and development because of its capacity to metabolize retinol to retinoic acid. Allelic gene differences exist among inbred mouse strains which control structure and tissue-specific regulation of Adh4. C57BL/6 mice are unique and have no detectable ADH4 enzyme activity in epididymis and low levels in seminal vesicle, ovary and uterus compared to other strains. C57BL/6 mice express Adh4 in stomach at levels similar to other strains. The goal of this research was to investigate this genetic variation at the molecular level. Northern analysis revealed that the content of ADH4 mRNA in tissues correlate with the enzyme expression pattern. Interestingly, C57BL/6 mice express an ADH4 mRNA in stomach which is smaller than expressed in C3H and other mice. An analysis of the 5'- and 3'-ends of the mRNA using RACE analysis determined that the ADH4 mRNA in C57BL/6 mice is truncated in the 3'-untranslated region. Sequence analysis of RACE products showed that the truncation is due to a single nucleotide mutation which produces an early polyadenylation signal. Additional RACE and Northern analysis revealed that at least five different polyadenylation sites are used in the Adh4 gene. Using 3'-end polymorphisms found between C57BL/6 and C3H strains and RT-PCR, it was shown that the lack of expression in epididymis in C57BL/6 mice is cis-acting in F(1) hybrid animals. The DNA sequence of the proximal promoter (-600/+42 nt) was determined in several mouse strains differing in tissue-specific expression patterns and did not reveal any nucleotide substitutions correlating with expression pattern suggesting further upstream or downstream sequences may be involved.