ALBERT

Notes: [Auszug] TWO single gene mutations in the mouse show similar timing of expression during post-natal development. The two mutant al eles were found to produce nearly normal phenotypes until some critical time after birth following which normal differentiation is arrested. The first of these genes, originally ...

Notes: [Auszug] “A mouse is miracle enough to stagger sextillions of infidels” Leaves of Grass, Walt Whitman The mouse has become our experimental surrogate. It is the creature we turn to to do the experiments, so important in reaching an understanding of ourselves, that are either technically ...

Notes: [Auszug] A genetic complex, termed [Bgl], mapping near the Mod-l locus on chromosome 9, contains within it elements governing the structure, development and rate of synthesis of /3-galac-tosidase4'9"15. The structural gene Bgl-e is defined by an electrophoretic polymorphism which affects the enzyme in all ...

Notes: Abstract A gene complex consists of a structural gene with its associated regulatory information; together they behave as the functional and evolutionary unit of mammalian chromosomes. The use of congenic lines, in which alternate forms, or haplotypes, of a gene complex are transferred into a common genetic background by repeated backcrossing, provides a means of comparing the regulatory properties of different haplotypes of a gene complex without the complications introduced by extraneous genetic differences. We have now carried out such a study of the A, B, and H haplotypes of the β-glucuronidase gene complex, [Gus], in mice. These haplotypes were derived from strains A/J, C57BL/6J, and C3H/HeJ and were compared against the C57BL/6J genetic background. Enzyme structure was compared in terms of charge (isoelectric point), stability (rate of thermal denaturation), substrate affinity (for 4 MU glucuronide), and antigenicity (reactivity with a standard antibody). Compared to the B form, the enzyme coded by the A haplotype has a lower isoelectric point, and that coded by the H haplotype is less stable. The decreased stability is the result of a lower activation energy for the thermal denaturation reaction. These differences were maintained in the congenic strains. All three enzyme forms showed identical substrate affinities. Antigenicity per enzyme unit was also identical for all three, indicating that none lacks an antigenic site possessed by the others and that they all possess the same catalytic activity per molecule. The expression of alleles of the Gus-t temporal locus within the gene complex was not affected by transfer into the C57BL/6 genetic background. The same developmental switches in enzyme activity were seen in each case. Transfer into the C57Bl/6 background also did not affect expression of the Gus-r regulator determining androgen inducibility of β-glucuronidase synthesis in kidney epithelial cells. However, enzyme accumulation in induced cells was altered when the haplotypes were transferred into the C57BL/6 genetic background. Since the rate of synthesis was not affected, it suggests that the genetic differences between strains that are not linked to the [Gus] complex affect the rate of enzyme loss by degradation or secretion. β-Glucuronidase in liver is present in both lysosomes and endoplasmic reticulum (microsomes). The relative amount of enzyme at each site depended on both the indentity of the structural allele and the function of unlinked genetic modifiers. Within the C57BL/6 background the percentage of total enzyme present in the microsome fraction was the order A〉B〉H. For the H form of the enzyme the percentage was appreciably greater in the C3H genetic background compared to C57BL/6. As expected, then, the [Gus] complex contains all of the genetic determinants of enzyme structure detected by thermal stability and isoelectric point measurements. Additionally, the complex contains all of the genetically determined differences between strains in the regulation of β-glucuronidase synthesis, including the programming of synthesis during development and the responsiveness of the [Gus] complex to hormonal stimulation. In contrast, genetic determinants of posttranslational processing are located elsewhere, including factors affecting enzyme localization and secretion/degradation. These results illustrate the utility of congenic strains for minimizing other genetic variables in characterizing the regulatory properties of alternate haplotypes of a gene complex.

Notes: Abstract The magnitude and kinetics of β-glucuronidase induction in mouse kidney are determined by a cis-acting regulatory gene, Gus-r, that is closely linked to the enzyme structural gene. The accumulation of β-glucuronidase mRNA during induction is much slower than the turnover time of the mRNA, suggesting progressive acquisition of mRNA synthesizing capacity during induction. Counts of the numbers of induced cells present at various times of induction in strains carrying three different alleles of Gus-r show that all potentially responsive cells respond immediately. The level of induction is progressive in individual cells and does not involve continued recruitment of new cells into the induced population. It appears that during induction each chromosome becomes progressively more active in directing the synthesis of β-glucuronidase.

Notes: Abstract A set of recombinant-inbred strains developed from mouse strains BALB/c and C57BL/6 includes two β-glucuronidase phenotypes that are not seen in either of the progenitor strains. These new recombinant phenotypes indicate that glucuronidase levels are regulated by genes additional to the Gur locus, which is closely linked to the glucuronidase structural gene (Gus) and is known to regulate the rate of glucuronidase synthesis. In this study, induced rates of glucuronidase synthesis were determined for these recombinant-inbred strains, and rate constants for enzyme loss were calculated. The rate of synthesis was found to segregate with the Gus gene in all of the strains, and only the determinants for rate of enzyme loss recombined to give new phenotypes. It was concluded that at least two genes affect the rate of enzyme loss, that these genes are not closely linked to each other or to the Gur-Gus region on chromosome 5, and that no major determinants of glucuronidase synthesis segregate independently of Gur.

Notes: Abstract Both enzyme activity and mRNA concentration of β-glucuronidase were measured in kidneys of mice treated with testosterone and the synthetic estrogen, diethylstilbestrol. Six congenic strains, all having a C57BL6/J genetic background but each having a different haplotype of the β-glucuronidase gene complex, were compared. In each strain the induction caused by androgen was partially repressed by estrogen. The extent of this antagonism varied among the six haplotypes and was not coordinate with the extent of induction by androgen alone. Antagonism appears to be regulated by at least two alleles of a new locus,Gus-e, within the β-glucuronidase gene complex. Repression by estrogen, like induction by androgen, appears to take place primarily at the transcriptional level. Kinetic studies revealed that estrogen causes the androgen response curve to plateau earlier and at a lower level. This suggests that estrogen increases the rate of gene deactivation rather than decreasing the rate of gene activation, Isoelectric focusing of β-glucuronidase fromGus-e a andGus-e b mice and their F1 progeny revealed that the genes are regulated incis. Together, these findings support a model in which both sex hormones exert their effects on separate DNA response elements located in close proximity to the gene or within the gene itself.

Notes: Abstract Previous studies have suggested that the binding of mouse glucuronidase to endoplasmic reticulum membrane is stabilized by the membrane protein egasyn. Using a radioimmunoassay for egasyn, we have now examined the inheritance of egasyn levels in mice. Mice of the ibred strain C57BL/6J, which have normal levels of microsomal glucuronidase, contained 56±10 μg egasyn per gram of liver. Mice of the inbred strain YBR, which carry the Eg 0 mutation resulting in the absence of microsomal glucuronidase, did not contain detectable levels of egasyn. The F1 progeny of these two strains contained intermediate levels of egasyn, 25±4 μg egasyn per gram of liver. Progeny from the backcross of these F1 animals to YBR were distributed equally into two discrete phenotypic classes. One class lacked both egasyn and microsomal glucuronidase, while the other class contained 25±3 μg egasyn per gram of liver and contained normal levels of microsomal glucuronidase. Thus egasyn levels are determined by the Eg locus and show additive inheritance. These results suggest that the Eg gene codes for egasyn and that it is the inability to produce egasyn that results in a deficiency of microsomal glucuronidase in the Eg 0 mutant.

Notes: Abstract Acid hydrolases are present in normal human urine in appreciable amounts. Their source appears to be lysosomes released by kidney proximal tubule epithelial cells. For a given lysosomal enzyme the total amount excreted is the product of two parameters, a general one describing the rate of lysosome secretion and a specific one describing the relative concentration of that enzyme in lysosomes. There is considerable population variation in both parameters. Studies of β-glucuronidase, β-galactosidase, β-hexosaminidase, and α-galactosidase in monozygotic and dizygotic twins show that an appreciable part of this variation is genetic in origin. This appears to be true for both total enzyme excretion and lysosome composition. Although it was not possible to test directly whether this is also true for the rate of lysosome secretion, the fact that the two former parameters are both heritable strongly suggests that the rate of lysosome excretion is also a heritable trait. Taken together with previous data, the results suggest polygenic control of these biochemical traits. It is particularly significant that β-glucuronidase excretion in normal individuals is a heritable trait since the excretion of this enzyme has frequently been used as a measure of normal and pathological physiological changes.