Two Distinct Pancreatic Amylase Genes Are Active in Ybr Mice

The genetic determinants of pancreatic amylase expression in YBR mice differ in two respects from those of other inbred strains. First, there are two nonallelic amylase isozymes present in YBR pancreas, while most mouse strains express a single pancreatic amylase protein. In addition, the in vivo rate of total pancreatic amylase synthesis is 50% of that in other strains. Both these traits are determined by genetic sites in the region of the Amy-2 locus on mouse chromosome 3. To determine the molecular basis for the presence of two isozymes in this strain, we have compared portions of their amino acid sequences. Two differences between isozymes A₁ and B₁ were identified among the 77 residues compared. This result demonstrates that two distinct amylase genes are expressed in YBR pancreas.     

Simultaneous expression of salivary and pancreatic amylase genes in cultured mouse hepatoma cells.

The tissue-specific expression of two types of mouse amylase genes does not overlap in vivo; the Amy-1 locus is transcribed in the parotid gland and the liver, while expression of Amy-2 is limited to the pancreas. We identified a mouse hepatoma cell line, Hepa 1-6, in which both amylase genes can be simultaneously expressed. Amy-1 is constitutively active in these cells and is inducible by dexamethasone at the level of mRNA. We demonstrated that the liver-specific promoter of Amy-1 is utilized by the dexamethasone-treated hepatoma cells, and that glucocorticoid consensus sequences are present upstream of this promoter. Amy-2 is not detectable constitutively, but can be activated if the cells are cultured in serum-free medium containing dexamethasone. Expression of Amy-2 in a nonpancreatic cell type has not previously been observed. We speculate that induction of Amy-1 and activation of Amy-2 may involve different regulatory mechanisms. Hepa 1-6 cells provide an experimental system for molecular analysis of these events.     

Inheritance of a Parotid Secretory Protein in Mice and Its Use in Determining Salivary Amylase Quantitative Variants

Among inbred strains of mice, a major protein, PSP, produced and secreted by the parotid glands, shows variation in electrophoretic mobility and in the peptides produced by cyanogen bromide treatment. This variation is inherited as a single Mendelian factor with two alleles showing co-dominant expression. In genetic crosses, it segregates independently from the amylase complex and is closely linked to the agouti locus on chromosome 2. The protein ratios between amylase and PSP in saliva, obtained by scanning of electrophoretic gel separations, were found to reflect genetic differences in salivary amylase production in strains YBR/Cv and C3H/As.     

Independent regulation of nonallelic pancreatic amylase genes in diabetic mice

Administration of streptozotocin produces a diabetic condition in mice characterized by a specific decrease in amylase synthesis in the pancreas as well as a substantial reduction in amylase mRNA concentration. We have studied this effect in mice of the congenic strains C3H.AmyYBR and C3H.AmyCE with multiple active copies of the pancreatic amylase structural gene. When mice of these strains are treated with streptozotocin, the magnitude of reduction in the synthesis of each amylase isozyme is different. These differences are reflected in the relative activities of isozyme-specific mRNAs in an in vitro translation assay. Administration of insulin results in partial restoration of normal phenotypes. The results provide genetic evidence that individual copies of the amylase structural gene are associated with divergent cis-acting insulin-responsive sequences.     

Electrophoretic behavior of amylase in mouse: the parotid gland is major source of serum amylase

Amylase activity in the saliva, salivary glands, serum, liver (perfused and non perfused) and pancreas was assayed and isoamylases were separated by electrophoresis in these organs using C57BR/cdJ and M. m. molossinus (Kor) mice. Amylase isozymes in the saliva, parotid gland, serum and liver were identical in both strains, respectively. Amylase activity in the liver was lower than that in the serum and liver amylase disappeared almost by perfusion. Major serum amylase was released from the parotid gland in intact animals.     

Multiple non-allelic genes encoding pancreatic alpha-amylase of mouse are expressed in a strain-specific fashion.

The number of active Amy-2 genes has been estimated in strain CE/J mice which produce four distinct electrophoretic forms of alpha-amylase in their pancreas. cDNA cloning and DNA sequence analysis discloses five distinct mRNA sequences which differ by approximately 1% of their nucleotides. Two of these mRNAs specify the same protein. Changes in the nucleotide sequences result in amino acid replacements that alter the net charges of the deduced proteins. This has allowed a tentative assignment of individual mRNAs to isozymes detected by electrophoresis. Quantitative Southern blot hybridization using a DNA probe specific for the first exon of Amy-2 reveals the presence of greater than 10 Amy-2 related sequences per haploid CE/J genome. Models which could account for the mouse strain-specific differences with respect to the number of pancreatic alpha-amylase isozymes and their variable but genetically determined quantitative ratios are discussed.     

Genetic variation in mouse salivary amylase rate of synthesis

Heterozygotes from matings of the mouse strains YBR/Cv and C3H/As have about 3 times more YBR-amylase than C3H-amylase in the saliva. The determinant for this quantitative effect is located on linkage group XVI close to or within the structural gene for salivary amylase. The quantitative effect is the result of an increase in the rate of synthesis of YBR-amylase, and the determinant is cis acting. Studies of other mouse strains suggest that regulatory genetic elements may modulate salivary amylase production.This work was supported by the Danish Natural Science Research Council and a grant from the United States Public Health Service (Grant GM-19521).     

Genetic variation of amylases in deer mice

The inheritance of salivary amylase variants in the deer mouse, Peromyscus maniculatus, is controlled by codominant alleles, Amy-1a, Amy-1b, and Amy-1c, at a single autosomal locus. Pancreatic amylases were invariant and unaffected by salivary amylase genotypes. Salivary amylase zymograms of five myomorphic rodents are compared and evolutionary implications are discussed.     

Genetic Regulation of SN-Glycerol-3-Phosphate Dehydrogenase in Brown Adipose Tissue

The levels of sn-glycerol-3-phosphate dehydrogenase (GPDH) were determined in the brown adipose tissue (BAT) of different inbred strains of mice. The BAT of the BALB/cJ strain contains twice as much enzyme activity per milligram protein as do other strains. The appearance of this difference is developmentally dependent, since it is not detected in BAT until 25-30 days postpartum. Genetic analysis of this strain difference has shown that the mechanism of inheritance involves at least two genes, one of which is linked to the Gdc-1 structural locus on chromosome 15. Determinations of GPDH synthesis by immunoprecipitation of GPDH protein labeled in vivo with [3H]leucine, and of GPDH mRNA by Northern blot analysis, establish that in BALB/cJ mice higher rates of enzyme synthesis are determined by elevated levels of GPDH mRNA. It was also found that cold stress increases GPDH mRNA levels in all the strains examined.