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Biomedical subjects

I M Schaefer

Publications and source records attributed to I M Schaefer.

8 recordsLinked to original sources
PubMed1992

Tissue-specific, developmental, hormonal, and dietary regulation of rat phosphoenolpyruvate carboxykinase-human growth hormone fusion genes in transgenic mice.

The cytosolic phosphoenolpyruvate carboxykinase (PEPCK) gene is expressed in multiple tissues and is regulated in a complex tissue-specific manner. To map the cis-acting DNA elements that direct this tissue-specific expression, we made transgenic mice containing truncated PEPCK-human growth hormone (hGH) fusion genes. The transgenes contained PEPCK promoter fragments with 5' endpoints at -2088, -888, -600, -402, and -207 bp, while the 3' endpoint was at +69 bp. Immunohistochemical analysis showed that the -2088 transgene was expressed in the correct cell types (hepatocytes, proximal tubular epithelium of the kidney, villar epithelium of the small intestine, epithelium of the colon, smooth muscle of the vagina and lungs, ductal epithelium of the sublingual gland, and white and brown adipocytes). Solution hybridization of hGH mRNA expressed from the transgenes indicated that white and brown fat-specific elements are located distally (-2088 to -888 bp) and that liver-, gut-, and kidney-specific elements are located proximally (-600 to +69 bp). However, elements outside of the region tested are necessary for the correct developmental pattern and level of PEPCK expression in kidney. Both the -2088 and -402 transgenes responded in a tissue-specific manner to dietary stimuli, and the -2088 transgene responded to glucocorticoid stimuli. Thus, different tissues utilize distinct cell-specific cis-acting elements to direct and regulate the PEPCK gene.

Adipose Tissue↗
PubMed1991

Culture at high density increases phosphoenolpyruvate carboxykinase messenger RNA in H4IIEC3 hepatoma cells.

We report that the concentration of phosphoenolpyruvate carboxykinase (PEPCK) mRNA increased 5- to 10-fold when H4IIEC3 rat hepatoma cells were cultured at high compared to low density. The magnitude and direction of this response were mRNA specific, as the mRNAs encoding tyrosine aminotransferase and albumin increased approximately 20%, whereas the mRNAs encoding beta-actin and alpha-tubulin decreased 40% and 20%, respectively. Paracrine or autocrine mechanisms were not responsible for the density effect, since conditioned medium or frequent medium changes had only a modest effect on the abundance of PEPCK mRNA. Culture of H4IIEC3 cells at low density or on collagen promoted a flattened morphology and low PEPCK mRNA levels. At high density, cells assumed a cuboidal shape on both plastic and collagen and expressed high PEPCK mRNA levels. Induction of PEPCK mRNA by high density culture did not involve increased intracellular cAMP, since treatment with 8-(4-chlorophenylthio)-cAMP was synergistic with density. High cell density increased PEPCK run-on transcription approximately 3-fold, while PEPCK mRNA increased more than 6-fold. These observations suggest that high culture density increases PEPCK mRNA by increasing its transcription and possibly stabilizing PEPCK mRNA. The response could be coupled to the regulation of cell shape, as a close relationship between cell shape and gene expression has previously been shown to be important in the development and maintenance of tissues and organs. The PEPCK gene in H4IIEC3 cells could provide a useful model in which to study the poorly understood mechanisms involved in coordinating form and function.

Animals↗
PubMed1986

Characterization of the two nonallelic genes encoding mouse preproinsulin.

We have cloned and sequenced the two mouse preproinsulin genes. The deduced amino acid sequences of the mature mouse insulins are identical to the published protein sequences. However, the nucleotide sequence indicates that the mouse I C-peptide has a deletion of two amino acids compared with the mouse II C-peptide. We used an S1 nuclease assay to confirm the presence of the deletion and to measure the ratio of transcripts from gene I to transcripts from gene II. The mouse preproinsulin I gene, like the rat gene I, is missing the second intervening sequence that normally interrupts the C-peptide region in other insulin genes. Comparison of the 5' flanking sequences of the mouse and rat genes II indicates that they are homologous for at least 1000 base pairs. The preproinsulin I genes also share homology in their 5' flanking DNAs; however, their homology to the preproinsulin II genes extends for only about 500 base pairs.

Amino Acid Sequence↗
PubMed1984

Mouse kidney renin gene is on chromosome one.

The structural gene for mouse kidney renin (Ren-1) was localized to chromosome 1 by Southern blot analysis of mouse-hamster somatic cell hybrids with a cloned mouse submandibular renin cDNA probe. The submandibular renin gene (Ren-2) also lies on chromosome 1; so it may be, in those mouse lines which carry it, a tandem duplication of the kidney-type Ren-1 gene.

Animals↗
PubMed1984

Human renin gene is on chromosome 1.

DNA sequences encoding kidney renin were localized to region p21----qter of human chromosome 1 by Southern blot analysis of mouse-human somatic cell hybrids with a cloned human renin DNA probe. The renin gene may be a member of a chromosome 1 linkage group which is conserved in mouse and man. Available evidence suggests this gene is present in one copy per haploid genome. Thus those renin-like molecules detected immunologically in tissues other than the kidney (such as brain, placenta, uterus, pituitary, vasculature, and adrenal) may be derived from this single gene. Since renin messenger RNA in human kidney is about 1550 nucleotides long, reported molecular weights in excess of 45,000 for circulating renin represent posttranslational or postsecretory modifications of the polypeptide.

Chromosome Mapping↗
PubMed1984

Human renin gene: structure and sequence analysis.

The complete protein precursor of human kidney renin has been determined from the sequence of cloned genomic DNA. The gene spans 12 kilobases of DNA and is interrupted by eight intervening sequences. The nine regions (exons) encoding the protein were mapped with a mouse renin cDNA probe, synthetic oligonucleotide probes, and by hybridization of genomic restriction fragments to a 1600-nucleotide human kidney mRNA. The predicted 403-amino acid preprorenin consists of mature renin and a 66-residue amino-terminal prepropeptide. The DNA sequence 5' to the first exon indicates the location of a transcriptional promoter (T-A-T-A-A-A) for a mRNA encoding preprorenin. An additional transcriptional promoter site is located within the first intron, which, if used, would express a shortened nonsecreted prorenin. The structure of the human renin gene is similar to that of human pepsinogen, a closely related aspartyl protease enzyme. This observation suggests that renin and pepsinogen have a common evolutionary origin.

Amino Acid Sequence↗
PubMed1984

Construction and partial characterization of recombinant cDNA clones for chicken type I collagen messenger RNAs.

Messenger RNAs for the alpha 1 and alpha 2 chains of type I procollagen were partially purified from total embryonic chicken calvaria using gel chromatography on Sepharose 4B and used to construct recombinant cDNA clones corresponding to both mRNAs. Restriction site mapping, nucleotide sequencing and hybridization to RNA blots were used to show that clones pCAL1 and pCAL2 contain inserted sequences corresponding to the mRNAs for chicken alpha 1 and alpha 2 procollagen chains, respectively.

Animals↗