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  • 1
    ISSN: 1432-1432
    Keywords: Squalus acanthias ; Carbamoyl-phosphate synthetase ; Promoter ; Rana catesbeiana ; TATA box ; TACAAA ; C/EBP
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Carbamoyl-phosphate synthetase III (CPSase III) ofSqualus acanthias (spiny dogfish) is a nuclear-encoded mitochondrial enzyme that catalyzes glutamine-dependent formation of carbamoyl phosphate for urea synthesis. In this paper we report the results of cloning a 10-kb segment of genomic DNA which includes the region flanking the 5′ end of the spiny dogfish CPSase III gene. A total of 1,295 base pairs of sequence straddling the start codon was obtained. Primer extension experiments revealed that the transcription start site is the G located 114 residues upstream of the translation start codon ATG. The first exon has 240 base pairs, including the 5′ untranslated region, the coding sequence for the signal peptide (38 amino acids), and the four N-terminal amino acids of the mature enzyme. The boundary of the first exon and the first intron of the CPSase III gene is concordant with that of rat and frog (Rana catesbeiana) CPSase I, which have been suggested to have evolved from CPSase III. The putative TATA box sequence, TACAAA, is located at position −31 with an uncommonly found C at the third position. Two C/EBP binding site sequences, ATTCTGCAAG (−405 to −397) and GTGCAGTAAG (−168 to −160), were identified in the promoter region, which suggests that spiny dogfish CPSase III might be subjected to transactivation of transcription by C/EBP-related proteins, as has been reported for rat CPSase I. The preparation and binding of a recombinant RcC/EBP-1 protein (theR. catesbeiana homolog of the mammalian C/EBPα) to the two spiny dogfish C/EBP binding sequences are described. Two putative heatshock binding elements were also identified in the promoter region.
    Type of Medium: Electronic Resource
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  • 2
    ISSN: 0192-253X
    Keywords: Thyroid hormone ; carbamyl phosphate synthetase ; Rana catesbeiana ; metamorphosis ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: During both spontaneous and thyroid hormone (TH)-induced metamorphosis, the Rana catesbeiana tadpole undergoes postembryonic developmental changes in its liver which are necessary for its transition from an ammonotelic larva to a ureotelic adult. Although this transition ultimately results from marked increases in the activities and/or de novo synthesis of the urea cycle enzymes, the precise molecular means by which TH exerts this tissue-specific response are presently unknown. Recent reports, using RNA from whole Xenopus laevis tadpole homogenates and indirect means of measuring TH receptor (TR) mRNAs, suggest a correlation between the up-regulation of TRβ-mRNAs and the general morphological changes occurring during amphibian metamorphosis. To assess whether or not this same relationship exists in a TH-responsive tissue, such as liver, we isolated and characterized a cDNA clone containing the complete nucleotide sequence for a R. catesbeiana urea cycle enzyme, ornithine transcarbamylase (OTC), as well as a genomic clone containing a portion of the hormone-binding domain of a R. catesbeiana TRβ gene. Through use of these homologous sequences and a heterologous cDNA fragment encoding rat carbamyl phosphate synthetase (CPS), we directly determined the relative levels of the TRβ, OTC, and CPS mRNAs in liver from spontaneous and TH-induced tadpoles. Our results establish that TH affects an up-regulation of mRNAs for its own receptor prior to up-regulating CPS and OTC mRNAs. Moreover, results with cultured tadpole liver demonstrate that TH, in the absence of any other hormonal influence, can affect an up-regulation of both the TRβ and OTC mRNAs. © 1992 Wiley-Liss, Inc.
    Additional Material: 9 Ill.
    Type of Medium: Electronic Resource
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