ABSTRACT
The liver of a young girl which had been successfully transplanted was investigated at the ornithine transcarbamylase (OTC, EC 2.1.3.3) gene expression level. Northern blot hybridization using a human OTC cDNA probe showed a greater than 80% decrease in specific OTC mRNA although having the same molecular size as a normal control. OTC polypeptide was simultaneously synthesized with a normal molecular size but at a low level (20%) as shown by immunoblotting. The OTC enzyme from the deficient liver exhibited very little catalytic activity (7.2% as compared to the normal subject). These results may support several explanations of this disease such as mutation of the OTC gene promoter leading to a low transcriptional activity or mutation of the encoding sequence which results in a modified translation product but with a normal size. mRNA instability may also occur.
Subject(s)
Mitochondria, Liver/enzymology , Ornithine Carbamoyltransferase Deficiency Disease , Blotting, Northern , Catalysis , Child, Preschool , Cloning, Molecular , DNA/metabolism , Female , Humans , Nucleic Acid Hybridization , Ornithine Carbamoyltransferase/analysis , Ornithine Carbamoyltransferase/metabolism , Peptides/analysis , Peptides/chemistry , RNA/analysis , RNA, Messenger/metabolismABSTRACT
Ovariectomized guinea-pigs were treated with oestradiol-17 beta (ES), oestrone sulphate (E1S) and progesterone (P) and the in-vitro incorporation of 35SO4 was studied in uterine fragments. The net uptake of 35O4 into tissue was only increased by oestradiol-17 beta plus progesterone. The incorporation of 35SO4 in the tissue-associated proteins was increased after treatment with E2 and E1S compared with untreated controls (3.1- and 2.5-fold, respectively). For secreted proteins, all hormone treatments induced an increase in protein sulphation, the highest increase occurring when progesterone was administered after oestrogens. Tyrosine 35SO4 was identified in protein extracts from tissues and media and values were greater after hormone treatments. The biggest increase in tyrosine 35SO4 was observed in secreted proteins in the E1S + P treatment group. The patterns of 35S-sulphate-labelled proteins were examined by SDS-polyacrylamide gel electrophoresis. In tissue extracts, the most striking differences related to the hormone treatments were observed in the Mr 94,000-190,000 region. A sulphated protein band of Mr 102,000 was specifically found in the E2 + P group and a band of Mr 125,000 only in the E1S + P group. The Mr 125,000 band was also found in tissue proteins from the E1S + P-treated animals after the incorporation of 35SO4 in vivo. This protein band may be a marker of the action of oestrone sulphate plus progesterone. For secreted proteins, those with a molecular weight greater than 100,000 were more abundant in the oestrogen plus progesterone-treated groups than in the oestrogen-treated groups. The content of tyrosine sulphate in each protein band ranged from 8 to 25% of the total radioactivity. No protein sulphated exclusively on the tyrosine residues was found. These studies provide the first description of the effects of steroid hormones on sulphated proteins in the guinea-pig uterus and suggest that oestrone sulphate is a potent biologically active hormone in the uterus.
Subject(s)
Gonadal Steroid Hormones/pharmacology , Proteins/metabolism , Sulfates/metabolism , Uterus/metabolism , Animals , Culture Techniques , Estradiol/pharmacology , Estrone/analogs & derivatives , Estrone/pharmacology , Female , Guinea Pigs , Ovariectomy , Progesterone/pharmacology , Tyrosine/metabolism , Uterus/drug effectsABSTRACT
Tyrosine sulfation was studied in guinea-pig uterus by in vitro labelling with [35S]sulfate, after estradiol-17 beta (E2) and E2 plus progesterone (P) treatment. [35S]Sulfated tyrosine was identified in tissue and secreted proteins, ranged from 9.3 to 21.0% of total protein sulfation and was higher in secreted proteins than in tissue proteins. Sulfate incorporation into tyrosine increased with hormone treatments. The highest level was found in secreted proteins under the combined effect of E2 plus P. The effect of P may be related to both the increase of cellular uptake of sulfate and the increase of tyrosine sulfation of secreted proteins. These results are consistent with the effect of P on endometrium secretions.
Subject(s)
Tyrosine/metabolism , Uterus/metabolism , Animals , Estradiol/pharmacology , Female , Guinea Pigs , In Vitro Techniques , Progesterone/pharmacology , Proteins/metabolism , Sulfates/metabolism , Uterus/drug effectsABSTRACT
In vitro metabolism of androstenedione in gonads of juvenile and adult Helix aspersa has been investigated. The conversion of [3H]androstenedione into testosterone, 5 alpha-dihydrotestosterone, androsterone, and estriol was demonstrated. In juvenile animals testosterone (59.8%) is the major metabolite whereas in adult animals androsterone (18.8%) is. The following endogenous steroids have been identified by gas chromatography-mass spectrometry in adult gonads: androsterone, dehydroepiandrosterone, androstenedione, 3 alpha-androstanediol, estrone, estradiol-17 beta, and estriol. The levels of testosterone, 5 alpha-dihydrotestosterone, androstenedione, and progesterone have been measured by RIAs in gonads and hemolymph. Their levels vary with the physiological stage: the gonadal and circulating levels of testosterone decrease with the sexual maturation whereas the 5 alpha-dihydrotestosterone increases. These differences observed in metabolism and in level of steroids between the juvenile and the adult snails allow us to suppose that these steroids have a biological role.