Richard Gagné

Assignment of the human 3β-hydroxysteroid dehydrogenase gene (HSDB3) to the p13 band of chromosome 1

Three-beta-hydroxysteroid dehydrogenase (HSDB3) is the enzyme which catalyses the oxidative conversion of delta 5-3 beta-hydroxy steroids to the delta 4-3-keto configuration and is therefore involved in the biosynthesis of all classes of hormonal steroids, namely progesterone, glucocorticoids, mineralocorticoids, androgens, and estrogens. Deficiency of the enzyme is associated with congenital adrenal hyperplasia and is usually lethal in early life. Despite its crucial role, chromosome assignment of the gene for this enzyme has not been reported. Using in situ hybridization, we report that hybridization with labeled human HSDB3-specific cDNA yielded 27% of silver grains associated with chromosome 1 with a maximal concentration in the p13 band.

Detection of succinylacetone and the use of its measurement in mass screening for hereditary tyrosinemia

A technique designed to measure quantitatively succinylacetone (4,6-dioxoheptanoic acid) is presented. It essentially involves the inhibition of delta-aminolevulinate dehydratase (EC 4.2.1.24) by succinylacetone. Prior to their use in the assay, the samples are heated at 100 degrees C for 30 min in order to transform all succinylacetoacetate (3,5-dioxooctanedioic acid) to succinylacetone. By this transformation of the first abnormal metabolite specific to hereditary tyrosinemia to the second and last one, which is a powerful inhibitor of delta-aminolevulinate dehydratase, we determine in one sensitive assay the total amount of both. Succinylacetone was measured in sera and urines from 19 patients with hereditary tyrosinemia. All sera and urines contained succinylacetone at concentrations ranging, respectively, from 2 to 100 mumol/l and from 190 to 6000 mumol/g creatinine. The technique was also adapted to dried blood spots on paper and was used as a test complementary to blood tyrosine determination in mass screening for hereditary tyrosinemia. A total of 2412 samples having concentrations of 60 mg/l or more of tyrosine were assayed, and ten showed the presence of succinylacetone. These were all from newborns with hereditary tyrosinemia. The test has proven to virtually eliminate false positives, and, thereby, much clerical work and parental anxiety.

Aspect cytologique et localisation intranucléaire de l'hétérochromatine constitutive des chromosomes C9 chez l'homme

Using a specific technique which stains the secondary constriction of number 9 chromosomes in man, we observed that this segment frequently appears to be composed of multiple small units packed together in metaphase as well as in interphase. During prophase of the first meiotic division in man, this segment looks like a diffuse structure, suggesting division and multiplication of these units. On the other hand, the C9 bodies observed in interphase with this cytological procedure are frequently associated with the nucleolus. This heterochromatin is less compact than previously shown and may represent repetitive DNA with a specific role.

The human corticosteroid binding globulin gene is located on chromosome 14q31-q32.1 near two other serine protease inhibitor genes

SummaryHuman corticosteroid binding globulin (CBG) cDNA fragments were radiolabeled and hybridized in situ to metaphase chromosome preparations. The results localized the CBG gene to the q31–q32.1 region of human chromosome 14. This location also contains the genes for two closely related serine protease inhibitors: alpha1-proteinase inhibitor and alpha1-antichymotrypsin. It is therefore likely that these genes evolved by duplication events, and it would appear that this region contains a series of functionally related genes.

Purification, Cloning, Complementary DNA Structure, and Predicted Amino Acid Sequence of Human Estradiol 17β‐Dehydrogenase

Seventeen-P-hydroxysteroid dehydrogenase ( 17P-HSD) is the enzyme that catalyzes the interconversion of dehydroepiandrosterone (DHEA) and Sandrostenediol, 4-androstenedione and testosterone, and estrone and estradiol. This enzyme is present in many tissues such as the placenta,’,? testis,3 kidney,4 liver,’ skin? and ovary.’ This enzyme is likely to play a key role in the regulation of intracellular concentrations of sex hormones. The androgen testosterone and the estrogen 17Pestradiol are known to play an essential role in the development, growth, and function of all tissues responsible for reproduction and fertility in men and women. Moreover, sex steroids are involved in the regulation of hormone-sensitive cancer growth, especially prostate,* b r e a ~ t , ~ and endometrial cancer.IO Despite its purification from many tissues”-I4 and its partial biochemical chara c t e r i ~ a t i o n , ~ ~ ’ ~ little was known about the structure of this enzyme. We report the purification, cloning, and deduced amino acid sequence of estradiol 17pdehydrogenase (E2DH) from human placenta. The availability of cDNAs for EzDH offers the opportunity of studying in detail the factors controlling the expression of this crucial enzyme not only in gonadal but also in several peripheral estrogen target tissues.

Localization of the catalytic subunit Cγ of the cAMP-dependent protein kinase gene (PRKACG) to human chromosome region 9q13

A cDNA for a new catalytic subunit (C gamma) of the cAMP-dependent protein kinase (PKA) was recently isolated from a human testis cDNA library. This subunit was shown to be expressed only in testis, and has so far not been demonstrated in other species. In the present study, we have determined the chromosomal localization of this gene employing a cDNA for C gamma as a probe. Southern blot analysis of genomic DNA from human x mouse somatic cell hybrids allowed us to assign this gene (PRKACG) to human chromosome 9. In situ hybridization to metaphase chromosomes confirmed the somatic cell hybrid data and regionally mapped the C gamma gene of PKA to human chromosome 9q13.

Localization of the human gene for the type I cyclic GMP-dependent protein kinase to chromosome 10.

We have recently characterized cDNAs and genomic DNA fragments for human type I cGMP-dependent protein kinase (cGK). By probing human x hamster hybrid cell lines with a 1.2-kb intron fragment from the human type I cGK gene, we identified a 5.9-kb BglII restriction fragment and localized it to human chromosome 10. In situ hybridization analyses using 3H-labeled cDNA and genomic DNA probes for the human type I cGK to human metaphase chromosomes supported the somatic cell hybrid data and indicated that the gene (PRKG1B; protein kinase, cGMP-dependent) maps to 10p11.2----q11.2.

Assignment of the gene encoding the catalytic subunit Cβ of CAMP-dependent protein kinase to the p36 band on chromosome

SummaryA cDNA for the human catalytic subunit (Cβ) of cAMP-dependent protein kinase (PKA) has been cloned from a testis cDNA library. In the present study, we have determined the chromosomal localization of this gene using a cDNA for Cβ as a probe. Southern blot analysis of genomic DNA from human/mouse cell hybrids revealed that the presence or absence of a 20-kbXbaI fragment, which hybridized with the Cβ probe, was concordant with the presence of human chromosome 1.In situ hybridization to metaphase chromosome confirmed the somatic cell hybrid data and regionally mapped the Cβ gene of PKA to the p36 band on chromosome 1.

A case of tyrosinaemia type I with normal level of succinylacetone in the amniotic fluid.

Prenatal diagnosis of tyrosinaemia type I can be achieved in cultured amniotic cells and in chorionic villus material by testing the activity of fumarylacetoacetate hydrolase and by DNA analysis, and in amniotic fluid by succinylacetone measurement. This specific metabolite can be measured either by gas chromatography–mass spectrometry or by |gd‐aminolevulinate dehydratase inhibition assay. In a series of 65 at‐risk cases tested with the enzyme inhibition assay, one case out of the 18 with the disease had a normal level of succinylacetone. This case is presented.

DNAse I digestion of fixed chromosomes specifically reveals NORs in man

Abstract Chromosomal preparations were digested with different concentrations of DNAse I for various periods of time and then stained with Giemsa. It was found that this endonuclease rapidly modifies the structure of the chromosomes which then lose most of their stainability. However, small chromosomal segments corresponding to the nucleolar organizers (NORs) in man have been consistently observed. Although DNAse I treatment has permitted us to observe that some NORs are stained less intensely than others, comparisons with silver nitrate-stained NORs have shown a strict correspondence, in metaphase as well as in interphase. Chromosomal proteins seem responsible for this staining.