scholarly journals Expression Cloning of a Novel Estrogenic Mouse 17β-Hydroxysteroid Dehydrogenase/ 17-Ketosteroid Reductase (m17HSD7), Previously Described as a Prolactin Receptor-Associated Protein (PRAP) in Rat

1998 ◽  
Vol 12 (7) ◽  
pp. 1048-1059 ◽  
Author(s):  
Pasi Nokelainen ◽  
Hellevi Peltoketo ◽  
Reijo Vihko ◽  
Pirkko Vihko
Keyword(s):  
Mammary Gland ◽  
Corpus Luteum ◽  
Prolactin Receptor ◽  
Hydroxysteroid Dehydrogenase ◽  
Mouse Mammary Gland ◽  
Expression Cloning ◽  
Calculated Molecular Mass ◽  
Hek 293 ◽  
Receptor Associated Protein ◽  
Enzymatic Characteristics

Abstract 17β-Hydroxysteroid dehydrogenases/17-ketosteroid reductases (17HSDs) modulate the biological activity of certain estrogens and androgens by catalyzing reductase or dehydrogenase reactions between 17-keto- and 17β-hydroxysteroids. In the present study, we demonstrate expression cloning of a novel type of 17HSD, chronologically named 17HSD type 7, from the HC11 cell line derived from mouse mammary gland. The cloned cDNA, 1.7 kb in size, encodes a protein of 334 amino acids with a calculated molecular mass of 37,317 Da. The primary structure contains segments characteristic of enzymes belonging to the short-chain dehydrogenase/reductase superfamily. Strikingly, mouse 17HSD type 7 (m17HSD7) shows 89% identity with a recently cloned rat protein called PRL receptor-associated protein (PRAP). The function of PRAP has not yet been demonstrated. The enzymatic characteristics of m17HSD7 and RT-PCR-cloned rat PRAP (rPRAP) were analyzed in cultured HEK-293 cells, where both of the enzymes efficiently catalyzed conversion of estrone (E1) to estradiol (E2). With other substrates tested no detectable 17HSD or 20α-hydroxysteroid dehydrogenase activities were found. Kinetic parameters for m17HSD7 further indicate that E1 is a preferred substrate for this enzyme. Relative catalytic efficiencies (Vmax/Km values) for E1 and E2 are 244 and 48, respectively. As it is the case with rPRAP, m17HSD7 is most abundantly expressed in the ovaries of pregnant animals. Further studies show that the rat enzyme is primarily expressed in the middle and second half of pregnancy, in parallel with E2 secretion from the corpus luteum. The mRNA for m17HSD7 is also apparent in the placenta, and a slight signal for m17HSD7 is found in the ovaries of adult nonpregnant mice, in the mammary gland, liver, kidney, and testis. Altogether, because of their similar primary structures, enzymatic characteristics, and the tissue distribution of m17HSD7 and rPRAP, we suggest that rPRAP is rat 17HSD type 7. Furthermore, the results indicate that 17HSD7 is an enzyme of E2 biosynthesis, which is predominantly expressed in the corpus luteum of the pregnant animal.

scholarly journals Reproductive Cycle Regulation of Nuclear Import, Euchromatic Localization, and Association with Components of Pol II Mediator of a Mammalian Double-Bromodomain Protein

2002 ◽  
Vol 16 (8) ◽  
pp. 1727-1737 ◽  
Author(s):  
Thomas E. Crowley ◽  
Emily M. Kaine ◽  
Manabu Yoshida ◽  
Anindita Nandi ◽  
Debra J. Wolgemuth
Keyword(s):  
Mammary Gland ◽  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Reproductive Cycle ◽  
Large Subunit ◽  
Mouse Mammary Gland ◽  
Receptor Associated Protein ◽  
E2f Transcription Factor ◽  
Mammary Epithelia ◽  
Gland Function

Abstract Fsrg1 (female sterile homeotic-related gene 1) is the mouse homolog of the human RING3 protein, which has been shown to associate with the E2 promoter binding factor (E2F) transcription factor and to have a possible role in cell cycle-linked transcriptional regulation. The Fsrg1 protein is 60% identical in sequence to the RNA polymerase II mediator subunit Fsrg4, another member of this subfamily of double bromodomain-containing proteins that are homologs of Drosophila female sterile homeotic. Antibodies against murine Fsrg1 were generated and used in immunoblot and immunoprecipitation experiments to identify proteins interacting with Fsrg1 and RING3. In the presence of acetylated but not nonacetylated histone H3 and H4 peptides, RING3 was shown to interact with E2F, mediator components cyclin-dependent kinase 8 and thyroid receptor-associated protein 220, and the RNA polymerase II large subunit. Fsrg1 mRNA had been previously shown to be expressed at high levels in the epithelium of the adult mouse mammary gland. To determine the physiological relevance of these potential associations, we examined the patterns of expression of Fsrg1 mRNA and protein in the adult mammary epithelia during the reproductive cycle as the tissue is responding to estrogen, progesterone, and prolactin. Changes in the nuclear vs. cytoplasmic localization of Fsrg1 were observed and correlated with physiological changes in mammary gland function. The observations suggested that Fsrg1 may be involved in the transcriptional activities of genes involved in proliferation of the mammary epithelia during pregnancy and in orchestrating postlactation involution and apoptosis. Localization of Fsrg1 on euchromatin, the transcribed portion of the chromosomes, is consistent with its hypothesized function as a transcription regulator.

scholarly journals Cloning and Characterization of a 5′ Regulatory Region of the Prolactin Receptor-Associated Protein/17β Hydroxysteroid Dehydrogenase 7 Gene

Endocrinology ◽  
2005 ◽  
Vol 146 (6) ◽  
pp. 2807-2816 ◽  
Author(s):  
Michael Risk ◽  
Aurora Shehu ◽  
Jifang Mao ◽  
Carlos O. Stocco ◽  
Laura T. Goldsmith ◽  
...  
Keyword(s):  
Cell Line ◽  
Chorionic Gonadotropin ◽  
Human Chorionic Gonadotropin ◽  
Prolactin Receptor ◽  
Hydroxysteroid Dehydrogenase ◽  
Proximal Promoter ◽  
Luteal Cell ◽  
Nuclear Factor ◽  
Receptor Associated Protein ◽  
Nuclear Factor Y

Abstract Prolactin receptor-associated protein (PRAP) originally cloned in our laboratory was shown to be a novel, luteal isoform of 17β hydroxysteroid dehydrogenase 7 (17βHSD7). In this study, we cloned the promoter region of rat PRAP/17βHSD7 and investigated the mechanisms regulating both basal activity and LH-induced repression of this promoter. Truncated and site-specific mutants of PRAP/17βHSD7 promoter identified two enhancer regions that contained highly conserved Sp1 binding site and bound Sp1 from nuclear extracts of both corpora lutea and a rat luteal cell line. Repression of PRAP/17βHSD7 expression and promoter activity by human chorionic gonadotropin/forskolin was localized to a −52-bp proximal segment of the promoter. This region contained a conserved CCAAT site and bound nuclear factor Y; binding of this transcription factor was inhibited by human chorionic gonadotropin in vivo. Furthermore, mutation of the nuclear factor Y site in the −52-bp promoter-reporter construct abolished forskolin-mediated inhibition of the promoter in a rat luteal cell line. In summary, we have identified the promoter elements involved in the basal expression of PRAP/17βHSD7. We have also found that LH-mediated repression of this gene is at the level of transcription and involves inhibition of nuclear factor YA binding to the CCAAT site within the proximal promoter.

Progesterone and EGF inhibit mouse mammary gland prolactin receptor and beta-casein gene expression

1994 ◽  
Vol 267 (5) ◽  
pp. C1467-C1472 ◽  
Author(s):  
S. Nishikawa ◽  
R. C. Moore ◽  
N. Nonomura ◽  
T. Oka
Keyword(s):  
Gene Expression ◽  
Mammary Gland ◽  
Prolactin Receptor ◽  
Mammary Epithelium ◽  
Mouse Mammary Gland ◽  
Mrna Levels ◽  
Casein Gene ◽  
Beta Casein

Regulation of mouse mammary gland long-form prolactin receptor (PRL-RL) mRNA levels by progesterone and epidermal growth factor (EGF) and the relationship between PRL-RL and beta-casein gene expression were examined in vivo and in vitro. PRL-RL and beta-casein mRNA levels increased approximately 6- and 15-fold from the pregnant to the lactating period, respectively, when normalized to the level of beta-actin mRNA. Ovariectomy of pregnant mice rapidly reduced the serum concentration of progesterone and increased the level of PRL-RL and beta-casein mRNAs approximately three- and fourfold compared with sham-operated animals 24 h after the operation. Injection of progesterone, but not estrogen, inhibited the increase in both mRNA levels. PRL-RL and beta-casein mRNA levels in cultured mammary epithelium increased in response to insulin, hydrocortisone, and prolactin, whereas progesterone or EGF caused inhibition. The combination of EGF and progesterone produced a greater inhibition than either hormone alone. These results indicate that both progesterone and EGF serve as negative regulators of lactogenesis.

scholarly journals Effects of Weaning and Suckling on γ-Casein and Prolactin Receptor mRNA Levels in the Mouse Mammary Gland during Lactation

1998 ◽  
Vol 69 (8) ◽  
pp. 728-733
Author(s):  
Jae-Young KIM ◽  
Yasushi MIZOGUCHI ◽  
Takeshi KURAISHI ◽  
Hirohito YAMAGUCHI ◽  
Jumpei ENAMI ◽  
...  
Keyword(s):  
Mammary Gland ◽  
Prolactin Receptor ◽  
Mouse Mammary Gland ◽  
Mrna Levels ◽  
Receptor Mrna

scholarly journals The Expression and Activity of Rhodanese, 3-Mercaptopyruvate Sulfurtransferase, Cystathionine γ-Lyase in the Most Frequently Chosen Cellular Research Models

Biomolecules ◽  
2021 ◽  
Vol 11 (12) ◽  
pp. 1859
Author(s):  
Marta Kaczor-Kamińska ◽  
Kamil Kaminski ◽  
Maria Wróbel
Keyword(s):  
Mammary Gland ◽  
Cell Lines ◽  
Colorectal Adenocarcinoma ◽  
Mouse Mammary Gland ◽  
Mouse Fibroblast ◽  
Screening Tests ◽  
Limited Data ◽  
Hek 293 ◽  
Mercaptopyruvate Sulfurtransferase ◽  
Sulfur Level

This paper provides information concerning the activity and expression levels of three sulfurtransferases (STRs): rhodanese (TST, EC: 2.8.1.1), 3-mercaptopyruvate sulfurtransferase (MPST, EC: 2.8.1.2) and cystathionine γ-lyase (CTH, EC: 4.4.1.1) in various cell lines. Since very limited data are available in the scientific literature on this subject, the available data are included in this paper. These shortages often force the researchers to carry out their own screening tests that allow them to choose an appropriate model for their further studies. This work supplements the existing deficiencies in this area and presents the activity and expression of STRs in the eight most frequently chosen cell lines: the mouse mammary gland cell line (NMuNG, ATCC: CRL-1636), mouse mammary gland tumor (4T1, ATCC: CRL-2539), mouse fibroblast (MEF, ATCC: SCRC-1008), mouse melanoma (B16-F1, ATCC: CRL-6323), human colorectal adenocarcinoma (Caco-2, ATCC: HTB-37), human embryonic kidney (HEK-293, ATCC: CRL-1573), human osteosarcoma (MG-63, ATCC: CRL-1427) and rat myocardium (H9c2, ATCC: CRL-1446). Changes in STRs activity are directly related to the bioavailability of cysteine and the sulfane sulfur level, and thus the present authors also measured these parameters, as well as the level of glutathione (its reduced (GSH) and oxidized (GSSG) form) and the [GSH]/[GSSG] ratio that determines the antioxidant capacity of the cells. STRs demonstrate diverse functionality and clinical relevance; therefore, we also performed an analysis of genetic variation of STRs genes that revealed a large number of polymorphisms. Although STRs still provide challenges in several fields, responding to them could not only improve the understanding of various diseases, but may also provide a way to treat them.

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