Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.7.49 (reverse transcriptase)
 31,746 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Androgens have significant beneficial effects on the skeleton. However, studies on the effects of androgens on osteoblasts are limited due to the absence of appropriate model systems that combine completeness of the osteoblastic phenotype, rapid proliferation rate, and stable expression of the androgen receptor (AR). Thus, we stably transfected the conditionally immortalized human fetal osteoblastic cell line (hFOB) with the human wild-type AR (hAR) cDNA. Compared to nontransfected hFOB cells, constitutive hAR mRNA expression in three independent hAR-transfected hFOB clones (hFOB/AR) was 15-fold higher in hFOB/AR-16, 62-fold higher in hFOB/AR-2, and 72-fold higher in hFOB/AR-6 cells, respectively, as assessed by semiquantitative reverse transcriptase-polymerase chain reaction (RT-PCR). Detectable constitutive levels of hAR mRNA by Northern blot analysis were present in hFOB/AR-2 and hFOB/AR-6 cells, but not in hFOB/AR-16 or hFOB cells, respectively. Treatment with 5 alpha-dihydrotestosterone (5 alpha-DHT) (10(-8) M) for 24 h did not alter hAR mRNA steady state levels in the hFOB/AR cell lines. Nuclear binding studies demonstrated 152 +/- 73 (mean +/- SEM) functional hARs/nucleus in non-transfected hFOB cells, 3,940 +/- 395 functional hARs/nucleus in hFOB/AR-2 cells, and 3,987 +/- 823 hARs/nucleus in hFOB/AR-6 cells, respectively. Treatment with 5 alpha-DHT increased the expression of a transiently transfected androgen response element-chloramphenicol acetyltransferase (ARE-CAT) reporter construct in hFOB/AR-6 cells in a dose- and time-dependent manner; no such effect was observed in transiently transfected hFOB cells lacking exogenously transfected hARs. Moreover, 5 alpha-DHT-induced ARE-CAT expression was inhibited by the selective androgen receptor antagonist, hydroxyflutamide. In summary, we have developed and characterized androgen-responsive osteoblastic cell lines derived from normal human fetal bone that express physiological levels of functional hARs. These cell lines should provide a suitable model for further studies on the effects of androgens on osteoblast function, including the identification of potential androgen-regulated growth factors and cytokines.
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PMID:Development and characterization of a conditionally immortalized human osteoblastic cell line stably transfected with the human androgen receptor gene. 928 32

The kinetic parameters, steroid substrate specificity and identities of reaction products were determined for four homogeneous recombinant human 3alpha-hydroxysteroid dehydrogenase (3alpha-HSD) isoforms of the aldo-keto reductase (AKR) superfamily. The enzymes correspond to type 1 3alpha-HSD (AKR1C4), type 2 3alpha(17beta)-HSD (AKR1C3), type 3 3alpha-HSD (AKR1C2) and 20alpha(3alpha)-HSD (AKR1C1), and share at least 84% amino acid sequence identity. All enzymes acted as NAD(P)(H)-dependent 3-, 17- and 20-ketosteroid reductases and as 3alpha-, 17beta- and 20alpha-hydroxysteroid oxidases. The functional plasticity of these isoforms highlights their ability to modulate the levels of active androgens, oestrogens and progestins. Salient features were that AKR1C4 was the most catalytically efficient, with k(cat)/K(m) values for substrates that exceeded those obtained with other isoforms by 10-30-fold. In the reduction direction, all isoforms inactivated 5alpha-dihydrotestosterone (17beta-hydroxy-5alpha-androstan-3-one; 5alpha-DHT) to yield 5alpha-androstane-3alpha,17beta-diol (3alpha-androstanediol). However, only AKR1C3 reduced Delta(4)-androstene-3,17-dione to produce significant amounts of testosterone. All isoforms reduced oestrone to 17beta-oestradiol, and progesterone to 20alpha-hydroxy-pregn-4-ene-3,20-dione (20alpha-hydroxyprogesterone). In the oxidation direction, only AKR1C2 converted 3alpha-androstanediol to the active hormone 5alpha-DHT. AKR1C3 and AKR1C4 oxidized testosterone to Delta(4)-androstene-3,17-dione. All isoforms oxidized 17beta-oestradiol to oestrone, and 20alpha-hydroxyprogesterone to progesterone. Discrete tissue distribution of these AKR1C enzymes was observed using isoform-specific reverse transcriptase-PCR. AKR1C4 was virtually liver-specific and its high k(cat)/K(m) allows this enzyme to form 5alpha/5beta-tetrahydrosteroids robustly. AKR1C3 was most prominent in the prostate and mammary glands. The ability of AKR1C3 to interconvert testosterone with Delta(4)-androstene-3,17-dione, but to inactivate 5alpha-DHT, is consistent with this enzyme eliminating active androgens from the prostate. In the mammary gland, AKR1C3 will convert Delta(4)-androstene-3,17-dione to testosterone (a substrate aromatizable to 17beta-oestradiol), oestrone to 17beta-oestradiol, and progesterone to 20alpha-hydroxyprogesterone, and this concerted reductive activity may yield a pro-oesterogenic state. AKR1C3 is also the dominant form in the uterus and is responsible for the synthesis of 3alpha-androstanediol which has been implicated as a parturition hormone. The major isoforms in the brain, capable of synthesizing anxiolytic steroids, are AKR1C1 and AKR1C2. These studies are in stark contrast with those in rat where only a single AKR with positional- and stereo-specificity for 3alpha-hydroxysteroids exists.
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PMID:Human 3alpha-hydroxysteroid dehydrogenase isoforms (AKR1C1-AKR1C4) of the aldo-keto reductase superfamily: functional plasticity and tissue distribution reveals roles in the inactivation and formation of male and female sex hormones. 1099 48

Androgen independence is the major cause of endocrine therapy failure in advanced prostate cancer (PC). To examine the effects of human androgen receptor (AR) expression on growth of human PC cells, transfection of full-length AR cDNA in an androgen-insensitive human prostatic adenocarcinoma cell line (DU145) was performed. Transcriptional activity of AR was confirmed by the MMTV luciferase assay and AR expression was assessed by reverse transcriptase polymerase chain reaction, Western blotting, and immunocytochemistry. Two stable transfectant cell lines expressing functional AR were established and passaged over 60 times. Under standard culture conditions, AR expression in transfected cells was predominantly cytoplasmic. Exposure to dihydrotestosterone (DHT; 60 pM-10 nM) resulted in a rapid (maximal at 30 minutes) translocation of AR to the nucleus. Treatment with DHT (5 nM) caused a significant reduction in cell-cell adhesion and aggregation accompanied by a decrease in E-cadherin expression. This was associated with up to 40% inhibition of proliferation and approximately two-fold increase in apoptosis. These results suggest that gene transfer-mediated AR expression in DU145 cells confers sensitivity to DHT, modulates cell-cell adhesion through E-cadherin, and suppresses cell growth by inhibiting proliferation and promoting apoptosis. This provides amodelfor studies ofAR-regulated cell signalling and identification of novel androgen-regulated genes in PC.
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PMID:Ligand activation of the androgen receptor downregulates E-cadherin-mediated cell adhesion and promotes apoptosis of prostatic cancer cells. 1451 6

To investigate the dynamics of circRNA expression in pig testes, we designed specific strategies to individually study circRNA production from intron lariats and circRNAs originating from back-splicing of two exons. By applying these methods on seven Total-RNA-seq datasets sampled during the testicular puberty, we detected 126 introns in 114 genes able to produce circRNAs and 5,236 exonic circRNAs produced by 2,516 genes. Comparing our RNA-seq datasets to datasets from the literature (embryonic cortex and postnatal muscle stages) revealed highly abundant intronic and exonic circRNAs in one sample each in pubertal testis and embryonic cortex, respectively. This abundance was due to higher production of circRNA by the same genes in comparison to other testis samples, rather than to the recruitment of new genes. No global relationship between circRNA and mRNA production was found. We propose ExoCirc-9244 (SMARCA5) as a marker of a particular stage in testis, which is characterized by a very low plasma estradiol level and a high abundance of circRNA in testis. We hypothesize that the abundance of testicular circRNA is associated with an abrupt switch of the cellular process to overcome a particular challenge that may have arisen in the early stages of steroid production. We also hypothesize that, in certain circumstances, isoforms and circular transcripts from different genes share functions and that a global regulation of circRNA production is established. Our data indicate that this massive production of circRNAs is much more related to the structure of the genes generating circRNAs than to their function. Abbreviations: PE: Paired Ends; CR: chimeric Read; SR: Split Read; circRNA: circular RNA; NC: non conventional; ExoCirc-RNA: exonic circular RNA; IntroLCirc-: name of a porcine intronic lariat circRNA; ExoCirc-: name of a porcine exonic circRNA; IntronCircle-: name of a porcine intron circle; sisRNA: stable intronic sequence RNA; P: porcine breed Pietrain; LW: porcine breed Large White; RT: reverse transcription/reverse transcriptase; Total-RNA-seq: RNA-seq obtained from total RNA after ribosomal depletion; mRNA-seq: RNA-seq of poly(A) transcripts; TPM: transcripts per million; CR-PM: chimeric reads per million; RBP: RNA binding protein; miRNA: micro RNA; E2: estradiol; DHT: dihydrotestesterone.
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PMID:Analysis of pig transcriptomes suggests a global regulation mechanism enabling temporary bursts of circular RNAs. 3112 Mar 23