Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.3.1.28 (chloramphenicol acetyltransferase)
 5,100 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

In the present report we describe a heretofore unrecognized role for a Jak/STAT signaling pathway, namely the stimulation of expression of the aromatase P450 (CYP19) gene, and hence of estrogen biosynthesis, in human adipose tissue. Expression of this gene in adipose tissue as well as in adipose stromal cells maintained in the presence of serum and glucocorticoids is regulated by a distal TATA-less promoter, I.4, which contains a glucocorticoid response element, an Sp1 binding site, and an interferon-gamma activation site (GAS) element. The stimulatory action of serum (in the presence of dexamethasone) can be replaced by interleukin (IL)-11, leukemia inhibitory factor, and oncostatin-M, as well as by IL-6, providing the IL-6 soluble receptor is also present. Stimulation of the cells by these factors led to rapid phosphorylation of Jak1, but not Jak2 or Jak3, on tyrosine residues. STAT3 but not STAT1 was also phosphorylated and bound to the GAS element in the I.4 promoter region. When regions of this promoter were fused upstream of the chloramphenicol acetyltransferase reporter gene and transfected into the cells, mutagenesis or deletion of the GAS element led to complete loss of reporter gene expression. Since adipose tissue is the major site of estrogen biosynthesis in men and in postmenopausal women, this pathway involving a Jak/STAT signaling mechanism acting together with glucocorticoids and Sp1 appears to be the principal means whereby estrogen biosynthesis is regulated in the elderly.
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PMID:Aromatase P450 gene expression in human adipose tissue. Role of a Jak/STAT pathway in regulation of the adipose-specific promoter. 760 17

The biosynthesis of estrogens is catalyzed by aromatase P450 (P450arom), the product of the CYP19 gene. The tissue-specific expression of the CYP19 gene is regulated by means of tissue-specific promoters through the use of alternative splicing mechanisms. Thus, transcripts containing various 5'-untranslated termini are present in human placenta and other fetal tissues, ovary, brain, and adipose stromal cells. Sequence corresponding to untranslated exon 1.4 is present in 5'-termini of transcripts expressed in adipose tissue and fetal liver, as well as adipose stromal cells in primary culture in the presence of dexamethasone and fetal calf serum (FCS). Identification of hormone-responsive, tissue-specific promoter regions, as well as growth factor-response elements upstream of exon 1.4, may provide insight into the regulation of estrogen biosynthesis in adipose tissue, which is implicated in the development of breast and endometrial cancer. The goals of the present study were to define the 1.4 promoter region with respect to the start of transcription and to characterize the region(s) responsible for conferring glucocorticoid responsiveness on aromatase expression. The transcription initiation site was identified by means of primer extension and S1 nuclease protection analyses. No TATA-like sequence was evident upstream of this site. Various deletion mutations of the upstream flanking region of exon 1.4 and including part of exon 1.4 were made using polymerase chain reaction or restriction enzyme digestion. The genomic fragments were fused upstream of the chloramphenicol acetyltransferase (CAT) reporter gene. These constructs were transfected into adipose stromal cells and fetal hepatocytes in primary culture in medium containing FCS with or without dexamethasone. The -560/+10 base pair (bp) construct expressed CAT activity after a putative silencer element was deleted, and expression was induced by dexamethasone about 3-fold. Transfection of the -330/+170 bp construct, which contains an upstream glucocorticoid response element (GRE) as well as an Sp1-like sequence in untranslated exon 1.4, resulted in an 8-fold stimulation of expression of CAT activity by dexamethasone. The upstream GRE as well as the Sp1-like sequence in untranslated exon 1.4 were mutated separately, and together, to further confirm whether the GRE or Sp1 binding site play a role in the regulation of promoter 1.4-driven transcription. Mutation of either the GRE or Sp1 binding site, or both, in the -330/+170 bp construct, resulted in loss of dexamethasone-induced CAT reporter gene expression.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Characterization of the sequences of the human CYP19 (aromatase) gene that mediate regulation by glucocorticoids in adipose stromal cells and fetal hepatocytes. 777 80

CYP19, the human aromatase cytochrome P-450 (P450arom) gene, encodes an enzyme which converts androgens to estrogens by three successive hydroxylation reactions by coupling with NADPH-cytochrome P-450 reductase. In the present study, we have characterized two cis-acting transcriptional regulatory elements of CYP19, termed as hATRE-1 (human aromatase cytochrome P-450 gene transcriptional regulatory element-1) ([sequence: see text]) and hATRE-2 ([sequence: see text]). These sequences are located between -2238 and -2214, and between -2141 and -2098 relative to the major cap site of the gene, respectively. Transient expression analysis in human BeWo choriocarcinoma cells, in which CYP19 is expressed, shows that hATRE-1 represses the expression of the bacterial chloramphenicol acetyltransferase reporter gene driven by the promoter of CYP19, whereas hATRE-2 enhances the reporter gene expression in response to 12-O-tetradecanoylphorbol 13-acetate. Electrophoretic mobility shift analysis indicates that nuclear binding factors specific to hATRE-1 are present in BeWo cells, but not in HeLa cells nor in TYK-nu cells that lack the expression of CYP19. In contrast, nuclear binding factors to hATRE-2 are present not only in BeWo cells but also in the latter two types of cells. Nevertheless, hATRE-2 does not affect the reporter gene expression in HeLa cells and TYK-nu cells. These results indicate that hATRE-1 and hATRE-2 are cis-acting transcriptional regulatory elements involving in the regulation of the cell type-specific expression of CYP19.
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PMID:Identification and characterization of cis-acting regulatory elements for the expression of the human aromatase cytochrome P-450 gene. 813 35

Aromatase cytochrome P450, a member of the cytochrome P450 gene super family, catalyzes conversion of androgens to estrogens in a form of an enzyme-complex with NADPH-cytochrome P450 reductase. Transcription of the aromatase cytochrome P450 gene (CYP19) is regulated in part by tissue-specific promoters coupled with alternative splicing mechanisms. The transcription in human placenta is governed by a promoter activity of the 5' flanking region of exon I.1, which is mapped more than 40 kb upstream from the translational start codon observed in exon II. Transient expression analyses with chimeric constructs containing the 5' flanking sequences linked to the bacterial chloramphenicol acetyltransferase (CAT) gene in human BeWo choriocarcinoma cells localized a cell-type specific enhancer element between -242 and -166 relative to the major cap site. DNase I footprinting and transient expression analyses of the enhancer element indicate that it consists of two sub-elements and that both sub-elements are necessary for the maximum enhancement of the transcription. In addition to the enhancer element, a cis-acting element important for transcriptional enhancement of the gene in response to 12-O-tetradecanoylphorbol 13-acetate in BeWo cells is localized between -2141 and -2115. A nuclear factor binding to the element is identified as NF-IL6 (also termed as LAP and C/EBP beta). Transient expression analyses using the CAT constructs containing the NF-IL6 binding sites involvement of the factor in transcriptional regulation of CYP19.
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PMID:Identification and characterization of transcriptional regulatory elements of the human aromatase cytochrome P450 gene (CYP19). 860 36

The proximal promoter of the rat aromatase CYP19 gene contains two functional regions that, by 5'-deletion analyses, have been shown to confer hormone/ cAMP inducibility to chimeric genes in primary cultures of rat granulosa cells and constitutive expression in R2C Leydig cells. Promoter region A binds Steroidogenic Factor-1 (SF-1); region B binds cAMP-regulatory element binding protein (CREB) and two other factors (designated X and Y). Mutations were generated within the context of the intact promoter to selectively eliminate the binding of either SF-1, CREB, CREB plus factors X and Y, or all of the above. When expression vectors that failed to bind either CREB alone or CREB plus factors X and Y were transfected into granulosa cells, cAMP-dependent chloramphenicol acetyltransferase (CAT) activity was reduced 65% indicating that CREB alone, and not factors X and Y, mediates the cAMP response of this cAMP response element-like domain. Similarly, cAMP-dependent CAT activity was reduced 50% in constructs that failed to bind SF-1 and was abolished with vectors that were unable to bind either factor. In R2C Leydig cells, the absence of either CREB or SF-1 binding resulted in an almost complete loss in CAT activity. Both immunoreactive CREB and phosphorylated CREB (phospho-CREB) were present in extracts and nuclei of R2C cells. Immunoreactive phosphoCREB was low in granulosa cell extracts and nuclei but increased rapidly (90 min) in response to FSH/cAMP and was highest at 48 h, at a time when SF-1 was also phosphorylated and expression of the endogenous gene was elevated. Although the amount of CREB and SF-1 remained unchanged in response to FSH, LH mediated a rapid decrease in the amount of SF-1 (but not CREB) that is coincident with decreased aromatase mRNA in luteinizing granulosa cells. Taken together, the data indicate that expression of the aromatase gene is dependent on the additive interactions of regions A and B of the aromatase promoter in granulosa cells and the synergistic interactions of these same regions in R2C cells and that these interactions are dependent, in turn, on the phosphorylation of CREB and SF-1 and the content of these factors, as well as the presence of putative coregulatory molecules.
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PMID:Functional interactions, phosphorylation, and levels of 3',5'-cyclic adenosine monophosphate-regulatory element binding protein and steroidogenic factor-1 mediate hormone-regulated and constitutive expression of aromatase in gonadal cells. 905 76

Aromatase cytochrome P450 catalyses the reaction to convert androgens to estrogens by coupling with NADPH-cytochrome P450 reductase in the endoplasmic reticulum. The human aromatase cytochrome P450 gene (CYP19) is expressed in a variety of tissues under regulation of tissue-specific promoters. Previously, we localized a cell-type specific transcriptional enhancer element between -242 and -166 relative to the major cap site of the gene, by transient expression analysis in human BeWo choriocarcinoma cells. In the present study, we demonstrate that the enhancer element consists of two subelements, element I (located between -238 and -200), and element II (located between -196 and -176) as analysed by DNase I footprinting using the nuclear extracts of BeWo cells. The gel mobility shift assay shows that each of these subelements binds specific nuclear factor(s). The transient expression of the bacterial chloramphenicol acetyltransferase gene constructs involving the subelements in BeWo cells reveals that the elements activate reporter gene expression synergistically when present together, nevertheless each of the elements by itself also has an enhancer activity. The transient expression analysis further shows that element I is responsible for the transcriptional synergism with the binding site of a nuclear factor-interleukin-6 (NF-IL-6) (also known as CCAAT enhancer/binding protein beta), which is located between -2141 and -2115 relative to the major cap site of the gene. These results suggest that the enhancer element plays important roles in sustaining the high levels of CYP19 expression in placental cells in cooperation with other cis-acting transcritional regulatory elements.
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PMID:Cooperative regulation of the human aromatase cytochrome P450 gene transcription by placenta-specific cis-acting elements. 936 92