Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

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)
Mol Endocrinol 1995 Mar
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

The enzyme 5-enolpyruvylshikimate-3-phosphate (EPSP) synthase (EC 2.5.1.19), the target of the herbicide glyphosate [N-(phosphonomethyl)glycine], exists in two molecular forms in Euglena gracilis. One form has previously been characterized as a monofunctional 59 kDa protein. The other form constitutes a single domain of the multifunctional 165 kDa arom protein. The two enzyme forms are inversely regulated at the protein and mRNA levels during light-induced chloroplast development, as demonstrated by the determination of their enzyme activities after non-denaturing polyacrylamide gel electrophoresis and Northern hybridization analysis with a Saccharomyces cerevisiae ARO1 gene probe. The arom protein and its mRNA predominate in dark-grown cells, and the levels of both decline upon illumination. In contrast, the monofunctional EPSP synthase and its mRNA are induced by light, the increase in mRNA abundance preceding accumulation of the protein. The two enzymes are localized in different subcellular compartments, as demonstrated by comparing total protein patterns with those of isolated organelles. Glyphosate-adapted wild-type cells and glyphosate-tolerant cells of a plastid-free mutant of E. gracilis, W10BSmL, were used for organelle isolation and protein extraction, as these cell lines overproduce EPSP synthase and the arom protein, respectively. Evidence was obtained for the cytosolic localization of the arom protein and the plastid compartmentalization of the monofunctional EPSP synthase. These conclusions are further supported by the observation that EPSP synthase precursor, produced by in vitro translation of the hybrid-selected mRNA, was efficiently taken up and processed to mature size by isolated chloroplasts from photoautotrophic wild-type E. gracilis cells, while the in vitro-synthesized arom protein was not sequestered by isolated Euglena plastids.
Mol Gen Genet 1994 Dec 01
PMID:Cytosolic and plastid forms of 5-enolpyruvylshikimate-3-phosphate synthase in Euglena gracilis are differentially expressed during light-induced chloroplast development. 780 12

Aromatase activity may be detected in most, but not all, breast cancers, and in certain tumours there appears to be decreased sensitivity to the aromatase inhibitor 4-hydroxyandrostenedione (4-OHA). The aims of the present study were to measure aromatase activity, and its sensitivity to 4-OHA, in breast tumours, and to examine the CYP19 gene encoding the aromatase cytochrome P450 (P450arom) for the presence of mutations. In vitro aromatase activity and sensitivity to 4-OHA were measured by determining the conversion of tritiated testosterone to tritiated oestradiol in breast tumour tissue in the absence and presence of 4-OHA (10 nM). Genomic DNA was extracted from five tumours: one showing no detectable aromatase activity and four displaying evidence of aromatase activity (two sensitive and two insensitive to 4-OHA). Subsequent PCR-single-strand conformation polymorphism analysis revealed a variation in the mobility of single-stranded DNA for exons III, VII and X, corresponding, as shown by direct sequencing of PCR products, to common polymorphism of the aromatase gene. This study does not provide evidence for mutation in the coding exons of the P450arom gene which would account for either the absence of aromatase activity or its changed sensitivity to 4-OHA in breast cancers.
J Mol Endocrinol 1994 Dec
PMID:Analysis of the aromatase cytochrome P450 gene in human breast cancers. 789 51

Estrogens are synthesized from C19 steroids by a unique form of cytochrome P450, aromatase cytochrome P-450 (P-450AROM; the product of the CYP19 gene). We have shown that tissue-specific expression of human P-450AROM is determined, in part, by the use of alternative promoters. Previous methods of analysis for determining the specific 5'-termini of the different transcripts included S1 nuclease protection, primer extension, and Northern analysis. In the present study we have used the RACE procedure (rapid amplification of cDNA ends) to amplify and clone the 5' termini of P-450AROM transcripts expressed in human corpus luteum (CL). Sequencing of the resulting clones supports the results of the previously performed studies. Specifically, the proximal promoter, PII, is the predominant promoter utilized in CL, such that the start of transcription occurs 26 bp downstream of the putative TATA sequence. A minority of the clones possess an alternative 5'-end, namely I.3. Exon-specific Northern analysis confirms that the majority of the P-450AROM transcripts in CL tissue contain sequence specific for promoter II. Similarly, exon-specific Northern analysis indicates that transcripts in human follicles, as well as granulosa cells in culture, contain primarily sequence specific for promoter II.
Mol Cell Endocrinol 1993 Nov
PMID:Exon-specific northern analysis and rapid amplification of cDNA ends (RACE) reveal that the proximal promoter II (PII) is responsible for aromatase cytochrome P450 (CYP19) expression in human ovary. 814 90

The relationship of function to structure of aromatase cytochrome P450 (P450arom; the product of the CYP19 gene) has been examined by means of sequence alignment and site-directed mutagenesis. Comparison has been made between the sequence of P450arom and the two soluble bacterial cytochrome P450 isoforms, whose three-dimensional structure has been determined (P450BM3 and P450cam). From this comparison, it appears that although there is a similarity of overall structure in cytochromes P450, there is enough significant difference in the regions involved in substrate recognition and substrate binding that residues believed to be involved, even in the known structures, must be tested. With this in mind, we have generated a detailed alignment of P450arom, including the definition of putative alpha-helices and beta-sheets based on comparison of the alignments of P450BM3 and P450cam, generated from their three-dimensional structure, and have made mutations in regions we believe to be involved in substrate recognition at the solvent surface and orientation in the heme pocket. We have mutated F116 and F134 to determine if they are present in the heme pocket, and Q225 and L228 to determine if they are a part of the substrate recognition loop. Although F116E is essentially inactive and may be a folding mutant or may inhibit reductase binding, F134E is more active than the wild type and may be located in the heme pocket facilitating the hydrogen abstraction from C2 of androstenedione. Mutations at Q225 and L228 also result in the anticipated changes in the apparent Km and maximum velocity.(ABSTRACT TRUNCATED AT 250 WORDS)
Mol Endocrinol 1993 Dec
PMID:Functional domains of human aromatase cytochrome P450 characterized by linear alignment and site-directed mutagenesis. 814 67

The formation of estrogens from C19 steroids is catalyzed by a specific form of cytochrome P450, aromatase cytochrome P450 (P450arom; the product of the CYP19 gene). In previous studies we have demonstrated that in adult human tissues and placenta, expression of the CYP19 gene is regulated in part by means of tissue-specific promoters through the use of alternative splicing mechanisms. In addition to placenta, a number of fetal tissues express aromatase, including liver, intestine, skin, and brain. To characterize the CYP19 transcripts present in these and other fetal tissues, we have used reverse transcription and polymerase chain reaction to amplify sequences corresponding to the various untranslated exons from RNA extracted from these tissues. In addition, we have prepared cDNA libraries using RNA from these tissues by the method of rapid amplification of cDNA ends. Sequencing of clones derived from these libraries has been employed to confirm the presence of sequence corresponding to untranslated exons at the 5'-ends of P450arom transcripts. Based on these findings, we conclude that in fetal tissues other than placenta, transcripts containing sequence found in the exon we have previously named I.4 appear to be the most common. Such sequences have been found in cells in which P450arom expression is stimulated by glucocorticoids. Thus, the presence of such transcripts in fetal liver RNA is consistent with our previous observations that aromatase activity in fetal hepatocytes is stimulated by glucocorticoids. Secondly, transcripts are present in the fetal adrenal, although no aromatase activity has ever been detected in this tissue.(ABSTRACT TRUNCATED AT 250 WORDS)
Mol Endocrinol 1994 Feb
PMID:Expression of the gene encoding aromatase cytochrome P450 (CYP19) in fetal tissues. 817 Apr 77

Cells of the plastid-free mutant line of Euglena gracilis var. bacillaris, W10BSmL, can be adapted to glyphosate [N-(phosphonomethyl)glycine] by gradually increasing the concentration of the herbicide in the culture medium. The molecular basis of glyphosate tolerance is the selective ca. ten-fold overproduction of the multifunctional arom protein catalyzing steps 2-6 in the pre-chorismate pathway. Determination of 5-enolpyruvylshikimate-3-phosphate (EPSP) synthase (E.C.2.5.1.19), shikimate:NADP+ oxidoreductase (E.C.1.1.1.25) and shikimate kinase (E.C.2.7.1.71) activities after non-denaturing gel electrophoresis, in combination with two-dimensional separations, revealed an increase in all three enzyme activities associated with overproduction of a 165 kDa protein in cells adapted to 6 mM glyphosate. Further evidence for an involvement of the multifunctional arom protein in aromatic amino acid synthesis in the plastid-free W10BSmL cells was obtained by Northern hybridization with ARO1-, aroA-, aroL- and aroE-specific Saccharomyces cerevisiae gene probes encoding the entire arom protein or parts of the EPSP synthase, shikimate:NADP+ oxidoreductase and shikimate kinase domains, respectively. Overproduction in adapted relative to control cells of a 5.3 kb transcript that cross-hybridized with all of the different probes could be demonstrated. The elevated content of the arom transcript correlated with a selective amplification of two out of five genomic sequences that hybridized with the S. cerevisiae ARO1 gene probe in Southern blots. One of the amplified genomic fragments is assumed to encode the previously identified monofunctional 59 kDa EPSP synthase, which is thought to be an organellar protein, that accumulates to a certain extent in its enzymatically active precursor form of 64.5 kDa in the plastid-free W10BSmL cells.
Mol Gen Genet 1993 Jun
PMID:Overproduction by gene amplification of the multifunctional arom protein confers glyphosate tolerance to a plastid-free mutant of Euglena gracilis. 839 Nov 14

The activational effects of testosterone (T) on male copulatory behavior are mediated by its aromatization into estradiol. In quail, we have shown by stereotaxic implantation of steroids and metabolism inhibitors and by electrolytic lesions that the action of T and its aromatization take place in the sexually dimorphic medial preoptic nucleus (POM). The distribution and regulation of brain aromatase was studied in this species by product-formation assays measuring aromatase activity (AA) in microdissected brain regions and by immunocytochemistry (ICC). Aromatase-immunoreactive (ARO-ir) neurons were found in four brain regions: the POM, the septal region, the bed nucleus of the stria terminals (BNST) and the tuberal hypothalamus. ARO-ir cells actually outline the POM boundaries. ARO-ir material is found not only in the perikarya of neurons but also in the full extension of their cellular processes including the axons and the presynaptic boutons. This is confirmed at the light level by the demonstration of immunoreactive fibers and punctate structures in brain regions that are sometimes fairly distant from the closest ARO-ir cells. A lot of ARO-ir cells in the POM and BNST do not contain immunoreactive estrogen receptors (ER-ir) as demonstrated by double label ICC. These morphological data suggest an unorthodox role for the enzyme or the locally formed estrogens. In parallel with copulatory behavior, the preoptic AA decreases after castration and is restored by T to levels seen in sexually mature males. This probably reflects a change in enzyme concentration rather than a modulation of the activity in a constant number of molecules since the maximum enzymatic velocity (Vmax) only is affected while the affinity (Km) remains unchanged. In addition, T increases the number of ARO-ir neurons in POM and other brain areas suggesting that the concentration of the antigen is actually increased. This probably involves the direct activation of aromatase transcription as demonstrated by RT-PCR studies showing that aromatase mRNA is increased following T treatment of castrates. These activating effects of T seem to result from a synergistic action of androgenic and estrogenic metabolites of the steroid. The anatomical substrate for these regulations remains unclear at present especially in POM where ARO-ir cells do not in general contain ER-ir while androgen receptors appear to be rare based on both [3H] dihydrotestosterone autoradiography and ICC. Transynaptic mechanisms of control may be considered. A modulation of brain aromatase by catecholamines is also suggested by a few pharmacological studies.(ABSTRACT TRUNCATED AT 400 WORDS)
J Steroid Biochem Mol Biol 1993 Mar
PMID:Brain aromatase and the control of male sexual behavior. 847 66

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.
J Steroid Biochem Mol Biol 1996 Jan
PMID:Identification and characterization of transcriptional regulatory elements of the human aromatase cytochrome P450 gene (CYP19). 860 36

The enzyme aromatase converts testosterone (T) into 17 beta-estradiol and plays a pivotal role in the control of reproduction. In particular, the aromatase activity (AA) located in the preoptic area (POA) of male Japanese quail is a limiting step in the activation by T of copulatory behavior. Aromatase-immunoreactive (ARO-ir) cells of the POA are specifically localized within the cytoarchitectonic boundaries of the medial preoptic nucleus(POM), a sexually dimorphic and steroid-sensitive structure that is a necessary and sufficient site of steroid action in the activation of behavior. Stereotaxic implantation of aromatase inhibitors in but not around the POM strongly decreases the behavioral effects of a systemic treatment with T of castrated males. AA is decreased by castration and increased by aromatizable androgens and by estrogens. These changes have been independently documented at three levels of analysis: the enzymatic activity measured by radioenzymatic assays in vitro, the enzyme concentration evaluated semi-quantitatively by immunocytochemistry and the concentration of its messenger RNA quantified by reverse transcription-polymerase chain reaction (RT-PCR). These studies demonstrate that T acting mostly through its estrogenic metabolites regulates brain aromatase by acting essentially at the transcriptional level. Estrogens produced by central aromatization of T therefore have two independent roles: they activate male copulatory behavior and they regulate the synthesis of aromatase. Double label immunocytochemical studies demonstrate that estrogen receptors(ER) are found in all brain areas containing ARO-ir cells but the extent to which these markers are colocalized varies from one brain region to the other. More than 70% of ARO-ir cells contain detectable ER in the tuberal hypothalamus but less than 20% of the cells display this colocalization in the POA. This absence of ER in ARO-ir cells is also observed in the POA of the rat brain. This suggests that locally formed estrogens cannot control the behavior and the aromatase synthesis in an autocrine fashion in the cells where they were formed. Multi-neuronal networks need therefore to be considered. The behavioral activation could result from the action of estrogens in ER-positive cells located in the vicinity of the ARO-ir cells where they were produced (paracrine action). Alternatively, actions that do not involve the nuclear ER could be important. Immunocytochemical studies at the electron microscope level and biochemical assays of AA in purified synaptosomes indicate the presence of aromatase in presynaptic boutons. Estrogens formed at this level could directly affect the pre-and post-synaptic membrane or could directly modulate neurotransmission namely through their metabolization into catecholestrogens (CE) which are known to be powerful inhibitors of the catechol- omicron - methyl transferase (COMT). The inhibition of COMT should increase the catecholaminergic transmission. It is significant to note, in this respect, that high levels of 2-hydroxylase activity, the enzyme that catalyzes the transformation of estrogens in CE, are found in all brain areas that contain aromatase. On the other hand, the synthesis of aromatase should also be controlled by estrogens in an indirect, transynaptic manner very reminiscent of the way in which steroids indirectly control the production of LHRH. Fibers that are immunoreactive for tyrosine hydroxylase (synthesis of dopamine), dopamine beta-hydroxylase (synthesis of norepinephrine) or vasotocine have been identified in the close vicinity of ARO-ir cells in the POM and retrograde tracing has identified the origin of the dopaminergic and noradrenergic innervation of these areas. A few preliminary physiological experiments suggest that these catecholaminergic inputs regulate AA and presumably synthesis.
J Steroid Biochem Mol Biol 1996 Jan
PMID:Effects of testosterone and its metabolites on aromatase-immunoreactive cells in the quail brain: relationship with the activation of male reproductive behavior. 860 40


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