Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.1.21 (thymidine kinase)
 7,561 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Steroid hormones, which are ubiquitous regulators of physiologic processes, are produced primarily in the adrenals, gonads, and placenta. Each steroidogenic cell type produces different steroids due to cell-specific expression of various steroidogenic enzymes, but all steroidogenesis is initiated by P450scc, the mitochondrial enzyme that converts cholesterol to pregnenolone. We previously showed the unique segments of the P450scc promoter that are responsible for basal and cAMP-induced expression of this gene in the placenta are not employed for expression in the adrenal (C.C.D. Moore, D.W. Hum, and W.L. Miller, Mol. Endocrinol. 6, 2045-2058, 1992). We now show that sequences between -142 and -153 exhibit placental-specific activator activity. Sequences between -131 and -155 can confer activator activity to a 32-bp promoter from the thymidine kinase gene of herpes simplex virus in an orientation-independent fashion. Two protein complexes, termed IV and VII, interact specifically with DNA from -131 to -155. Mutating bases -142 to -151 abolishes formation of complex VII and partially inhibits complex IV, suggesting that the proteins forming these complexes bind neighboring segments of DNA. Mutating only two cytosines at bases 141 and 142 also eliminates the formation of complex VII and reduces the transcriptional activity of the activator by about 75-80%, indicating that complex VII is important for placental expression of P450scc. The sequence from -140 to -149 on the antisense strand resembles an NF-kappa B binding site. Antibodies to NF-kappa B subunit p50, but not to p52, p65, or c-Rel, will supershift some but not all of complex IV, whereas none of these antibodies interact with complex VII. A consensus NF-kappa B oligonucleotide does not form complex IV, suggesting that p50 interacts with the protein component, but not the DNA component of complex IV. Photoaffinity UV cross-linking yielded single bands of cross-linked DNA-protein complexes at approximately 85 kD for complex IV and approximately 70 kD for complex VII, indicating that separate proteins form complexes IV and VII. Southwestern blotting identified a single protein of 55 kD forming complex VII but did not identify the protein forming complex IV. Bandshifts and Southwestern blots with nuclear extracts from steroidogenic human placental JEG-3 cells and human adrenal NCI-H295 cells show that this 55-kD protein is found in placental but not adrenal cells. This 55-kD nuclear protein appears to be a trans-acting factor necessary for placental but not adrenal expression of P450scc.
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PMID:Characterization of placental transcriptional activation of the human gene for P450scc. 774 95

The CYP11A1 gene encodes the cholesterol side-chain cleavage enzyme P450scc, which catalyzes the synthesis of steroids from cholesterol. This gene is expressed only in steroidogenic organs such as the adrenal, gonad, placenta, and brain. We have characterized an upstream regulatory element of the human CYP11A1 gene, termed AdE, which contributed to its cell type-specific expression. The AdE sequence contains two protein binding regions, AdE1 and AdE2, which bind many proteins including NF1- and Sp1-like proteins as shown by electrophoretic mobility shift assay, footprinting, competition, antibody supershift, and mutagenesis of the binding sites. When cloned in front of the CYP11A1 promoter or the heterologous thymidine kinase promoter, AdE sequences enhanced expression of the reporter gene in steroidogenic cell lines of the adrenal, gonad, and placental origin but not in nonsteroidogenic cell lines such as COS-1 and Rat-1. The function of AdE1 and AdE2 was lower when present individually than together. The combined action of multiple transcription factors binding to the AdE sequence brings about the final activation of the CYP11A1 gene in a tissue-specific manner.
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PMID:Characterization of the upstream sequence of the human CYP11A1 gene for cell type-specific expression. 870 23

In preliminary studies we demonstrated that the CYP11B1 (11beta-hydroxylase) promoter could direct specific expression of a suicide gene in adrenocortical cancer cells, providing a potentially specific therapeutic option for adrenocortical cancer. In this present study we describe our attempts to enhance the activity of the CYP11B1 promoter while maintaining its specificity for adrenal cells. Using a putative enhancer element from the cholesterol side-chain cleavage (P450scc) gene, the activity of the CYP11B1 promoter in and its specificity for adrenocortical cells were enhanced. Treatment with 8-bromo-cAMP or forskolin resulted in further enhancement. In stably transfected Y-1 cells, in which the herpes simplex virus thymidine kinase (HSV-TK) gene was driven by the CYP11B1 promoter with the P450scc enhancer element, HSV-TK expression and ganciclovir sensitivity were augmented by treatment with 8-bromo-cAMP, forskolin, and ACTH. In summary, we report the construction of a suicide HSV-TK vector with preferential toxicity to adrenocortical cells. We propose that a similar strategy using differentiating agents may be useful in the gene therapy of tumors with unique differentiated properties, including those arising from other endocrine organs.
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PMID:Construction of gene therapy vectors targeting adrenocortical cells: enhancement of activity and specificity with agents modulating the cyclic adenosine 3',5'-monophosphate pathway. 1063 96

Cytochrome P450scc catalyzes the important first step in the steroid synthesis pathway; however, it is clear that additional factors regulating the temporal and spacial specific expression of the CYP11A1 gene remain to be identified. To isolate novel transcription factors that regulate this gene, a cis-acting element of the 5'-flanking region from nucleotides -155 to -131 (-155/-131) was used to screen a human placental lambda gt11 cDNA expression library, and an interacting clone was isolated. The open reading frame of the cDNA encodes several domains that are characteristic of transcription factors including an acidic region, a region rich in prolines and three zinc-finger motifs. Expression of the cDNA by in vitro transcription/translation and by transient transfection in HeLa cells yielded a protein of 132 kDa, which concurs with the predicted size. Transfection of the cDNA in placental JEG-3 and adrenal NCI-H295 cells, stimulate expression of a reporter construct controlled by the P450scc gene 5'-flanking region from nucleotides -1676 to +49. This transcriptional regulating protein of 132kDa (TReP-132) when expressed in HeLa cells was demonstrated to interact with the -155/-131 region in bandshift analysis, and tandem copies of this region was shown to confer activation of the heterologous HSV thymidine kinase minimal promoter. Coexpression of CBP/p300 with TReP-132 further increased promoter activity, and the proteins were demonstrated to interact physically. RNA analysis demonstrated the highest levels of expression in the adrenal cortex and testis; and transcript expression is also found in the steroidogenic JEG-3, NCI-H295, and MCF-7 cell lines, but not in non-steroidogenic HepG2 and HK293 cells. Subsequently it has been shown that TReP-132 interacts with steroidogenic factor-1 (SF-1) through specific domains; and along with the interaction with CBP/p300 these factors are postulated to form a complex to regulate expression of the P450scc gene.
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PMID:Function of the transcriptional regulating protein of 132 kDa (TReP-132) on human P450scc gene expression. 1253 Jun 63