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

The rat steroid cytochrome P450 17 alpha-hydroxylase/c17-20 lyase (rP450c17) gene is transcriptionally regulated in steroidogenic tissues. Previous studies showed that one DNA element located between -75 and -50 base pairs (bp) upstream from the transcriptional initiation site mediated both the basal and cAMP-regulated transcription of rP450c17. Using a series of mutant oligonucleotides in gel mobility shift assays and in functional assays, it is now shown that a core sequence of 12 bp, located at -58/-69 bp, is essential for nuclear protein binding and transcriptional activation. Mutant oligonucleotides cloned into a luciferase reporter gene construct containing a heterologous thymidine kinase promoter, transfected into mouse Leydig MA-10 and adrenocortical Y-1 cells, gave results consistent with those of gel shift assays. Mutants that abolished binding of the nuclear protein to DNA abolished the basal transcription of the gene as well as the responsiveness to cAMP, whereas those mutants that did not abolish binding of the nuclear protein to DNA still showed strong basal transcription as well as responsiveness to cAMP. Comparison of the binding sequence with the consensus binding site for the orphan nuclear receptor steroidogenic factor-1 (SF-1) showed that eight of nine bases were identical. However, the sequence from rP450c17 includes an additional three bases at the 5'-end, not previously demonstrated to be important for SF-1 binding. Recombinant rat SF-1 protein expressed in Escherichia coli binds to this sequence, and antibodies raised against rat SF-1 abolish binding of both recombinant SF-1 and the nuclear protein from Y-1 and MA-10 cells. These observations demonstrate that this region of the rP450c17 gene is responsible for both the basal transcription and cAMP inducibility and is bound by the orphan nuclear receptor SF-1. It is further shown that SF-1 can be phosphorylated in vitro by protein kinase A. This phosphorylation occurs at serine and threonine residues and results in decreased binding to the rP450c17 -58/-69 element. Since SF-1 mediates cAMP-induced transcriptional regulation of the rat P450c17 gene, phosphorylation of SF-1 via protein kinase A is likely to play a regulatory role in transcriptional activation.
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PMID:The orphan nuclear receptor steroidogenic factor-1 regulates the cyclic adenosine 3',5'-monophosphate-mediated transcriptional activation of rat cytochrome P450c17 (17 alpha-hydroxylase/c17-20 lyase). 882 55

The Dax-1 gene encodes a protein that is structurally related to members of the orphan nuclear receptor superfamily. Dax-1 is coexpressed with another orphan nuclear receptor, steroidogenic factor-1 (SF-1), in the adrenal, gonads, hypothalamus, and pituitary gland. Mutations in Dax-1 cause adrenal hypoplasia congenita, a disorder that is characterized by adrenal insufficiency and hypogonadotropic hypogonadism. These developmental and endocrine abnormalities are similar to those caused by disruption of the murine Ftz-F1 gene (which encodes SF-1), suggesting that these nuclear receptors act along the same developmental cascade. Cloning of the murine Dax-1 gene revealed a candidate SF-1-binding site in the Dax-1 promoter. In transient expression assays in SF-1-deficient JEG-3 cells, SF-1 stimulated expression of the Dax-1 promoter. However, deletion or mutation of the consensus SF-1-binding site did not eliminate SF-1 stimulation. Further analyses revealed the presence of a cryptic SF-1 site that creates an imperfect direct repeat of the SF-1 element. When linked to the minimal thymidine kinase promoter, each of the isolated SF-1 sites was sufficient to mediate transcriptional regulation by SF-1. Mutation of both SF-1 sites eliminated SF-1 binding and stimulation of the Dax-1 promoter. Unexpectedly, mutation of either half of the composite SF-1 sites increased basal activity in JEG-3 cells, suggesting interaction of a repressor protein. Gel shift analyses of the composite response element revealed an additional complex that was not supershifted by SF-1 antibodies. This complex was eliminated by mutation of either half-site, and it was supershifted by antibodies against chicken ovalbumin upstream promoter-transcription factor (COUP-TF). We propose that Dax-1 is stimulated by SF-1, and that SF-1 and COUP-TF provide antagonistic pathways that converge upon a common regulatory site.
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PMID:The murine Dax-1 promoter is stimulated by SF-1 (steroidogenic factor-1) and inhibited by COUP-TF (chicken ovalbumin upstream promoter-transcription factor) via a composite nuclear receptor-regulatory element. 965 5