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)

Human progesterone target tissues contain two progesterone receptors: B-receptors (hPRB), which are 933 amino acids in length, and A-receptors (hPRA), which lack the N-terminal 164 amino acids. The two isoforms differ functionally when they are occupied by agonists or antagonists. We postulated that the unique 164-amino acid, B-upstream segment (BUS) is in part responsible for the functional differences between the two isoforms and have constructed a series of hPR expression vectors encoding BUS fused to isolated down-stream functional domains of the receptors. These include the two transactivation domains: activation function-1 (AF1), located in a 90-amino acid segment just up-stream of the DNA-binding domain (DBD) and nuclear localization signal (NLS), and AF2, located in the hormone-binding domain. BUS is a highly phosphorylated domain, and contains the serine residues responsible for the hPRB triplet protein structure. The construct containing BUS-DBD-NLS binds tightly to DNA when aided by accessory nuclear factors. In HeLa cells, BUS-DBD-NLS strongly and autonomously activates transcription of chloramphenicol acetyltransferase (CAT) from a promoter containing two progesterone response elements (PRE2-TATAtk-CAT). Transcription levels with BUS-DBD-NLS are equivalent to those seen with full-length hPRB, and are higher than those seen with hPRA. BUS specifically requires an intact hPR DBD to be transcriptionally active. DBD mutants that cannot bind DNA or whose DNA binding specificity has been switched to an estrogen response element cannot cooperate in BUS transcriptional activity. The function of BUS-DBD-NLS is promoter and cell specific. It does not transactivate a CAT reporter driven by the mouse mammary tumor virus promoter in HeLa cells and poorly transactivates PRE2-TATAtk-CAT in PR-negative T47D breast cancer cells. However, in the breast cancer cells, BUS-DBD-NLS transactivation of PRE2-TATAtk-CAT can be reconstituted by either elevating cellular levels of cAMP or linking BUS and DBD to AF1 or AF2 of hPR, each of which alone is also inactive in these cells. We conclude that hPRB contains a unique third activation function (AF3) located within BUS and requiring the functional DBD of hPR. Depending on the promoter or cell tested, AF3 can activate transcription autonomously, or it can functionally synergize with AF1 or AF2. Autonomous AF3 function may explain the unexpected transactivating actions of antiprogestin-occupied hPRB, an issue of importance in hormone-resistant breast cancers and in tissue-specific agonist-like effects of hormone antagonists.
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PMID:A third transactivation function (AF3) of human progesterone receptors located in the unique N-terminal segment of the B-isoform. 785 52

When antagonist-occupied steroid receptors have agonist-like effects, the clinical consequences are grave. We present evidence that human progesterone B-receptors (hPRB) when occupied by progesterone antagonists, inappropriately activate transcription by an unusual mechanism that does not require the canonical progesterone response element (PRE). In HeLa cells cotransfected with a PRE-tk-chloramphenicol acetyltransferase reporter and a hPRB expression vector, strong transcription is seen not only when receptors are activated by the agonist R5020, but also in the presence of the three antiprogestins, RU486, ZK112993, and ZK98299. Human PRB occupied by ZK98299 do not bind to a PRE, suggesting that the transcriptional stimulation is independent of DNA binding. Indeed, a tk-chloramphenicol acetyltransferase promoter-reporter lacking the PRE loses transcriptional activation by the agonist, but retains transactivation by the three antagonists. The PRE-independent antagonist-induced transcription requires that hPRB have an intact DNA-binding domain, but hPR target gene specificity is not required, because a hPRB mutant that binds an estrogen response element still activates transcription. It appears that antagonist-occupied hPR activate transcription without binding to a PRE, perhaps by interacting with tethering proteins instead. Even a gene that is not a normal progesterone target could be aberrantly activated. Human cells contain equimolar amounts of hPRB and the N-terminally truncated natural isotype, hPRA. Unlike hPRB, hPRA are not transcriptionally activated by progesterone antagonists. We, therefore, tested the effects of antagonists when the two receptor isotypes are coexpressed and found that A-receptors can annul the inappropriate transcription by B-receptors. Thus, when both receptor forms are present, the hPRA phenotype is dominant. Moreover, pure hPRB/hPRA heterodimers, produced by fos/jun leucine zipper domain-hPR chimeras, also have the inactive transcriptional phenotype of hPRA. Our studies suggest not only that the two hPR isotypes are functionally quite different, but also that some of the agonist-like transcriptional effects of antagonist-occupied B-receptors proceed through novel mechanisms.
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PMID:Antagonist-occupied human progesterone B-receptors activate transcription without binding to progesterone response elements and are dominantly inhibited by A-receptors. 826 57