Mechanisms of nuclear localization of the progesterone receptor: Evidence for interaction between monomers

Guiochon‐Mantel, Anne; Loosfelt, Hugues; Lescop, Pierre; Sar, Sokhavuth; Atger, Michel; Perrot‐Applanat, Martine; Milgröm, Edwin

doi:10.1016/0092-8674(89)90052-4

Cited by 320 publications

(153 citation statements)

References 44 publications

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“…In particular, the cotransport of nuclear import-deficient nuclear receptors to the nucleus by wild type partners has been used to demonstrate that nuclear receptors dimerize in solution subsequent to ligand binding (11,44,50). In the absence of rapid nucleocytoplasmic shuttling, our results would imply that these receptor interactions occur more efficiently than has previously been appreciated.…”

Section: Figcontrasting

confidence: 48%

“…Heterokaryon fusion assays have indicated that, while localized to the nucleus, nuclear receptors, including liganded GR, traffic continuously and transiently to the cytoplasm (11)(12)(13)(14)(15)(16). Upon withdrawal of steroid, shuttling continues for GR as the receptor reassembles into chaperone complexes and slowly reaccumulates in the cytoplasm over a period of several hours (17)(18)(19).…”

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confidence: 99%

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Nuclear Export of the Glucocorticoid Receptor Is Accelerated by Cell Fusion-dependent Release of Calreticulin

Walther

Lamprecht

Ridsdale

et al. 2003

Journal of Biological Chemistry

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Nucleocytoplasmic exchange of nuclear hormone receptors is hypothesized to allow for rapid and direct interactions with cytoplasmic signaling factors. In addition to recycling between a naïve, chaperone-associated cytoplasmic complex and a liganded chaperone-free nuclear form, the glucocorticoid receptor (GR) has been observed to shuttle between nucleus and cytoplasm. Nuclear export of GR and other nuclear receptors has been proposed to depend on direct interactions with calreticulin, which is predominantly localized to the lumen of the endoplasmic reticulum. We show that rapid calreticulin-mediated nuclear export of GR is a specific response to transient disruption of the endoplasmic reticulum that occurs during polyethylene glycol-mediated cell fusion. Using live and digitonin-permeabilized cells we demonstrate that, in the absence of cell fusion, GR nuclear export occurs slowly over a period of many hours independent of direct interaction with calreticulin. Our findings temper expectations that nuclear receptors respond rapidly and directly to cytoplasmic signals in the absence of additional regulatory control. These results highlight the importance of verifying findings of nucleocytoplasmic trafficking using techniques in addition to heterokaryon cell fusion.Nuclear hormone receptors are dynamic transcription factors that move rapidly through the nucleus and that, based on the results of heterokaryon fusion assays, are believed to shuttle or exchange rapidly between nucleus and cytoplasm. Shuttling offers the potential for rapid modulation of receptor function in response to cytoplasmic signaling pathways. Nuclear receptors are imported into the nucleus through the karyopherin␣/␤ pathway (1, 2). How nuclear receptor export is accomplished is less clear, although recent reports have suggested an integral role for calreticulin (CRT) 1 (3, 4), a calcium binding protein localized to the lumen of the endoplasmic reticulum (5). The naïve glucocorticoid receptor (GR) is a cytoplasmic protein, which is held in a chaperone complex anchored by hsp90 and containing hsp70, immunophilins, and other factors, including p23, where it is poised to bind ligand (6). Upon ligand binding, the chaperone complex is dissociated, and the receptor moves rapidly to the nucleus to regulate specific gene transcription (7). Within the nucleus, the receptor becomes localized to specific sites but exchanges very rapidly with chromatin and remains highly mobile (8). Ligand binding and transcriptional regulation are transient events, with molecular chaperones also being involved in the disassembly of regulatory complexes (9, 10).Heterokaryon fusion assays have indicated that, while localized to the nucleus, nuclear receptors, including liganded GR, traffic continuously and transiently to the cytoplasm (11-16). Upon withdrawal of steroid, shuttling continues for GR as the receptor reassembles into chaperone complexes and slowly reaccumulates in the cytoplasm over a period of several hours (17)(18)(19). Depending on the cell type, the time req...

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Section: Figcontrasting

confidence: 48%

mentioning

confidence: 99%

Nuclear Export of the Glucocorticoid Receptor Is Accelerated by Cell Fusion-dependent Release of Calreticulin

Walther

Lamprecht

Ridsdale

et al. 2003

Journal of Biological Chemistry

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“…[38], who showed that binding of a glucocorticoid-recep-.or DNA-binding domain to HRE occurs preferentially at one ialf site and is followed by the glucocorticoid-receptor DNA-binding domain occupying the other half site [ 191 via protein-protein interactions [38]. Interaction between soluble progesterone-receptor monomers does not occur in vivo after deletion of the steroid-binding domain [8]. The latter conclusion was reached using indirect evidence based on nuclear cotransport experiments.…”

Section: Discussionmentioning

confidence: 99%

“…Previous experiments [8] have shown that this deletion does not modify the main biological properties of the receptor. COS-7 cells were cotransfected with expression vectors encoding wild-type receptors and Let 126-mutant receptors (Let 126-mutant lacks the Let 126 epitope).…”

Section: Dimerization Of the Wild-type Receptor And Absence Of Dimerimentioning

confidence: 99%

Specific binding of progesterone receptor to progesterone‐responsive elements does not require prior dimerization

Cohen‐Solal

Bailly

Rauch

et al. 1993

European Journal of Biochemistry

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Steroid-hormone receptors undergo, prior to binding to DNA, a hormone-dependent dimerization. It is generally accepted that this dimerization is indispensable for the high-affinity binding of hormone receptor to hormone-responsive elements.Using a progesterone-receptor mutant with the complete steroid-binding domain deleted (positions 663 -930), with or without the epitope required for binding the monoclonal antibody Let 126, we have shown that this receptor species was unable to undergo dimerization in solution. However, this mutant retained a high affinity (60-70% of the affinity of the wild-type receptor) for the progesterone-responsive elements of the mouse-mammary-tumor-virus long-terminal-repeat promoter and for a consensus palindromic. progesterone-responsive element, as measured by both DNase-I protection experiments and gel-shift experiments. This mutant also increased gene transcription. Thus, at least in the case of the progesterone receptor, prior dimerization is dispensable for receptor binding to regulatory DNA elements and for subsequent transcription activation.

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“…For GFP-⌬(NLS)-SRC-1/DsRed-PR and ⌬(NLS)-PR/HA-SRC-1 cotransport experiments, the corresponding expression plasmids were transfected at a ratio of 1:5. Cells were fixed and processed as previously described (7). For PR detection, we used the Mi60 monoclonal antibody (54) at a final concentration of 8 g/ml.…”

Section: Methodsmentioning

confidence: 99%

Subcellular Localization and Mechanisms of Nucleocytoplasmic Trafficking of Steroid Receptor Coactivator-1

Amazit¹,

Alj²,

Tyagi³

et al. 2003

Journal of Biological Chemistry

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Steroid hormone receptors are ligand-stimulated transcription factors that modulate gene transcription by recruiting coregulators to gene promoters. Subcellular localization and dynamic movements of transcription factors have been shown to be one of the major means of regulating their transcriptional activity. In the present report we describe the subcellular localization and the dynamics of intracellular trafficking of steroid receptor coactivator 1 (SRC-1). After its synthesis in the cytoplasm, SRC-1 is imported into the nucleus, where it activates transcription and is subsequently exported back to the cytoplasm. In both the nucleus and cytoplasm, SRC-1 is localized in speckles. The characterization of SRC-1 nuclear localization sequence reveals that it is a classic bipartite signal localized in the N-terminal region of the protein, between amino acids 18 and 36. This sequence is highly conserved within the other members of the p160 family. Additionally, SRC-1 nuclear export is inhibited by leptomycin B. The region involved in its nuclear export is localized between amino acids 990 and 1038. It is an unusually large domain differing from the classic leucine-rich NES sequences. Thus SRC-1 nuclear export involves either an alternate type of NES or is dependent on the interaction of SRC-1 with a protein, which is exported through the crm1/exportin pathway. Overall, the intracellular trafficking of SRC-1 might be a mechanism to regulate the termination of hormone action, the interaction with other signaling pathways in the cytoplasm and its degradation.

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Mechanisms of nuclear localization of the progesterone receptor: Evidence for interaction between monomers

Cited by 320 publications

References 44 publications

Nuclear Export of the Glucocorticoid Receptor Is Accelerated by Cell Fusion-dependent Release of Calreticulin

Nuclear Export of the Glucocorticoid Receptor Is Accelerated by Cell Fusion-dependent Release of Calreticulin

Specific binding of progesterone receptor to progesterone‐responsive elements does not require prior dimerization

Subcellular Localization and Mechanisms of Nucleocytoplasmic Trafficking of Steroid Receptor Coactivator-1

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