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| (1) Nuclear receptors |
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| Protein | Model(s)/putative function |
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| AR | The AR/SHP interaction leads to >95% inhibition of AR via the LXXLL motifs. Mechanisms involve inhibition of AR ligand-binding domain and AR N-terminal domain-dependent transactivation and competing with AR coactivators [23]. |
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| CAR, RAR, TR | HepG2 and JEG-3 cells/early evidence that SHP interacts with several receptor superfamily members and inhibits transactivation. CAR is an NR-inducing CYP2 and CYP3 genes involved in the metabolism of xenobiotics [10, 24]. |
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| DAX-1 | Human embryonic kidney 293 cells/beside individual homodimerization of DAX1 and SHP, this is the first evidence of DAX1-SHP heterodimerization in the nucleus of mammalian cells. Involvement of the LXXLL motifs and AF-2 domain of DAX1 in this interaction. Distinct functions for SHP (different from transcriptional repressor) are anticipated [141, 142]. |
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| ER | 293 human embryo kidney cells, Cos7 kidney cells/direct inhibitory binding of SHP to ERs via LXXLL-related motifs to the AF-2 domain [21]. RL95-2 human endometrial carcinoma cells/SHP inhibits the agonist activity of 4-hydroxytamoxifen displaying a potent inhibitory effect for Erα > ERβ. Direct interaction of SHP with ER and inhibition of ER transcriptional activity [143]. Prevention of tamoxifen-induced estrogen agonistic effects and neoplastic changes in the endometrium in women with breast cancer taking tamoxifen? |
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| ERRγ | HeLa (human cervical carcinoma), CV-1 (green monkey kidney), and HEK 293 (human embryonic kidney) cell lines/SHP inhibits ERRγ transactivation by physical interaction with the 3 members of the ERR subfamily. Interaction is dependent on N-terminal receptor interaction domain of SHP and AF-2 surface of ERRγ. Part of the autoregulatory mechanism of gene expression going through ERRγ/SHP/ERRγ. A potential role in some forms of moderate human obesity during SHP mutations [133]. |
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| GR | 293 human embryo kidney cells and COS-7 monkey kidney/SHP inhibits the transcriptional activity of GR via the LXXLL motif. Physiological role of SHP in glucocorticoid signaling and gluconeogenesis [22]. See also HNF4 [90] and Foxo1 [115]. |
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| HNF4 | Human ANG transgenic mice and HepG2 cells treated with bile acids/evidence that bile acids negatively regulate the human ANG gene through the FXR/SHP-mediated process (inhibition of the binding of HNF4 to the ANG promoter) [90]. Mechanisms: SHP binds the AF-2 region and the N-terminal region of HNF4 and inhibits the binding of HNF4 to DNA. Also, modulation of HNF4 activity by SHP has important metabolic effects and interacts with the pathway of gluconeogenesis [47] (see text and Foxo1) [115]. |
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| LRH-1 | HepG2 cells/SHP interacts directly with the orphan receptor LRH-1 (AF-2 surface) and competes with other coactivators, leading to repression of LRH-1 transcriptional activity [48]. Demonstration that repression of CYP7A1 and bile acid synthesis requires coordinate interaction/transcription of FXR/LRH-1/SHP autoregulatory cascade, essential for maintenance of bile acid-induced negative feedback, and therefore hepatic cholesterol metabolism [65] (see also Figure 2). |
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| LXRα | In vitro experiments and in vivo human colon Caco-2 cells/SHP directly inhibits the transcriptional activity of LXRα via the AF-2 domain. Relevance for direct downregulation of specific LXR target genes (controlling CYP7A1, ABCA1, ABCG1, ABCG5, ABCG8, CETP, ApoE, SREBP-1c) and therefore cholesterol-bile acid homeostasis [144]. |
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| Nur77 (NGFI-B) | HepG2 cells/Nur77 plays a key role in apoptosis of many cell types and cancer cells. Evidence that SHP functions to repress the transcriptional function of Nur77 (binding coactivator CBP, see elsewhere in this table). SHP plays a protective role in the Nur77-mediated apoptosis in liver. Mutations in SHP: a role also for affect initiation and progression of inflammatory liver diseases such as alcoholic hepatitis and hepatic viral infections? [32]. |
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| PPARα | In vitro binding assays and in vivo experiments/the promoter regions of the genes encoding the first two enzymes of the peroxisomal beta-oxidation pathway (AOx, HD), contain transcriptional regulatory sequences (PPRE) bound by the PPARα/RXRα heterodimeric complex. SHP-inhibited transcription by PPARα/RXRα heterodimers from the AOx-PPRE. SHP potentiated transcription by PPARα/RXRα heterodimers from the HD-PPRE (evidence of SHP-dependent upregulation PPARα-mediated gene transcription) [46]. |
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| PPARγ | In vitro experiments, COS-7 cells/Shp gene expressed also in adipose tissue. SHP induces PPAR activation via C terminus (direct binding to the DBD/hinge region of PPARγ) and inhibition of the repressor activity of NCoR. SHP may act as an endogenous enhancer of PPARγ by competing with NCoR [45]. Mutant SHP proteins display less enhancing activity for PPARy compared with wild-type SHP, and a human model leading to mild obesity and insulin resistance has been described in Japanese during naturally occurring mutations [111] (see also text and Table 3). |
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| PXR | In vitro experiments, human hepatocytes, mouse model on cholic acid-supplemented diet/SHP act as potent repressor of PXR transactivation. Upon sensing xenobiotics and bile acid precursors, PXR controls CYP3A gene induction and inhibits CYP7α, acting on both bile acid synthesis and catabolism. PXR function might be also inhibited in the presence of cholic acid, chenodeoxycholic acid-dependent SHP upregulation [1]. |
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| RXR | HepG2 cells/demonstration that SHP acts as a transcriptional repressor for RXR. Full inhibition by SHP requires its direct repressor activity [47]. |
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| SHP | Human embryonic kidney 293 cells/LXXLL motifs and AF-2 domain are involved in SHP homodimerization in the nucleus (similarly to DAX1-SHP heterodimerization). NR0B family members use similar mechanisms for homodimerization as well as heterodimerization. Distinct functions for SHP (different from transcriptional repressor) are anticipated [141, 142]. |
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| (2) Transcription factors |
|
| Protein | Model(s)/putative function |
|
| ARNT | RL95-2 human endometrial carcinoma cells/TCDD binds to AHR (a member of bHLH-PAS family of transcription factors). Studies on physical and functional interaction of SHP with the ligand AHR/ARNT heterodimer showed that SHP inhibits the transcriptional activity of ARNT (not AHR) in vitro. Consequent inhibition of binding of AHR/ARNT to XREs. [41]. Relevance for expression of several genes involved in drug and hormone metabolism [145]. |
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| BETA2/NeuroD | 293T, COS-7, CV-1 cells/BETA2/NeuroD is a member of tissue-specific class B bHLH proteins and cats as a positive regulator of insulin gene expression [146] and neuronal differentiation [147]. SHP physically interacts and inhibits helix-loop-helix transcription factor BETA2/NeuroD transactivation of an E-box reporter in mouse pancreas islets. The inhibitory effect of SHP requires its C-terminal repression domain, interference with coactivator p300 for binding to BETA2/NeuroD, and direct transcriptional repression function. Relevance for development of the nervous system and the maintenance and formation of pancreatic and enteroendocrine cells [148]. |
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| C/EBPα | HepG2 hepatoma cells/SHP interacts directly with C/EBPα and represses C/EBPα-driven PEPCK gene transcription. Overall, a role for SHP in regulation of hepatic gluconeogenes is driven by C/EBPα activation in the liver [149]. |
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| Foxo1 | C57BL/6J mice and HepG2 and HEK293T cells/treatment with chenodeoxycholic acid was associated with FXR-dependent SHP induction, downregulation of gluconeogenic gene expression (G6Pase, PEPCK, FBP1), interaction of the forkhead transcription factor Foxo1 with SHP, and repression of Foxo1-mediated G6Pase transcription (competition with CBP). A similar mechanism is postulated for SHP-driven HNF-4 repression of PEPCK, FBP1 transcription. A mechanism by which bile acids metabolism is linked to gluconeogenic gene expression via an SHP-dependent regulatory pathway [115]. |
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| HNF3 (Foxa) | HepG2, 293T, NIH3T3, and HeLa cells, primary hepatocytes/SHP physically interacts and inhibits the transcriptional activity of the forkhead transcription factor HNF3 (isoforms α, β, γ). Relevance for SHP-driven regulation of gluconeogenic genes encoding G6Pase, PEPCK, and bile acid synthesis (CYP7A1), via inhibition of DNA-binding of HNF3 [51]. |
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| Jun D | Two rat models of liver fibrosis and Hepatic Stellate cells (HSC)/promoting the ligand-induced FXR-SHP cascade (by the FXR ligand 6-EDCA, in rat models) and overexpressing SHP in HSC prevented fibrogenic changes in the liver. SHP binds JunD and inhibits DNA binding of adaptor protein (AP)-1 induced by thrombin. FXR ligands as therapeutic agents to treat liver fibrosis? [52]. |
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| NF-κB | Murine macrophage cell line RAW 264.7/SHP acts as a positive transcription coactivator of NF-κB and essential for NF-κB transactivation by palmitoyl lysophosphatidylcholine (one of the oxLDL constituents). Relevance for differentiation mechanism of resting macrophage cells into foam cells and resulting atherogenesis (see also [44]). |
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| Smad | HepG2, CV-1, and HeLa cells/SHP represses Smad3-induced transcription by competing for the coactivator p300. SHP therefore represses TGF-β-induced gene expression. Relevance for TGF-β-dependent regulation of cell growth, apoptosis, carcinogenesis, and regeneration following liver injury [40]. SHP-Smad3 interaction similar to SHP-BETA2/NeuroD [148]. |
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| TRAF6, p65 | Macrophages/a novel function of SHP in innate immunity involving Toll-like receptors (TLRs). SHP negatively regulates TLR signaling to NF-κB. Likely, SHP negatively regulates immune responses initiated by various pathogen-recognition receptors by forming a complex with TRAF6 and effect on TRAF6 ubiquitination. In the cytosol of LPS-stimulated cells. SHP also acts as specific transrepressor of the transcription factor p65 (part of the p50/p65 heterodimer found in NF-κB). An additional role for SHP in sepsis and inflammatory disease? [128, 129]. |
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| (3) Transcriptional coregulators |
|
| Protein | Model(s)/putative function |
|
| Brm, BAF155, BAF47, mSin3A, Swi/Snf | HepG2 cells/The CYP7A1 gene was used as a model system. SHP has direct interaction with corepressors at the level of native chromatin. SHP directly interacted and mediated the recruitment of mSin3A-Swi/Snf-Brm chromatin remodelling complex to the CYP7A1 promoter (TATA and BARE II region of the promoter). Also, the mSinA3/HDAC1 corepressor complex is inhibiting transcription by histone deacetylation. SHP also interacted with known proteins belonging to the Swi/Snf complex (BAF155, BAF47). This mechanism explains the complex and subtle SHP-driven inhibition of hepatic bile acid synthesis [50]. |
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| CBP | HepG2 cells, CV-1 cells/SHP binds coactivator CBP and competes with Nur77. The mechanism explains the repression of the transcriptional function of Nur77, which is fundamental in apoptosis in the liver [32]. |
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| EID-1 | Cos-7 cells/SHP specifically interacts with EID-1 providing inhibitory mechanisms. EID-1 (a non-HDAC cofactor) acts as inhibitor of the coregulator complex EID1–p300–CBP. Results clarify essential repression mechanisms of SHP involving coinhibitory factors (upstream targets) distinct from NRs corepressor [12, 150]. |
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| G9a, HDAC-1 | Caco-2, HepG2, HeLa, Cos-1 cells/SHP localized exclusively in nuclease-sensitive euchromatin regions. SHP can functionally interact with HDAC-1 (HDAC of class I) and the euchromatic histone 3 methylase G9a, and the unmodified K9-methylated histone 3 [151]. Additional data on mechanisms involved SHP-driven repressive activity, involving also target genes regulated by G9a and SHP-mediated inhibition of hepatic bile acid synthesis via coordinated chromatin modification at target genes [152]. |
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| GPS2 | Cos-7, HepG2, Huh7 cells/SHP negatively interacts with GPS2 (a stoichiometric subunit of the NR corepressor, N-Cor) complex, involved in bile acid synthesis and differential coregulation of CYP7A1 and CYP8B1 expression [67]. |
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| SIRT1 | HepG2, HEK293T (293T), and HeLa cells/SIRT1 is a HDAC of class III. SHP recruits SIRT1 (activating deacetylase activity of SIRT1) to repress LRH1 transcriptional activity as well as inhibition LRH1 target gene promoter activity and mRNA levels. A novel mechanism is described for SHP repressive action and control of bile acid homeostasis. SIRT1 in working concertedly with NRs and affecting chromatin remodeling in target gene promoters [42]. |
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| SMRT/NcoR | Hepatoma cell lines/studies on the role of SHP in CAR-mediated transactivation of the CYP2B gene. SHP might interact with subunits of functionally distinct coregulator complexes, including HDAC3-N-CoR-SMRT [24, 120]. |
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| (4) Others |
|
| Factor | Model(s)/putative function |
|
| miRNA-206 | SHP−/− mice/SHP as an important transcriptional activator of miRNA-206 gene expression via a cascade dual inhibitory mechanism involving AP1 but also YY1 and ERRγ. Relevance for multiple steps involving cellular development, proliferation, and differentiation [153]. |
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| RNA Pol II | Caco-2 cells/within the pathway of SHP-LXR interaction, it is shown that SHP can interact in vitro with RNA polymerase II but not with TFIID and TFIIE transcription initiation factor II D (TFIID), general transcription factor II E (TFIIE) (components of the basal transcription machinery). A further mechanism by which SHP could inhibit both basal and induced transactivation [144]. |
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