Determination of nuclear receptor corepressor interactions with the thyroid hormone receptor

The thyroid hormone receptor (TR) recruits the nuclear corepressors, nuclear receptor corepressor (NCoR) and silencing mediator of retinoid and thyroid hormone receptors (SMRT), to target DNA elements in the absence of ligand. While the TR preferentially recruits NCoR, the mechanism remains unclear. The corepressors interact with the TR via interacting domains (IDs) present in their C terminus which contain a conserved motif termed a CoRNR box. Despite their similarity, the corepressor IDs allow for nuclear receptor specificity. Here we demonstrate that NCoR stabilizes the TR homodimer when bound to DNA by preventing its dissociation from thyroid hormone response elements. This suggests that NCoR acts to hold the repression complex in place on target elements. The TR homodimer recruits NCoR through two of its three IDs, one of which is not present in SMRT. This unique ID, N3, contains a CoRNR box but lacks the extended helical motif present in each of the other IDs. Instead, N3 contains an isoleucine just proximal to this motif. This isoleucine is also conserved in N2 but not in the corresponding S2 domain in SMRT. On thyroid hormone response elements and in mammalian cells this residue is critical in both N3 and N2 for high-affinity TR binding. In addition, this residue also controls specificity for the interactions of TR with NCoR. Together these data suggest that the specific recruitment of NCoR by the TR through a unique motif allows for stabilization of the repression complex on target elements.     

Role of nuclear receptor corepressor RIP140 in metabolic syndrome

Obesity and its associated complications, which can lead to the development of metabolic syndrome, are a worldwide major public health concern especially in developed countries where they have a very high prevalence. RIP140 is a nuclear coregulator with a pivotal role in controlling lipid and glucose metabolism. Genetically manipulated mice devoid of RIP140 are lean with increased oxygen consumption and are resistant to high-fat diet-induced obesity and hepatic steatosis with improved insulin sensitivity. Moreover, white adipocytes with targeted disruption of RIP140 express genes characteristic of brown fat including CIDEA and UCP1 while skeletal muscles show a shift in fibre type composition enriched in more oxidative fibres. Thus, RIP140 is a potential therapeutic target in metabolic disorders. In this article we will review the role of RIP140 in tissues relevant to the appearance and progression of the metabolic syndrome and discuss how the manipulation of RIP140 levels or activity might represent a therapeutic approach to combat obesity and associated metabolic disorders. This article is part of a Special Issue entitled: Translating nuclear receptors from health to disease.     

TR surfaces and conformations required to bind nuclear receptor corepressor

Residues of the TR that are critical for binding the nuclear receptor corepressor (N-CoR) were identified by testing more than 100 separate mutations of the full-length human TRbeta that scan the surface of its ligand binding domain. The primary inferred interaction surface overlaps the surface described for binding of p160 coactivators, but differs by extending to a novel site underneath which helix 12 rests in the liganded TR, rather than including residues of helix 12. Nonconservative mutations of this surface diminished binding similarly to three isolated N-CoR receptor interaction domains (RIDs), but conservative mutations affected binding variably, consistent with a role for this surface in RID selectivity. The commonality of this surface in binding N-CoR was confirmed for the RXRs and ERs. Deletion of helix 12 increased N-CoR binding by the TR modestly, and by the RXR and ER to a much greater extent, indicating a competition between this helix and the corepressor that regulates the extent of corepressor binding by nuclear receptors. When helix 12 was deleted, N-CoR binding by the ER was stimulated by tamoxifen, and binding by the TR was stimulated by Triac, indicating that helix 12 is not the only feature that regulates corepressor binding. Two additional mutationsensitive surfaces were found alongside helix 1, near the previously described CoR box, and above helix 11, nearby but separate from residues that help link receptor in dimers. Based on effects of selected mutations on T(3) and coactivator binding, and on results of combined mutations of the three sites on corepressor binding, we propose that the second and third surfaces stabilize TR unliganded conformation(s) required for efficient N-CoR binding. In transfection assays mutations of all three surfaces impaired the corepressor-mediated functions of unliganded TR repression or activation. These detailed mapping results suggest approaches for selective modulation of corepressor interaction that include the shape of the molecular binding surface, the competitive occupancy by helix 12, pharmacological stimulation, and specific conformational stabilization.     

Identification and redistribution of lamins during nuclear differentiation in mouse spermatogenesis

Chromatin may be attached to the nuclear envelope through interaction of the nuclear membrane lamins A, B, and C. Such a hypothesis requires that these proteins are present in all cells with chromatin attachment to the nuclear envelope. We have investigated the distribution of the lamins during spermatogenesis in mouse, which exhibits extremes in nuclear envelope structural changes. By immunohistochemical techniques using human auto-antibodies and monoclonal antibodies against these molecules, we found that the lamins persist through all stages of spermatogenesis, though in highly variable amounts. They are also present during meiotic prophase (pachytene) when chromosomes are only locally attached to the nuclear envelope, analogous to the early prophase of somatic cells. Restructuring of the early spermatid nuclear envelope is accompanied by the appearance of a new lamin at the acrosomal fossa. In the epididymal spermatozoon the distribution of different lamins varies markedly over the nucleus suggesting special structural functions. The presence of lamins throughout spermatogenesis supports the concept that they are a general feature of the nuclear envelope structure, even where a lamina is not recognizable ultrastructurally.     

Interactions of nuclear receptor coactivator/corepressor proteins with the aryl hydrocarbon receptor complex.

MCF-7 human breast cancer cells express the aryl hydrocarbon receptor (AhR), and treatment with AhR agonists such as 2,3,7, 8-tetrachlorodibenzo-p-dioxin (TCDD) inhibits estrogen receptor (ER)-mediated responses. This study investigates physical and functional interactions of the AhR complex with a prototypical coactivator (estrogen receptor associating protein 140, ERAP 140) and corepressor (silencing mediator for retinoic acid and thyroid hormone receptor, SMRT) for ER and other members of the nuclear receptor superfamily. The AhR, AhR nuclear translocator (Arnt), and AhR/Arnt proteins were coimmunoprecipitated with 35S-ERAP 140 and 35S-SMRT and, in gel mobility shift assays, AhR/Arnt binding to 32P-dioxin response element (DRE) was enhanced by ERAP-140 and inhibited by SMRT; supershifted bands were not observed. In transactivation assays, coactivator and corepressor proteins enhanced or inhibited AhR-mediated gene expression; however, these responses varied with the amount of coactivator/corepressor expression. These results confirmed functional and physical interactions of AhR/Arnt with ERAP 140 and SMRT in breast cancer cells.     

The N-CoR-HDAC3 nuclear receptor corepressor complex inhibits the JNK pathway through the integral subunit GPS2

The corepressors N-CoR and SMRT partner with histone deacetylases (HDACs) in diverse repression pathways. We report here that GPS2, a protein involved in intracellular signaling, is an integral subunit of the N-CoR-HDAC3 complex. We have determined structural motifs that direct the formation of a highly stable and active deacetylase complex. GPS2 and TBL1, another component of the N-CoR-HDAC3 complex, interact cooperatively with repression domain 1 of N-CoR to form a heterotrimeric structure and are indirectly linked to HDAC3 via an extended N-CoR SANT domain that also activates latent HDAC3 activity. More importantly, we show here that the N-CoR-HDAC3 complex inhibits JNK activation through the associated GPS2 subunit and thus could potentially provide an alternative mechanism for hormone-mediated antagonism of AP-1 function.     

MicroRNA-23b downregulates peroxiredoxin III in human prostate cancer

To investigate the mechanism by which peroxiredoxin III (PRDX3) is altered in human prostate cancer (PCa), we used microRNA (miRNA) target prediction program and miRNA microarray to predict and identify miR-23b as a candidate miRNA that targets PRDX3. We showed that miR-23b suppresses PRDX3 protein expression in human DU145 cells under normal and hypoxic conditions. Additionally, the clinical significance of miR-23b and PRDX3 expression in PCa patients was also confirmed. In conclusion, our data suggest that the effects of PRDX3 in PCa progression may be caused by the regulation function of miR-23b, and consequently, miR-23b may be involved in the response of PCa cells to hypoxia stress.