Characterization of receptor-interacting protein 140 in retinoid receptor activities.

Receptor-interacting protein 140 (RIP140) contains multiple receptor interaction domains and interacts with retinoic acid receptors in a ligand-dependent manner. Nine LXXLL receptor-interacting motifs are organized into two clusters within this molecule, each differentially interacting with retinoic acid receptor (RAR) and retinoid X receptor (RXR). RAR interacts with the 5' cluster, whereas RXR interacts with both clusters. Additionally, a third ligand-dependent receptor-interacting domain is assigned to the very C terminus of this molecule, which contains no LXXLL motif. In mammalian cells, receptor heterodimerization is required for efficient interaction of RAR/RXR with RIP140. Furthermore, the heterodimeric, holoreceptors cooperatively interact with RIP140, which requires the activation function 2 domains of both receptors. By using different retinoic acid reporter systems, it is demonstrated that RIP140 strongly suppresses retinoic acid induction of reporter activities, but coactivator SRC-1 enhances it. Furthermore, an intrinsic repressive activity of RIP140 is demonstrated in a GAL4 fusion system. Unlike receptor corepressor, which interacts with antagonist-bound RAR/RXRs, RIP140 does not interact with antagonist-occupied RAR/RXR dimers. These data suggest that RIP140 represents a third coregulator category that is able to suppress the activation of certain agonist-bound hormone receptors.     

Effects of retinoid ligands on RIP140: molecular interaction with retinoid receptors and biological activity

Receptor interacting protein 140 (RIP140) interacts with retinoic acid receptor (RAR) and retinoid X receptor (RXR) constitutively, but hormone binding enhances this interaction. The ligand-independent interaction is mediated by the amino and central regions of RIP140 which contain a total of nine copies of the LXXLL motif, whereas the agonist-induced interaction is mediated by its carboxyl terminus which contains a novel motif (1063-1076, LTKTNPILYYMLQK). The ligand-independent interaction could be enhanced slightly by agonists, whereas the ligand-dependent interaction was strictly agonist dependent for both RAR and RXR. In the context of heterodimers, ligand occupancy of RXR played a more dominant role for both molecular interaction and biological activity of RIP140. Competition and mutation studies demonstrated an essential role for (1067)Asn and (1073)Met for a ligand-dependent interaction. A model was proposed to address the constitutive and agonist-dependent interaction of RIP140 with RAR/RXR.     

Limiting effects of RIP140 in estrogen signaling: potential mediation of anti-estrogenic effects of retinoic acid

The receptor interacting protein 140 (RIP140) belongs to a unique subclass of nuclear receptor coregulators with the ability to bind and repress the action of a number of agonist-bound hormone receptors. We have previously demonstrated that all-trans-retinoic acid (RA) induction of RIP140 constitutes a rate-limiting step in the regulation of retinoid receptor signaling. Here we demonstrate that RIP140 is also a limiting regulator of estrogen receptor signaling. Overexpression of RIP140 dose dependently inhibits estrogen-dependent reporter activity in human breast cancer cells. Furthermore, small interfering RNA to RIP140 enhances estrogen-dependent signaling. Our previous studies indicate that RIP140 is a direct target of RA. We report here that RA can abrogate estrogen-mediated cell cycle re-entry. In addition, RA treatment of estrogen-dependent breast cancer cells opposes estrogen receptor-dependent reporter activity, implying that a proportion of RA effects are anti-estrogenic. We provide evidence for a role for RIP140 in mediating anti-estrogenic effects of RA. RIP140 small interfering RNA blocks RA-mediated repression of estrogen receptor activity and provides a growth advantage to estrogen-dependent cells. Together these data implicate a regulatory role for RIP140 in mediating anti-estrogenic effects of RA in estrogen-dependent breast cancer cells and suggest that acute regulation of coregulator expression may be a general mechanism to integrate diverse hormone signals.     

Orphan nuclear receptor TR2, a mediator of preadipocyte proliferation, is differentially regulated by RA through exchange of coactivator PCAF with corepressor RIP140 on a platform molecule GRIP1

Orphan nuclear receptor TR2 is a preadipocyte proliferator. Knockdown of TR2 in 3T3-L1 preadipocytes reduced their proliferation efficiency, whereas specific elevation of TR2 in these cells facilitated their proliferation. All-trans retinoic acid (RA) stimulates cellular proliferation in 3T3-L1 preadipocytes by activating TR2 through an IR0-type RA response element, which further activates c-Myc expression. In post-differentiated adipocytes, RA becomes a repressive signal for TR2 and rapidly down-regulates its expression. The biphasic effect of RA on TR2 expression in 3T3-L1 is mediated by differential RA-dependent coregulator recruitment to the receptor/Glucocorticoid Receptor-Interacting Protein 1 (GRIP1) complex that binds IR0 on the TR2 promoter. RA induces the recruitment of histone acetyl transferase-containing/GRIP1/p300/CBP-associated factor (PCAF) complex to the TR2 promoter in undifferentiated cells, whereas it triggers recruitment of histone deacetylase-containing/GRIP1/receptor-interacting protein 140 (RIP140) complex in differentiated cells. GRIP1 directly interacts with RIP140 through its carboxyl terminal AD2 domain. GRIP1 interacts with PCAF and RIP140 directly and differentially, functioning as a platform molecule to mediate differential RA-induced coregulator recruitment to TR2 promoter target. This results in a biphasic effect of RA on the expression of TR2 in undifferentiated and differentiated cells, which is required for RA-stimulated preadipocyte proliferation.     

Cytoplasmic receptor-interacting protein 140 (RIP140) interacts with perilipin to regulate lipolysis

Receptor-interacting protein 140 (RIP140) is abundantly expressed in mature adipocyte and modulates gene expression involved in lipid and glucose metabolism. Protein kinase C epsilon and protein arginine methyltransferase 1 can sequentially stimulate RIP140 phosphorylation and then methylation, thereby promoting its export to the cytoplasm. Here we report a lipid signal triggering cytoplasmic accumulation of RIP140, and a new functional role for cytoplasmic RIP140 in adipocyte to regulate lipolysis. Increased lipid content, particularly an elevation in diacylglycerol levels, promotes RIP140 cytoplasmic accumulation and increased association with lipid droplets (LDs) by its direct interaction with perilipin. By interacting with RIP140, perilipin more efficiently recruits hormone-sensitive lipase (HSL) to LDs and enhances adipose triglyceride lipase (ATGL) forming complex with CGI-58, an activator of ATGL. Consequentially, HSL can more readily access its substrates, and ATGL is activated, ultimately enhancing lipolysis. In adipocytes, blocking cytoplasmic RIP140 accumulation reduces basal and isoproterenol-stimulated lipolysis and the pro-inflammatory potential of their conditioned media (i.e. activating NF-κB and inflammatory genes in macrophages). These results show that in adipocytes with high lipid contents, RIP140 increasingly accumulates in the cytoplasm and enhances triglyceride catabolism by directly interacting with perilipin. The study suggests that reducing nuclear export of RIP140 might be a useful means of controlling adipocyte lipolysis.