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Metastasis-associated protein 1 interacts with NRIF3, an estrogen-inducible nuclear receptor coregulator.

ABSTRACT: The transcriptional activity of estrogen receptor alpha (ER-alpha) is modified by regulatory action and interactions of coactivators and corepressors. Recent studies have shown that the metastasis-associated protein 1 (MTA1) represses estrogen receptor element (ERE)-driven transcription in breast cancer cells. With a yeast two-hybrid screen to clone MTA1-interacting proteins, we identified a known nuclear receptor coregulator (NRIF3) as an MTA1-binding protein. NRIF3 interacted with MTA1 both in vitro and in vivo. NRIF3 bound to the C-terminal region of MTA1, while MTA1 bound to the N-terminal region of NRIF3, containing one nuclear receptor interaction LXXLL motif. We showed that NRIF3 is an ER coactivator, hyperstimulated ER transactivation functions, and associated with the endogenous ER and its target gene promoter. MTA1 repressed NRIF3-mediated stimulation of ERE-driven transcription and interfered with NRIF3's association with the ER target gene chromatin. In addition, NRIF3 deregulation enhanced the responsiveness of breast cancer cells to estrogen-induced stimulation of growth and anchorage independence. Furthermore, we found that NRIF3 is an estrogen-inducible gene and activated ER associated with the ER response element in the NRIF3 gene promoter. These findings suggest that NRIF3, an MTA1-interacting protein, is an estrogen-inducible gene and that regulatory interactions between MTA1 and NRIF3 might be important in modulating the sensitivity of breast cancer cells to estrogen.

SUBMITTER: Talukder AH

PROVIDER: S-EPMC444867 | biostudies-literature | 2004 Aug

REPOSITORIES: biostudies-literature

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Metastasis-associated protein 1 interacts with NRIF3, an estrogen-inducible nuclear receptor coregulator.

Talukder Amjad H AH Gururaj Anupama A Mishra Sandip K SK Vadlamudi Ratna K RK Kumar Rakesh R

Molecular and cellular biology 20040801 15

The transcriptional activity of estrogen receptor alpha (ER-alpha) is modified by regulatory action and interactions of coactivators and corepressors. Recent studies have shown that the metastasis-associated protein 1 (MTA1) represses estrogen receptor element (ERE)-driven transcription in breast cancer cells. With a yeast two-hybrid screen to clone MTA1-interacting proteins, we identified a known nuclear receptor coregulator (NRIF3) as an MTA1-binding protein. NRIF3 interacted with MTA1 both in ...[more]

PMID: 15254226

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Project description:Although molecular signatures based on transcript expression in breast cancer samples have provided new insights into breast cancer classification and prognosis, there are acknowledged limitations in current signatures. To provide rational, pathway-based signatures of disrupted physiology in cancer tissues that may be relevant to prognosis, this study has directly quantitated changed gene expression, between normal breast and cancer tissue, as a basis for signature development. The nuclear receptor (NR) family of transcription factors, and their coregulators, are fundamental regulators of every aspect of metazoan life, and were rigorously quantified in normal breast tissues and ER? positive and ER? negative breast cancers. Coregulator expression was highly correlated with that of selected NR in normal breast, particularly from postmenopausal women. These associations were markedly decreased in breast cancer, and the expression of the majority of coregulators was down-regulated in cancer tissues compared with normal. While in cancer the loss of NR-coregulator associations observed in normal breast was common, a small number of NR (Rev-ERB?, GR, NOR1, LRH-1 and PGR) acquired new associations with coregulators in cancer tissues. Elevated expression of these NR in cancers was associated with poorer outcome in large clinical cohorts, as well as suggesting the activation of ER? -related, but ER?-independent, pathways in ER? negative cancers. In addition, the combined expression of small numbers of NR and coregulators in breast cancer was identified as a signature predicting outcome in ER? negative breast cancer patients, not linked to proliferation and with predictive power superior to existing signatures containing many more genes. These findings highlight the power of predictive signatures derived from the quantitative determination of altered gene expression between normal breast and breast cancers. Taken together, the findings of this study identify networks of NR-coregulator associations active in normal breast but disrupted in breast cancer, and moreover provide evidence that signatures based on NR networks disrupted in cancer can provide important prognostic information in breast cancer patients.

Project description:BACKGROUND:CDK4/6 inhibitors in combination with endocrine therapy (AE/AI/SERDs) are approved for the treatment of ER+ advanced breast cancer (BCa). However, not all patients benefit from CDK4/6 inhibitors therapy. We previously reported a novel therapeutic agent, ERX-11, that binds to the estrogen receptor (ER) and modulates ER-coregulator interactions. Here, we tested if the combination of ERX-11 with agents approved for ER+ BCa would be more potent. METHODS:We tested the effect of combination therapy using BCa cell line models, including those that have acquired resistance to tamoxifen, letrozole, or CDK4/6 inhibitors or have been engineered to express mutant forms of the ER. In vitro activity was tested using Cell Titer-Glo, MTT, and apoptosis assays. Mechanistic studies were conducted using western blot, reporter gene assays, RT-qPCR, and mass spectrometry approaches. Xenograft, patient-derived explants (PDEs), and xenograft-derived explants (XDE) were used for preclinical evaluation and toxicity. RESULTS:ERX-11 inhibited the proliferation of therapy-resistant BCa cells in a dose-dependent manner, including ribociclib resistance. The combination of ERX-11 and CDK4/6 inhibitor was synergistic in decreasing the proliferation of both endocrine therapy-sensitive and endocrine therapy-resistant BCa cells, in vitro, in xenograft models in vivo, xenograft-derived explants ex vivo, and in primary patient-derived explants ex vivo. Importantly, the combination caused xenograft tumor regression in vivo. Unbiased global mass spectrometry studies demonstrated profound decreases in proliferation markers with combination therapy and indicated global proteomic changes in E2F1, ER, and ER coregulators. Mechanistically, the combination of ERX-11 and CDK4/6 inhibitor decreased the interaction between ER and its coregulators, as evidenced by immunoprecipitation followed by mass spectrometry studies. Biochemical studies confirmed that the combination therapy significantly altered the expression of proteins involved in E2F1 and ER signaling, and this is primarily driven by a transcriptional shift, as noted in gene expression studies. CONCLUSIONS:Our results suggest that ERX-11 inhibited the proliferation of BCa cells resistant to both endocrine therapy and CDK4/6 inhibitors in a dose-dependent manner and that the combination of ERX-11 with a CDK4/6 inhibitor may represent a viable therapeutic approach.

Project description:NCoA6 (also referred to as NRC, ASC-2, TRBP, PRIP and RAP250) was originally isolated as a ligand-dependent nuclear receptor interacting protein. However, NCoA6 is a multifunctional coregulator or coactivator necessary for transcriptional activation of a wide spectrum of target genes. The NCoA6 gene is amplified and overexpressed in breast, colon and lung cancers. NCoA6 is a 250 kDa protein which harbors a potent N-terminal activation domain, AD1; and a second, centrally-located activation domain, AD2, which is necessary for nuclear receptor signaling. The intrinsic activation potential of NCoA6 is regulated by its C-terminal STL regulatory domain. Near AD2 is an LxxLL-1 motif which interacts with a wide spectrum of ligand-bound NRs with high-affinity. A second LxxLL motif (LxxLL-2) located towards the C-terminal region is more restricted in its NR specificity. The potential role of NCoA6 as a co-integrator is suggested by its ability to enhance transcriptional activation of a wide variety of transcription factors and from its in vivo association with a number of known cofactors including CBP/p300. NCoA6 has been shown to associate with at least three distinct coactivator complexes containing Set methyltransferases as core polypeptides. The composition of these complexes suggests that NCoA6 may play a fundamental role in transcriptional activation by modulating chromatin structure through histone methylation. Knockout studies in mice suggest that NCoA6 is an essential coactivator. NCoA6-/- embryos die between 8.5-12.5 dpc from general growth retardation coupled with developmental defects in the heart, liver, brain and placenta. NCoA6-/- MEFs grow at a reduced rate compared to WT MEFs and spontaneously undergo apoptosis, indicating the importance of NCoA6 as a prosurvival and anti-apoptotic gene. Studies with NCoA6+/- and conditional knockout mice suggest that NCoA6 is a pleiotropic coregulator involved in growth, development, wound healing and maintenance of energy homeostasis.

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Metastasis-associated protein 1 interacts with NRIF3, an estrogen-inducible nuclear receptor coregulator.

Publications

Metastasis-associated protein 1 interacts with NRIF3, an estrogen-inducible nuclear receptor coregulator.

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