ERα promotes SUMO1 transcription by binding with the ERE and enhances SUMO1-mediated protein SUMOylation in breast cancer
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ABSTRACT:
SUBMITTER: Wang Q
PROVIDER: S-EPMC10506115 | biostudies-literature | 2023 Jul
REPOSITORIES: biostudies-literature
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Project description:Estrogen receptor α (ERα)-positive breast cancer patients are typically treated with ERα inhibitors, including selective estrogen receptor modulators (SERMs) and selective estrogen receptor degraders (SERDs). However, the distinct pharmacological properties of various ERα inhibitors remain incompletely understood. In this study, we employed formaldehyde cross-linking followed by ERα immunoprecipitation and mass spectrometry to reveal that fulvestrant, the first FDA-approved SERD, induces the interaction between ERα and SUMO E3 ligases PIAS1 and PIAS2. Biochemical and genomic assays confirmed that fulvestrant induces SUMOylation of ERα, which inhibits ERα's binding to chromatin DNA. In addition, raloxifene (a SERM) and elacestrant (the first FDA-approved oral SERD) were identified as compounds that similarly induce ERα SUMOylation and inhibit its chromatin interaction. Our findings reveal a mechanism by which select ERα inhibitors disrupt ERα function through SUMOylation, offering insights for the development of next-generation ERα-targeted therapies.
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Project description:Utilization of small molecules in modulation of stem cell self-renewal is a promising approach to expand stem cells for regenerative therapy. Here, we identify Icaritin, a phytoestrogen molecule enhances self-renewal of mouse embryonic stem cells (mESCs). Icaritin increases mESCs proliferation while maintains their self-renewal capacity in vitro and pluripotency in vivo. This coincides with upregulation of key pluripotency transcription factors OCT4, NANOG, KLF4 and SOX2. The enhancement of mESCs self-renewal is characterized by increased population in S-phase of cell cycle, elevation of Cylin E and Cyclin-dependent kinase 2 (CDK2) and downregulation of p21, p27 and p57. PCR array screening reveals that caudal-related homeobox 2 (Cdx2) and Rbl2/p130 are remarkably suppressed in mESCs treated with Icaritin. siRNA knockdown of Cdx2 or Rbl2/p130 upregulates the expression of Cyclin E, OCT4 and SOX2, and subsequently increases cell proliferation and colony forming efficiency of mESCs. We then demonstrate that Icaritin co-localizes with estrogen receptor alpha (ERα) and activates its nuclear translocation in mESCs. The promotive effect of Icaritin on cell cycle and pluripotency regulators are eliminated by siRNA knockdown of ERα in mESCs. The results suggest that Icaritin enhances mESCs self-renewal by regulating cell cycle machinery and core pluripotency transcription factors mediated by ERα.
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Project description:BackgroundChromothripsis is an event of genomic instability leading to complex chromosomal alterations in cancer. Frequent long-range chromatin interactions between transcription factors (TFs) and targets may promote extensive translocations and copy-number alterations in proximal contact regions through inappropriate DNA stitching. Although studies have proposed models to explain the initiation of chromothripsis, few discussed how TFs influence this process for tumor progression.MethodsThis study focused on genomic alterations in amplification associated regions within chromosome 17. Inter-/intra-chromosomal rearrangements were analyzed using whole genome sequencing data of breast tumors in the Cancer Genome Atlas (TCGA) cohort. Common ERα binding sites were defined based on MCF-7, T47D, and MDA-MB-134 breast cancer cell lines using univariate K-means clustering methods. Nanopore sequencing technology was applied to validate frequent rearrangements detected between ATC loci on 17q23 and an ERα hub on 20q13. The efficacy of pharmacological inhibition of a potentially druggable target gene on 17q23 was evaluated using breast cancer cell lines and patient-derived circulating breast tumor cells.ResultsThere are five adjoining regions from 17q11.1 to 17q24.1 being hotspots of chromothripsis. Inter-/intra-chromosomal rearrangements of these regions occurred more frequently in ERα-positive tumors than in ERα-negative tumors. In addition, the locations of the rearrangements were often mapped within or close to dense ERα binding sites localized on these five 17q regions or other chromosomes. This chromothriptic event was linked to concordant upregulation of 96 loci that predominantly regulate cell-cycle machineries in advanced luminal tumors. Genome-editing analysis confirmed that an ERα hub localized on 20q13 coordinately regulates a subset of these loci localized on 17q23 through long-range chromosome interactions. One of these loci, Tousled Like Kinase 2 (TLK2) known to participate in DNA damage checkpoint control, is an actionable target using phenothiazine antipsychotics (PTZs). The antiproliferative effect of PTZs was prominent in high TLK2-expressing cells, compared to low expressing cells.ConclusionThis study demonstrates a new approach for identifying tumorigenic drivers from genomic regions highly susceptible to ERα-related chromothripsis. We found a group of luminal breast tumors displaying 17q-related chromothripsis for which antipsychotics can be repurposed as treatment adjuncts.
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Project description:Post-translational modifications directly control protein activity and, thus, they represent an important means to regulate the responses of cells to different stimuli. Protein SUMOylation has recently been recognised as one such modification, and it has been associated with various diseases, including different types of cancer. However, the precise way that changes in SUMOylation influence the tumorigenic properties of cells remains to be fully clarified. Here, we show that blocking the SUMO pathway by depleting SUMO1 and UBC9, or by exposure to ginkgolic acid C15:1 or 2-D08 (two different SUMOylation inhibitors), induces cell death, also inhibiting the invasiveness of tumour cells. Indeed, diminishing the formation of SUMO1 complexes induces autophagy-mediated cancer cell death through increasing the expression of Tribbles pseudokinase 3 (TRIB3). Moreover, we found that blocking the SUMO pathway inhibits tumour cell invasion by decreasing RAC1 SUMOylation. These findings shed new light on the mechanisms by which SUMO1 modifications regulate the survival, and the migratory and invasive capacity of tumour cells, potentially establishing the bases to develop novel anti-cancer treatments based on the inhibition of SUMOylation.
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Project description:Upon binding to estrogens, the ERα nuclear receptor acts as a transcription factor and mediates a multitude of cellular functions central to health and disease. Herein, using isothermal titration calorimetry (ITC) and circular dichroism (CD) in conjunction with molecular modeling (MM), we analyze the effect of symmetric introduction of single nucleotide variations within each half-site of the estrogen response element (ERE) on the binding of ERα nuclear receptor. Our data reveal that ERα exudes remarkable tolerance and binds to all genetic variants in the physiologically relevant nanomolar-micromolar range with the consensus ERE motif affording the highest affinity. We provide rationale for how genetic variations within the ERE motif may reduce its affinity for ERα by orders of magnitude at atomic level. Our data also suggest that the introduction of genetic variations within the ERE motif allows it to sample a much greater conformational space. Surprisingly, ERα displays no preference for binding to ERE variants with higher AT content, implying that any advantage due to DNA plasticity may be largely compensated by unfavorable entropic factors. Collectively, our study bears important consequences for how genetic variations within DNA promoter elements may fine-tune the physiological action of ERα and other nuclear receptors.
| S-EPMC3044919 | biostudies-literature
Project description:Estrogen and estrogen receptor alpha (ERα)-induced gene transcription is tightly associated with ERα-positive breast carcinogenesis. ERα-occupied enhancers, particularly super-enhancers, have been suggested to play a vital role in regulating such transcriptional events. However, the landscape of ERα-occupied super-enhancers (ERSEs) as well as key ERα-induced target genes associated with ERSEs remain to be fully characterized. Here, we defined the landscape of ERSEs in ERα-positive breast cancer cell lines, and demonstrated that bromodomain protein BRD4 is a master regulator of the transcriptional activation of ERSEs and cognate ERα target genes. RET, a member of the tyrosine protein kinase family of proteins, was identified to be a key ERα target gene of BRD4-regulated ERSEs, which, in turn, is vital for ERα-induced gene transcriptional activation and malignant phenotypes through activating the RAS/RAF/MEK2/ERK/p90RSK/ERα phosphorylation cascade. Combination therapy with BRD4 and RET inhibitors exhibited additive effects on suppressing ERα-positive breast cancer both in vitro and in vivo, comparable with that of standard endocrine therapy tamoxifen. Furthermore, combination therapy re-sensitized a tamoxifen-resistant ERα-positive breast cancer cell line to tamoxifen treatment. Taken together, our data uncovered the critical role of a super-enhancer-associated positive feedback loop constituting BRD4/ERα-RET-ERα in ERα-positive breast cancer, and suggested that targeting components in this loop would provide a new therapeutic avenue for treating ERα-positive breast cancer in the clinic.
| S-EPMC9561272 | biostudies-literature
Project description:Tamoxifen is widely used as a medication for estrogen receptor α (ERα)-positive breast cancer, despite the ~50% incidence of tamoxifen resistance. To overcome such resistance, combining tamoxifen with other agents is considered an effective approach. Here, through in vitro studies with ER-positive MCF7 cells and ER-negative MDA-MB-231 cells, validated by the use of xenograft mice, we investigated the potential of tumor necrosis factor α (TNFα) to enhance tamoxifen sensitivity and identified NCOR1 as a key downstream regulator. TNFα specifically degraded nuclear receptor corepressor 1 (NCOR1) in MCF7 cells. Moreover, knockdown of NCOR1, similar to TNFα treatment, suppressed cancer cell growth and promoted apoptosis only in MCF7 cells and MCF7 xenograft mice through the stabilization of p53, a tumor suppressor protein. Interestingly, NCOR1 knockdown with TNFα treatment increased the occupancy of p53 at the p21 promoter, while decreasing that of ERα. Notably, NCOR1 formed a complex with p53 and ERα, which was disrupted by TNFα. Finally, combinatorial treatment with tamoxifen, TNFα and short-hairpin (sh)-NCOR1 resulted in enhanced suppression of tumor growth in MCF7 xenograft mice compared to single tamoxifen treatment. In conclusion, TNFα promoted tamoxifen sensitivity through the dissociation of the ERα-p53-NCOR1 complex, pointing at NCOR1 as a putative therapeutic target for overcoming tamoxifen resistance in ERα-positive breast cancer.
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Project description:Flavonoids are plant-derived compounds that occur abundantly in fruits and vegetables and have been shown to possess potent anti-cancer, antioxidant, and anti-inflammatory properties. However, their direct targets and molecular mechanism of action are not well characterized, hampering exploitation of the beneficial properties of flavonoids for drug development. Small ubiquitin-related modifier 1 (SUMO1) is attached to target proteins as part of a post-translational modification system implicated in a myriad of cellular processes from nuclear trafficking to transcriptional regulation. Using a combination of surface plasmon resonance, differential scanning fluorimetry and fluorescence quenching studies, we provide evidence for direct binding of the dietary flavonoid fisetin to human SUMO1. Our NMR chemical shift perturbation analyses reveal that binding to fisetin involves four conserved amino acid residues (L65, F66, E67, M82) previously shown to be important for conjugation of SUMO1 to target proteins. In vitro sumoylation experiments indicate that fisetin blocks sumoylation of tumor suppressor p53, consistent with fisetin negatively affecting post-translational modification and thus the biological activity of p53. A series of differential scanning fluorimetry experiments suggest that high concentrations of fisetin result in destabilization and unfolding of SUMO1, presenting a molecular mechanism by which flavonoid binding affects its activity. Overall, our data establish a novel direct interaction between fisetin and SUMO1, providing a mechanistic explanation for the ability of fisetin to modulate multiple key signaling pathways inside cells.
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Project description:Antiestrogens (AEs) are widely used for treatment of estrogen receptor alpha (ERα)-positive breast cancer, but display variable degrees of partial agonism in estrogen target tissues and breast cancer (BC) cells. The fact that BC cells resistant to selective ER modulators (SERMs) like tamoxifen (Tam) can still be sensitive to pure AEs, also called selective ER downregulators, suggests different mechanisms of action, some of which may contribute to the more complete suppression of estrogen target genes by pure AEs. We report herein that pure AEs such as fulvestrant induce transient binding of ERα to DNA, followed by rapid release after 30-40 min without loss of nuclear localization. Loss of DNA binding preceded receptor degradation and was not prevented by proteasome inhibition. Chromatin was less accessible in the presence of fulvestrant than with estradiol or Tam as early as 20 min following treatment, suggesting that chromatin remodeling by pure AEs at ERα target regions prevents transcription in spite of receptor binding. SUMO2/3 marks were detected on chromatin at the peak of ERα binding in cells treated with pure AEs, but not SERMs. Furthermore, decreasing SUMOylation by overexpressing the deSUMOylase SENP1 significantly delayed receptor release from DNA and de-repressed expression of estrogen target genes in the presence of fulvestrant, both in ERα-expressing MCF-7 cells and in transiently transfected ER-negative SK-BR-3 cells. Finally, mutation V534E, identified in a breast metastasis resistant to hormonal therapies, prevented ERα modification and resulted in increased transcriptional activity of estrogen target genes in the presence of fulvestrant in SK-BR-3 cells. Together, our results establish a role for SUMOylation in achieving a more complete transcriptional shut-off of estrogen target genes by pure AEs vs. SERMs in BC cells.
| S-EPMC6514857 | biostudies-literature
Project description:Two opposing clusters of transcription factors (TF) have been associated with the differential risks of estrogen receptor positive or negative breast cancers, but the mechanisms underlying the opposing functions of the two clusters are undefined. In this study, we identified NFIB and YBX1 as novel interactors of the estrogen receptor (ESR1). NFIB and YBX1 are both risk TF associated with progression of ESR1-negative disease. Notably, they both interacted with the ESR1-FOXA1 complex and inhibited the transactivational potential of ESR1. Moreover, signaling through FGFR2, a known risk factor in breast cancer development, augmented these interactions and further repressed ESR1 target gene expression. We therefore show that members of two opposing clusters of risk TFs associated with ESR1-positive and -negative breast cancer can physically interact. We postulate that this interaction forms a toggle between two developmental pathways affected by FGFR2 signaling, possibly offering a junction to exploit therapeutically.Significance: Binding of the transcription factors NFIB and YBX1 to the estrogen receptor can promote an estrogen-independent phenotype that can be reverted by inhibiting FGFR2 signaling. Cancer Res; 78(2); 410-21. ©2017 AACR.
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