Akt SUMOylation regulates cell proliferation and tumorigenesis.

PubMed

Li, Rong; Wei, Jie; Jiang, Cong; Liu, Dongmei; Deng, Lu; Zhang, Kai; Wang, Ping

2013-09-15

Proto-oncogene Akt plays essential roles in cell proliferation and tumorigenesis. Full activation of Akt is regulated by phosphorylation, ubiquitination, and acetylation. Here we report that SUMOylation of Akt is a novel mechanism for its activation. Systematically analyzing the role of lysine residues in Akt activation revealed that K276, which is located in a SUMOylation consensus motif, is essential for Akt activation. Ectopic or endogenous Akt1 could be modified by SUMOylation. RNA interference-mediated silencing of UBC9 reduced Akt SUMOylation, which was promoted by SUMO E3 ligase PIAS1 and reversed by the SUMO-specific protease SENP1. Although multiple sites on Akt could be SUMOylated, K276 was identified as a major SUMO acceptor site. K276R or E278A mutation reduced SUMOylation of Akt but had little effect on its ubiquitination. Strikingly, these mutations also completely abolished Akt kinase activity. In support of these results, we found that expression of PIAS1 and SUMO1 increased Akt activity, whereas expression of SENP1 reduced Akt1 activity. Interestingly, the cancer-derived mutant E17K in Akt1 that occurs in various cancers was more efficiently SUMOylated than wild-type Akt. Moreover, SUMOylation loss dramatically reduced Akt1 E17K-mediated cell proliferation, cell migration, and tumorigenesis. Collectively, our findings establish that Akt SUMOylation provides a novel regulatory mechanism for activating Akt function. ©2013 AACR.

Sumoylation of Sir2 differentially regulates transcriptional silencing in yeast.

PubMed

Hannan, Abdul; Abraham, Neethu Maria; Goyal, Siddharth; Jamir, Imlitoshi; Priyakumar, U Deva; Mishra, Krishnaveni

2015-12-02

Silent information regulator 2 (Sir2), the founding member of the conserved sirtuin family of NAD(+)-dependent histone deacetylase, regulates several physiological processes including genome stability, gene silencing, metabolism and life span in yeast. Within the nucleus, Sir2 is associated with telomere clusters in the nuclear periphery and rDNA in the nucleolus and regulates gene silencing at these genomic sites. How distribution of Sir2 between telomere and rDNA is regulated is not known. Here we show that Sir2 is sumoylated and this modification modulates the intra-nuclear distribution of Sir2. We identify Siz2 as the key SUMO ligase and show that multiple lysines in Sir2 are subject to this sumoylation activity. Mutating K215 alone counteracts the inhibitory effect of Siz2 on telomeric silencing. SUMO modification of Sir2 impairs interaction with Sir4 but not Net1 and, furthermore, SUMO modified Sir2 shows predominant nucleolar localization. Our findings demonstrate that sumoylation of Sir2 modulates distribution between telomeres and rDNA and this is likely to have implications for Sir2 function in other loci as well. © The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.

Sumoylation Dynamics During Keratinocyte Differentiation

PubMed Central

Deyrieux, Adeline F.; Rosas-Acosta, Germán; Ozbun, Michelle A.; Wilson, Van G.

2012-01-01

Summary SUMO modification regulates the activity of numerous transcription factors that have a direct role in cell cycle progression, apoptosis, cellular proliferation, and development, but its role in differentiation processes is less clear. Keratinocyte differentiation requires the coordinated activation of a series of transcription factors, and as several critical keratinocyte transcription factors are known to be SUMO substrates, we investigated the role of sumoylation in keratinocyte differentiation. In a human keratinocyte cell line model (HaCaT cells), calcium-induced differentiation led to the transient and coordinated transcriptional activation of the genes encoding critical sumoylation system components, including SAE1, SAE2, Ubc9, SENP1, Miz-1 (PIASxβ), SUMO2, and SUMO3. The increased gene expression resulted in higher levels of the respective proteins and changes in the pattern of sumoylated substrate proteins during the differentiation process. Similar to the HaCaT results, stratified human foreskin keratinocytes showed an upregulation of Ubc9 in the suprabasal layers. Lastly, abrogation of sumoylation by Gam1 expression severely disrupted normal HaCaT differentiation, consistent with an important role for sumoylation in the proper progression of this biological process. PMID:17164289

Sumoylation Modulates the Activity of Spalt-like Proteins during Wing Development in Drosophila*

PubMed Central

Sánchez, Jonatan; Talamillo, Ana; Lopitz-Otsoa, Fernando; Pérez, Coralia; Hjerpe, Roland; Sutherland, James D.; Herboso, Leire; Rodríguez, Manuel S.; Barrio, Rosa

2010-01-01

The Spalt-like family of zinc finger transcription factors is conserved throughout evolution and is involved in fundamental processes during development and during embryonic stem cell maintenance. Although human SALL1 is modified by SUMO-1 in vitro, it is not known whether this post-translational modification plays a role in regulating the activity of this family of transcription factors. Here, we show that the Drosophila Spalt transcription factors are modified by sumoylation. This modification influences their nuclear localization and capacity to induce vein formation through the regulation of target genes during wing development. Furthermore, spalt genes interact genetically with the sumoylation machinery to repress vein formation in intervein regions and to attain the wing final size. Our results suggest a new level of regulation of Sall activity in vivo during animal development through post-translational modification by sumoylation. The evolutionary conservation of this family of transcription factors suggests a functional role for sumoylation in vertebrate Sall members. PMID:20562097

Sumoylation in p27kip1 via RanBP2 promotes cancer cell growth in cholangiocarcinoma cell line QBC939.

PubMed

Yang, Jun; Liu, Yan; Wang, Bing; Lan, Hongzhen; Liu, Ying; Chen, Fei; Zhang, Ju; Luo, Jian

2017-09-07

Cholangiocarcinoma is one of the deadly disease with poor 5-year survival and poor response to conventional therapies. Previously, we found that p27kip1 nuclear-cytoplasmic translocation confers proliferation potential to cholangiocarcinoma cell line QBC939 and this process is mediated by crm-1. However, no other post-transcriptional regulation was found in this process including sumoylation in cholangiocarcinoma. In this study, we explored the role of sumoylation in the nuclear-cytoplasmic translocation of p27kip1 and its involvement of QBC939 cells' proliferation. First, we identified K73 as the sumoylation site in p27kip1. By utilizing plasmid flag-p27kip1, HA-RanBP2, GST-RanBP2 and His-p27kip1 and immunoprecipitation assay, we validated that p27kip1 can serve as the sumoylation target of RanBP2 in QBC939. Furthermore, we confirmed crm-1's role in promoting nuclear-cytoplasmic translocation of p27kip1 and found that RanBP2's function relies on crm-1. However, K73R mutated p27kip1 can't be identified by crm-1 or RanBP2 in p27kip1 translocation process, suggesting sumoylation of p27kip1 via K73 site is necessary in this process by RanBP2 and crm-1. Phenotypically, the overexpression of either RanBP2 or crm-1 can partially rescue the anti-proliferative effect brought by p27kip1 overexpression in both the MTS and EdU assay. For the first time, we identified and validated the K73 sumoylation site in p27kip1, which is critical to RanBP2 and crm-1 in p27kip1 nuclear-cytoplasmic translocation process. Taken together, targeted inhibition of sumoylation of p27kip1 may serve as a potentially potent therapeutic target in the eradication of cholangiocarcinoma development and relapses.

Solubility shift and SUMOylaltion of promyelocytic leukemia (PML) protein in response to arsenic(III) and fate of the SUMOylated PML

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hirano, Seishiro, E-mail: seishiro@nies.go.jp; Graduate School of Pharmaceutical Sciences, Chiba University; Tadano, Mihoko

2015-09-15

Promyelocytic leukemia (PML), which is a tumor suppressor protein that nevertheless plays an important role in the maintenance of leukemia initiating cells, is known to be biochemically modified by As{sup 3+}. We recently developed a simple method to evaluate the modification of PML by As{sup 3+} resulting in a change in solubility and the covalent binding of small ubiquitin-like modifier (SUMO). Here we semi-quantitatively investigated the SUMOylation of PML using HEK293 cells which were stably transfected with PML-VI (HEK-PML). Western blot analyses indicated that PML became insoluble in cold RadioImmunoPrecipitation Assay (RIPA) lysis buffer and was SUMOylated by both SUMO2/3more » and SUMO1 by As{sup 3+}. Surprisingly SUMO1 monomers were completely utilized for the SUMOylation of PML. Antimony (Sb{sup 3+}) but not bismuth (Bi{sup 3+}), Cu{sup 2+}, or Cd{sup 2+} biochemically modified PML similarly. SUMOylated PML decreased after removal of As{sup 3+} from the culture medium. However, unSUMOylated PML was still recovered in the RIPA-insoluble fraction, suggesting that SUMOylation is not requisite for changing the RIPA-soluble PML into the RIPA-insoluble form. Immunofluorescence staining of As{sup 3+}-exposed cells indicated that SUMO2/3 was co-localized with PML in the nuclear bodies. However, some PML protein was present in peri-nuclear regions without SUMO2/3. Functional Really Interesting New Gene (RING)-deleted mutant PML neither formed PML nuclear bodies nor was biochemically modified by As{sup 3+}. Conjugation with intracellular glutathione may explain the accessibility of As{sup 3+} and Sb{sup 3+} to PML in the nuclear region evading chelation and entrapping by cytoplasmic proteins such as metallothioneins. - Highlights: • As{sup 3+} is a carcinogen and also a therapeutic agent for leukemia. • PML becomes insoluble in RIPA and SUMOylated by As{sup 3+}. • Sb{sup 3+} modifies PML similar to As{sup 3+}. • Functional RING motif is necessary for As

Sumoylation of the Tumor Suppressor Promyelocytic Leukemia Protein Regulates Arsenic Trioxide-Induced Collagen Synthesis in Osteoblasts.

PubMed

Xu, Wen-Xiao; Liu, Sheng-Zhi; Wu, Di; Qiao, Guo-Fen; Yan, Jinglong

2015-01-01

Promyelocytic leukemia (PML) protein is a tumor suppressor that fuses with retinoic acid receptor-α (PML-RARα) to contribute to the initiation of acute promyelocytic leukemia (APL). Arsenic trioxide (ATO) upregulates expression of TGF-β1, promoting collagen synthesis in osteoblasts, and ATO binds directly to PML to induce oligomerization, sumoylation, and ubiquitination. However, how ATO upregulates TGF-β1 expression is uncertain. Thus, we suggested that PML sumoylation is responsible for regulation of TGF-β1 protein expression. Kunming mice were treated with ATO, and osteoblasts were counted under scanning electron microscopy. Masson's staining was used to quantify collagen content. hFOB1.19 cells were transfected with siRNA against UBC9 or RNF4, and then treated with ATO or FBS. TGF-β1, PML expression, and sumoylation were quantified with Western blot, and collagen quantified via immunocytochemistry. ATO enhanced osteoblast accumulation, collagen synthesis, and PML-NB formation in vivo. Knocking down UBC9 in hFOB1.19 cells inhibited ATO- and FBS-induced PML sumoylation, TGF-β1 expression, and collagen synthesis. Conversely, knocking down RNF4 enhanced ATO- and FBS-induced PML sumoylation, TGF-β1 expression, and collagen synthesis. These data suggest that PML sumoylation is required for ATO-induced collagen synthesis in osteoblasts. © 2015 S. Karger AG, Basel.

Preparation of sumoylated substrates for biochemical analysis.

PubMed

Knipscheer, Puck; Klug, Helene; Sixma, Titia K; Pichler, Andrea

2009-01-01

Covalent modification of proteins with SUMO (small ubiquitin related modifier) affects many cellular processes like transcription, nuclear transport, DNA repair and cell cycle progression. Although hundreds of SUMO targets have been identified, for several of them the function remains obscure. In the majority of cases sumoylation is investigated via "loss of modification" analysis by mutating the relevant target lysine. However, in other cases this approach is not successful since mapping of the modification site is problematic or mutation does not cause an obvious phenotype. These latter cases ask for different approaches to investigate the target modification. One possibility is to choose the opposite approach, a "gain in modification" analysis by producing both SUMO modified and unmodified protein in vitro and comparing them in functional assays. Here, we describe the purification of the ubiquitin conjugating enzyme E2-25K, its in vitro sumoylation with recombinant enzymes and the subsequent separation and purification of the modified and the unmodified forms.

SUMOylation-disrupting WAS mutation converts WASp from a transcriptional activator to a repressor of NF-κB response genes in T cells.

PubMed

Sarkar, Koustav; Sadhukhan, Sanjoy; Han, Seong-Su; Vyas, Yatin M

2015-10-01

In Wiskott-Aldrich syndrome (WAS), immunodeficiency and autoimmunity often comanifest, yet how WAS mutations misregulate chromatin-signaling in Thelper (TH) cells favoring development of auto-inflammation over protective immunity is unclear. Previously, we identified an essential promoter-specific, coactivator role of nuclear-WASp in TH1 gene transcription. Here we identify small ubiquitin-related modifier (SUMO)ylation as a novel posttranslational modification of WASp, impairment of which converts nuclear-WASp from a transcriptional coactivator to a corepressor of nuclear factor (NF)-κB response genes in human (TH)1-differentiating cells. V75M, one of many disease-causing mutations occurring in SUMO*motif (72-ψψψψKDxxxxSY-83) of WASp, compromises WASp-SUMOylation, associates with COMMD1 to attenuate NF-κB signaling, and recruits histone deacetylases-6 (HDAC6) to p300-marked promoters of NF-κB response genes that pattern immunity but not inflammation. Consequently, proteins mediating adaptive immunity (IFNG, STAT1, TLR1) are deficient, whereas those mediating auto-inflammation (GM-CSF, TNFAIP2, IL-1β) are paradoxically increased in TH1 cells expressing SUMOylation-deficient WASp. Moreover, SUMOylation-deficient WASp favors ectopic development of the TH17-like phenotype (↑IL17A, IL21, IL22, IL23R, RORC, and CSF2) under TH1-skewing conditions, suggesting a role for WASp in modulating TH1/TH17 plasticity. Notably, pan-histone deacetylase inhibitors lift promoter-specific repression imposed by SUMOylation-deficient WASp and restore misregulated gene expression. Our findings uncovering a SUMOylation-based mechanism controlling WASp's dichotomous roles in transcription may have implications for personalized therapy for patients carrying mutations that perturb WASp-SUMOylation. © 2015 by The American Society of Hematology.

Production and Purification of Recombinant SUMOylated Proteins Using Engineered Bacteria.

PubMed

Brockly, Frédérique; Piechaczyk, Marc; Bossis, Guillaume

2016-01-01

SUMO is a ubiquitin-like protein that is covalently conjugated to numerous cellular proteins to modify their function and fate. Although large progresses have been made in the identification of SUMOylated proteins, the molecular consequences of their SUMOylation are generally unknown. This is, most often, due to the low abundance of SUMOylated proteins in the cell, usually less than 1 % of a given protein being modified at steady state. To gain insights into the role of specific SUMOylation targets, SUMO conjugation can be reconstituted in vitro using purified proteins. However, for most substrates, the efficiency of in vitro SUMOylation is too low to obtain sufficient amounts of their SUMOylated forms for biochemical studies. Here, we describe a detailed protocol to purify large amounts of recombinant SUMOylated proteins using bacteria modified to express His-tagged SUMO as well as the SUMO-activating and -conjugating enzymes.

Expression, Localization of SUMO-1, and Analyses of Potential SUMOylated Proteins in Bubalus bubalis Spermatozoa

PubMed Central

Brohi, Rahim Dad; Wang, Li; Hassine, Najla Ben; Cao, Jing; Talpur, Hira Sajjad; Wu, Di; Huang, Chun-Jie; Rehman, Zia-Ur; Bhattarai, Dinesh; Huo, Li-Jun

2017-01-01

Mature spermatozoa have highly condensed DNA that is essentially silent both transcriptionally and translationally. Therefore, post translational modifications are very important for regulating sperm motility, morphology, and for male fertility in general. Protein sumoylation was recently demonstrated in human and rodent spermatozoa, with potential consequences for sperm motility and DNA integrity. We examined the expression and localization of small ubiquitin-related modifier-1 (SUMO-1) in the sperm of water buffalo (Bubalus bubalis) using immunofluorescence analysis. We confirmed the expression of SUMO-1 in the acrosome. We further found that SUMO-1 was lost if the acrosome reaction was induced by calcium ionophore A23187. Proteins modified or conjugated by SUMO-1 in water buffalo sperm were pulled down and analyzed by mass spectrometry. Sixty proteins were identified, including proteins important for sperm morphology and motility, such as relaxin receptors and cytoskeletal proteins, including tubulin chains, actins, and dyneins. Forty-six proteins were predicted as potential sumoylation targets. The expression of SUMO-1 in the acrosome region of water buffalo sperm and the identification of potentially SUMOylated proteins important for sperm function implicates sumoylation as a crucial PTM related to sperm function. PMID:28659810

SUMOylation of the KRAB zinc-finger transcription factor PARIS/ZNF746 regulates its transcriptional activity.

PubMed

Nishida, Tamotsu; Yamada, Yoshiji

2016-05-13

Parkin-interacting substrate (PARIS), a member of the family of Krüppel-associated box (KRAB)-containing zinc-finger transcription factors, is a substrate of the ubiquitin E3 ligase parkin. PARIS represses the expression of peroxisome proliferator-activated receptor γ coactivator-1α (PGC-1α), although the underlying mechanisms remain largely unknown. In the present study, we demonstrate that PARIS can be SUMOylated, and its SUMOylation plays a role in the repression of PGC-1a promoter activity. Protein inhibitor of activated STAT y (PIASy) was identified as an interacting protein of PARIS and shown to enhance its SUMOylation. PIASy repressed PGC-1a promoter activity, and this effect was attenuated by PARIS in a manner dependent on its SUMOylation status. Co-expression of SUMO-1 with PIASy completely repressed PGC-1a promoter activity independently of PARIS expression. PARIS-mediated PGC-1a promoter repression depended on the activity of histone deacetylases (HDAC), whereas PIASy repressed the PGC-1a promoter in an HDAC-independent manner. Taken together, these results suggest that PARIS and PIASy modulate PGC-1a gene transcription through distinct molecular mechanisms. Copyright © 2016 Elsevier Inc. All rights reserved.

Inhibition of HSF2 SUMOylation via MEL18 upregulates IGF-IIR and leads to hypertension-induced cardiac hypertrophy.

PubMed

Huang, Chih-Yang; Kuo, Chia-Hua; Pai, Pei-Ying; Ho, Tsung-Jung; Lin, Yueh-Min; Chen, Ray-Jade; Tsai, Fuu-Jen; Vijaya Padma, V; Kuo, Wei-Wen; Huang, Chih-Yang

2018-04-15

Cardiac hypertrophy is a major characteristic of early-stage hypertension-related heart failure. We have found that the insulin-like growth factor receptor II (IGF-IIR) signaling was critical for hypertensive angiotensin II-induced cardiomyocyte hypertrophy and apoptosis. Moreover, this IGF-IIR signaling was elegantly modulated by the heat shock transcription factors (HSFs) during heart failure. However, the detailed mechanism by which HSFs regulates IGF-IIR during hypertension-induced cardiac hypertrophy remains elusive. In this study, we found that heat shock transcription factor 2 (HSF2) activated IGF-IIR to induce cardiac hypertrophy for hypertension-induced heart failure. The transcriptional activity of HSF2 appeared to be primarily mediated by SUMOylation via conjugation with small ubiquitin-like modifier-1 (SUMO-1). The SUMOylation of HSF2 was severely attenuated by MEL18 (also known as polycomb group ring finger 2 or PCGF2) in the heart of spontaneously hypertensive rats (SHR). Inhibition of HSF2 SUMOylation severely induced cardiac hypertrophy via IGF-IIR-mediated signaling in hypertensive rats. Angiotensin II receptor type I blocker (ARB) treatment in spontaneously hypertensive rats restored HSF2 SUMOylation and alleviated the cardiac defects. Thus, our study uncovered a novel MEL18-SUMO-1-HSF2-IGF-IIR pathway in the heart that profoundly influences cardiac hypertrophy for hypertension-induced heart failure. Copyright © 2017 Elsevier B.V. All rights reserved.

A Novel Proteomics Approach to Identify SUMOylated Proteins and Their Modification Sites in Human Cells*

PubMed Central

Galisson, Frederic; Mahrouche, Louiza; Courcelles, Mathieu; Bonneil, Eric; Meloche, Sylvain; Chelbi-Alix, Mounira K.; Thibault, Pierre

2011-01-01

The small ubiquitin-related modifier (SUMO) is a small group of proteins that are reversibly attached to protein substrates to modify their functions. The large scale identification of protein SUMOylation and their modification sites in mammalian cells represents a significant challenge because of the relatively small number of in vivo substrates and the dynamic nature of this modification. We report here a novel proteomics approach to selectively enrich and identify SUMO conjugates from human cells. We stably expressed different SUMO paralogs in HEK293 cells, each containing a His6 tag and a strategically located tryptic cleavage site at the C terminus to facilitate the recovery and identification of SUMOylated peptides by affinity enrichment and mass spectrometry. Tryptic peptides with short SUMO remnants offer significant advantages in large scale SUMOylome experiments including the generation of paralog-specific fragment ions following CID and ETD activation, and the identification of modified peptides using conventional database search engines such as Mascot. We identified 205 unique protein substrates together with 17 precise SUMOylation sites present in 12 SUMO protein conjugates including three new sites (Lys-380, Lys-400, and Lys-497) on the protein promyelocytic leukemia. Label-free quantitative proteomics analyses on purified nuclear extracts from untreated and arsenic trioxide-treated cells revealed that all identified SUMOylated sites of promyelocytic leukemia were differentially SUMOylated upon stimulation. PMID:21098080

Glucocorticoid-induced tethered transrepression requires SUMOylation of GR and formation of a SUMO-SMRT/NCoR1-HDAC3 repressing complex

PubMed Central

Hua, Guoqiang; Ganti, Krishna Priya; Chambon, Pierre

2016-01-01

Upon binding of a glucocorticoid (GC), the GC receptor (GR) can exert one of three transcriptional regulatory functions. We recently reported that SUMOylation of the GR at position K293 in humans (K310 in mice) within the N-terminal domain is indispensable for GC-induced evolutionary conserved inverted repeated negative GC response element (IR nGRE)-mediated direct transrepression. We now demonstrate that the integrity of this GR SUMOylation site is mandatory for the formation of a GR-small ubiquitin-related modifiers (SUMOs)-SMRT/NCoR1-HDAC3 repressing complex, which is indispensable for NF-κB/AP1-mediated GC-induced tethered indirect transrepression in vitro. Using GR K310R mutant mice or mice containing the N-terminal truncated GR isoform GRα-D3 lacking the K310 SUMOylation site, revealed a more severe skin inflammation than in WT mice. Importantly, cotreatment with dexamethasone (Dex) could not efficiently suppress a 12-O-tetradecanoylphorbol-13-acetate (TPA)–induced skin inflammation in these mutant mice, whereas it was clearly decreased in WT mice. In addition, in mice selectively ablated in skin keratinocytes for either nuclear receptor corepressor 1 (NCoR1)/silencing mediator for retinoid or thyroid-hormone receptors (SMRT) corepressors or histone deacetylase 3 (HDAC3), Dex-induced tethered transrepression and the formation of a repressing complex on DNA-bound NF-κB/AP1 were impaired. We previously suggested that GR ligands that would lack both (+)GRE-mediated transactivation and IR nGRE-mediated direct transrepression activities of GCs may preferentially exert the therapeutically beneficial GC antiinflammatory properties. Interestingly, we now identified a nonsteroidal antiinflammatory selective GR agonist (SEGRA) that selectively lacks both Dex-induced (+)GRE-mediated transactivation and IR nGRE-mediated direct transrepression functions, while still exerting a tethered indirect transrepression activity and could therefore be clinically lesser

Epstein-Barr Virus Latent Membrane Protein 1 Regulates the Function of Interferon Regulatory Factor 7 by Inducing Its Sumoylation

PubMed Central

Bentz, Gretchen L.; Shackelford, Julia

2012-01-01

Epstein-Barr virus (EBV) latent membrane protein 1 (LMP1) induces multiple signal transduction pathways during latent EBV infection via its C-terminal activating region 1 (CTAR1), CTAR2, and the less-studied CTAR3. One mechanism by which LMP1 regulates cellular activation is through the induction of protein posttranslational modifications, including phosphorylation and ubiquitination. We recently documented that LMP1 induces a third major protein modification by physically interacting with the SUMO-conjugating enzyme Ubc9 through CTAR3 and inducing the sumoylation of cellular proteins in latently infected cells. We have now identified a specific target of LMP1-induced sumoylation, interferon regulatory factor 7 (IRF7). We hypothesize that during EBV latency, LMP1 induces the sumoylation of IRF7, limiting its transcriptional activity and modulating the activation of innate immune responses. Our data show that endogenously sumoylated IRF7 is detected in latently infected EBV lymphoblastoid cell lines. LMP1 expression coincided with increased sumoylation of IRF7 in a CTAR3-dependent manner. Additional experiments show that LMP1 CTAR3-induced sumoylation regulates the expression and function of IRF7 by decreasing its turnover, increasing its nuclear retention, decreasing its DNA binding, and limiting its transcriptional activation. Finally, we identified that IRF7 is sumoylated at lysine 452. These data demonstrate that LMP1 CTAR3 does in fact function in intracellular signaling, leading to biologic effects. We propose that CTAR3 is an important signaling region of LMP1 that regulates protein function by sumoylation. We have shown specifically that LMP1 CTAR3, in cooperation with CTAR2, can limit the ability of IRF7 to induce innate immune responses by inducing the sumoylation of IRF7. PMID:22951831

SUMOylated MAFB promotes colorectal cancer tumorigenesis

PubMed Central

Xie, Yin-Yin; Sun, Xiao-Jian; Zhao, Ren; Huang, Qiu-Hua

2016-01-01

The transcription factor, v-maf avian musculoaponeurotic fibrosarcoma oncogene homolog B (MAFB), promotes tumorigenesis in some cancers. In this study, we found that MAFB levels were increased in clinical colorectal cancer (CRC) samples, and higher expression correlated with more advanced TNM stage. We identified MAFB amplifications in a majority of tumor types in an assessment of The Cancer Genome Atlas database. Altered MAFB levels due to gene amplification, deletion, mutation, or transcription upregulation occurred in 9% of CRC cases within the database. shRNA knockdown experiments demonstrated that MAFB deficiency blocked CRC cell proliferation by arresting the cell cycle at G0/G1 phase in vitro. We found that MAFB could be SUMOylated by SUMO1 at lysine 32, and this modification was critical for cell cycle regulation by MAFB in CRC cells. SUMOylated MAFB directly regulated cyclin-dependent kinase 6 transcription by binding to its promoter. MAFB knockdown CRC cell xenograft tumors in mice grew more slowly than controls, and wild-type MAFB-overexpressing tumors grew more quickly than tumors overexpressing MAFB mutated at lysine 32. These data suggest that SUMOylated MAFB promotes CRC tumorigenesis through cell cycle regulation. MAFB and its SUMOylation process may serve as novel therapeutic targets for CRC treatment. PMID:27829226

Detection of Protein SUMOylation In Situ by Proximity Ligation Assays.

PubMed

Sahin, Umut; Jollivet, Florence; Berthier, Caroline; de Thé, Hugues; Lallemand-Breitenbach, Valérie

2016-01-01

Sumoylation is a posttranslational process essential for life and concerns a growing number of crucial proteins. Understanding the influence of this phenomenon on individual proteins or on cellular pathways in which they function has become an intense area of research. A critical step in studying protein sumoylation is to detect sumoylated forms of a particular protein. This has proven to be a challenging task for a number of reasons, especially in the case of endogenous proteins and in vivo studies or when studying rare cells such as stem cells. Proximity ligation assays that allow detection of closely interacting protein partners can be adapted for initial detection of endogenous sumoylation or ubiquitination in a rapid, ultrasensitive, and cheap manner. In addition, modified forms of a given protein can be detected in situ in various cellular compartments. Finally, the flexibility of this technique may allow rapid screening of drugs and stress signals that may modulate protein sumoylation.

SUMOylated IRF-1 shows oncogenic potential by mimicking IRF-2

DOE Office of Scientific and Technical Information (OSTI.GOV)

Park, Sun-Mi; School of Biological Sciences and Biotechnology, Chonnam National University, Gwangju 500-757; Chae, Myounghee

2010-01-01

Interferon regulatory factor-1 (IRF-1) is an interferon-induced transcriptional activator that suppresses tumors by impeding cell proliferation. Recently, we demonstrated that the level of SUMOylated IRF-1 is elevated in tumor cells, and that SUMOylation of IRF-1 attenuates its tumor-suppressive function. Here we report that SUMOylated IRF-1 mimics IRF-2, an antagonistic repressor, and shows oncogenic potential. To demonstrate the role of SUMOylated IRF-1 in tumorigenesis, we used SUMO-IRF-1 recombinant protein. Stable expression of SUMO-IRF-1 in NIH3T3 cells resulted in focus formation and anchorage-independent growth in soft agar. Inoculation of SUMO-IRF-1-transfected cells into athymic nude mice resulted in tumor formation and infiltration ofmore » adipose tissues. Finally, we demonstrated that SUMO-IRF-1 transforms NIH3T3 cells in a dose-dependent manner suggesting that SUMOylated IRF-1 may act as an oncogenic protein in tumor cells.« less

Cooling-induced SUMOylation of EXOSC10 down-regulates ribosome biogenesis

PubMed Central

Bastide, Amandine; Peretti, Diego; Roobol, Anne; Roobol, Jo; Mallucci, Giovanna R.; Smales, C. Mark; Willis, Anne E.

2016-01-01

The RNA exosome is essential for 3′ processing of functional RNA species and degradation of aberrant RNAs in eukaryotic cells. Recent reports have defined the substrates of the exosome catalytic domains and solved the multimeric structure of the exosome complex. However, regulation of exosome activity remains poorly characterized, especially in response to physiological stress. Following the observation that cooling of mammalian cells results in a reduction in 40S:60S ribosomal subunit ratio, we uncover regulation of the nuclear exosome as a result of reduced temperature. Using human cells and an in vivo model system allowing whole-body cooling, we observe reduced EXOSC10 (hRrp6, Pm/Scl-100) expression in the cold. In parallel, both models of cooling increase global SUMOylation, leading to the identification of specific conjugation of SUMO1 to EXOSC10, a process that is increased by cooling. Furthermore, we define the major SUMOylation sites in EXOSC10 by mutagenesis and show that overexpression of SUMO1 alone is sufficient to suppress EXOSC10 abundance. Reducing EXOSC10 expression by RNAi in human cells correlates with the 3′ preribosomal RNA processing defects seen in the cold as well as reducing the 40S:60S ratio, a previously uncharacterized consequence of EXOSC10 suppression. Together, this work illustrates that EXOSC10 can be modified by SUMOylation and identifies a physiological stress where this regulation is prevalent both in vitro and in vivo. PMID:26857222

Defining the SUMO System in Maize: SUMOylation Is Up-Regulated during Endosperm Development and Rapidly Induced by Stress1[OPEN

PubMed Central

Augustine, Robert C.; Rytz, Thérèse C.

2016-01-01

In response to abiotic and biotic challenges, plants rapidly attach small ubiquitin-related modifier (SUMO) to a large collection of nuclear proteins, with studies in Arabidopsis (Arabidopsis thaliana) linking SUMOylation to stress tolerance via its modification of factors involved in chromatin and RNA dynamics. Despite this importance, little is known about SUMOylation in crop species. Here, we describe the plant SUMO system at the phylogenetic, biochemical, and transcriptional levels with a focus on maize (Zea mays). In addition to canonical SUMOs, land plants encode a loosely constrained noncanonical isoform and a variant containing a long extension upstream of the signature β-grasp fold, with cereals also expressing a novel diSUMO polypeptide bearing two SUMO β-grasp domains in tandem. Maize and other cereals also synthesize a unique SUMO-conjugating enzyme variant with more restricted expression patterns that is enzymatically active despite a distinct electrostatic surface. Maize SUMOylation primarily impacts nuclear substrates, is strongly induced by high temperatures, and displays a memory that suppresses subsequent conjugation. Both in-depth transcript and conjugate profiles in various maize organs point to tissue/cell-specific functions for SUMOylation, with potentially significant roles during embryo and endosperm maturation. Collectively, these studies define the organization of the maize SUMO system and imply important functions during seed development and stress defense. PMID:27208252

Sumoylation of the Epstein-Barr Virus BZLF1 Protein Inhibits Its Transcriptional Activity and Is Regulated by the Virus-Encoded Protein Kinase▿

PubMed Central

Hagemeier, Stacy R.; Dickerson, Sarah J.; Meng, Qiao; Yu, Xianming; Mertz, Janet E.; Kenney, Shannon C.

2010-01-01

The Epstein-Barr virus (EBV) immediate-early protein BZLF1 (Z) mediates the switch between latent and lytic EBV infection. Z not only activates early lytic viral gene transcription but also plays a direct role in lytic viral genome replication. Although a small fraction of Z is known to be sumoylated, the effects of this posttranslational modification on various different Z functions have not been well defined. In this report, we show that only the lysine at amino acid residue 12 is required for the sumoylation of Z, and that Z can be sumoylated by SUMO isoforms 1, 2, and 3. We also demonstrate that the sumo-defective Z mutants ZK12A and ZK12R have enhanced transcriptional activity. The sumoylated and nonsumoylated forms of Z were found to have a similar cellular location, both being localized primarily within the nuclear matrix. The Z sumo-defective mutants were, however, partially defective for disrupting promyelocytic leukemia (PML) bodies compared to the ability of wild-type Z. In addition, we show that lytic viral genome replication does not require the sumoylation of Z, although a Z mutant altered at both amino acids 12 and 13 is replication defective. Furthermore, we show that the sumoylation of Z is greatly increased (from less than 1 to about 11%) in lytically induced 293 cells infected with an EBV mutant virus deleted for the EBV-encoded protein kinase (EBV-PK) compared to that of 293 cells infected with wild-type EBV, and that the overexpression of EBV-PK leads to the reduced sumoylation of Z in EBV-negative cells. Our results suggest that the sumoylation of Z helps to promote viral latency, and that EBV-PK inhibits Z sumoylation during viral reactivation. PMID:20181712

Development of a high-throughput screen to detect inhibitors of TRPS1 sumoylation.

PubMed

Brandt, Martin; Szewczuk, Lawrence M; Zhang, Hong; Hong, Xuan; McCormick, Patricia M; Lewis, Tia S; Graham, Taylor I; Hung, Sunny T; Harper-Jones, Amber D; Kerrigan, John J; Wang, Da-Yuan; Dul, Edward; Hou, Wangfang; Ho, Thau F; Meek, Thomas D; Cheung, Mui H; Johanson, Kyung O; Jones, Christopher S; Schwartz, Benjamin; Kumar, Sanjay; Oliff, Allen I; Kirkpatrick, Robert B

2013-06-01

Small ubiquitin-like modifier (SUMO) belongs to the family of ubiquitin-like proteins (Ubls) that can be reversibly conjugated to target-specific lysines on substrate proteins. Although covalently sumoylated products are readily detectible in gel-based assays, there has been little progress toward the development of robust quantitative sumoylation assay formats for the evaluation of large compound libraries. In an effort to identify inhibitors of ubiquitin carrier protein 9 (Ubc9)-dependent sumoylation, a high-throughput fluorescence polarization assay was developed, which allows detection of Lys-1201 sumoylation, corresponding to the major site of functional sumoylation within the transcriptional repressor trichorhino-phalangeal syndrome type I protein (TRPS1). A minimal hexapeptide substrate peptide, TMR-VVK₁₂₀₁TEK, was used in this assay format to afford high-throughput screening of the GlaxoSmithKline diversity compound collection. A total of 728 hits were confirmed but no specific noncovalent inhibitors of Ubc9 dependent trans-sumoylation were found. However, several diaminopyrimidine compounds were identified as inhibitors in the assay with IC₅₀ values of 12.5 μM. These were further characterized to be competent substrates which were subject to sumoylation by SUMO-Ubc9 and which were competitive with the sumoylation of the TRPS1 peptide substrates.

Sumoylation of CCAAT/enhancer-binding protein α is implicated in hematopoietic stem/progenitor cell development through regulating runx1 in zebrafish.

PubMed

Yuan, Hao; Zhang, Tao; Liu, Xiaohui; Deng, Min; Zhang, Wenqing; Wen, Zilong; Chen, Saijuan; Chen, Zhu; de The, Hugues; Zhou, Jun; Zhu, Jun

2015-03-11

The small ubiquitin-related modifier (SUMO) participates in various cellular processes, including maintenance of genome integrity, nuclear transport, transcription and signal transduction. However, the biological function of sumoylation in hematopoiesis has not been fully explored. We show here that definitive hematopoietic stem/progenitor cells (HSPCs) are depleted in SUMO-deficient zebrafish embryos. Impairment of sumoylation attenuates HSPC generation and proliferation. The hyposumoylation triggered HSPC defects are CCAAT/enhancer-binding protein α (C/ebpα) dependent. Critically, a SUMO-C/ebpα fusion rescues the defective hematopoiesis in SUMO-deficient embryos, at least in part through restored runx1 expression. While C/ebpα-dependent transcription is involved in myeloid differentiation, our studies here reveal that C/ebpα sumoylation is essential for HSPC development during definitive hematopoiesis.

TRPV1 SUMOylation regulates nociceptive signaling in models of inflammatory pain.

PubMed

Wang, Yan; Gao, Yingwei; Tian, Quan; Deng, Qi; Wang, Yangbo; Zhou, Tian; Liu, Qiang; Mei, Kaidi; Wang, Yingping; Liu, Huiqing; Ma, Ruining; Ding, Yuqiang; Rong, Weifang; Cheng, Jinke; Yao, Jing; Xu, Tian-Le; Zhu, Michael X; Li, Yong

2018-04-18

Although TRPV1 channels represent a key player of noxious heat sensation, the precise mechanisms for thermal hyperalgesia remain unknown. We report here that conditional knockout of deSUMOylation enzyme, SENP1, in mouse dorsal root ganglion (DRG) neurons exacerbated thermal hyperalgesia in both carrageenan- and Complete Freund's adjuvant-induced inflammation models. TRPV1 is SUMOylated at a C-terminal Lys residue (K822), which specifically enhances the channel sensitivity to stimulation by heat, but not capsaicin, protons or voltage. TRPV1 SUMOylation is decreased by SENP1 but upregulated upon peripheral inflammation. More importantly, the reduced ability of TRPV1 knockout mice to develop inflammatory thermal hyperalgesia was rescued by viral infection of lumbar 3/4 DRG neurons of wild-type TRPV1, but not its SUMOylation-deficient mutant, K822R. These data suggest that TRPV1 SUMOylation is essential for the development of inflammatory thermal hyperalgesia, through a mechanism that involves sensitization of the channel response specifically to thermal stimulation.

Shigella entry unveils a calcium/calpain-dependent mechanism for inhibiting sumoylation

PubMed Central

Lhocine, Nouara; Andrieux, Alexandra; Nigro, Giulia; Mounier, Joëlle

2017-01-01

Disruption of the sumoylation/desumoylation equilibrium is associated with several disease states such as cancer and infections, however the mechanisms regulating the global SUMO balance remain poorly defined. Here, we show that infection by Shigella flexneri, the causative agent of human bacillary dysentery, switches off host sumoylation during epithelial cell infection in vitro and in vivo and that this effect is mainly mediated by a calcium/calpain-induced cleavage of the SUMO E1 enzyme SAE2, thus leading to sumoylation inhibition. Furthermore, we describe a mechanism by which Shigella promotes its own invasion by altering the sumoylation state of RhoGDIα, a master negative regulator of RhoGTPase activity and actin polymerization. Together, our data suggest that SUMO modification is essential to restrain pathogenic bacterial entry by limiting cytoskeletal rearrangement induced by bacterial effectors. Moreover, these findings identify calcium-activated calpains as powerful modulators of cellular sumoylation levels with potentially broad implications in several physiological and pathological situations. PMID:29231810

DeSUMOylation switches Kaiso from activator to repressor upon hyperosmotic stress.

PubMed

Zhenilo, Svetlana; Deyev, Igor; Litvinova, Ekaterina; Zhigalova, Nadezhda; Kaplun, Daria; Sokolov, Alexey; Mazur, Alexander; Prokhortchouk, Egor

2018-02-22

Kaiso is a member of the BTB/POZ zinc finger family, which is involved in cancer progression, cell cycle control, apoptosis, and WNT signaling. Depending on promoter context, it may function as either a transcriptional repressor or activator. Previous studies found that Kaiso might be SUMOylated due to heat shock, but the biological significance of Kaiso SUMOylation is unclear. Here, we find that K42 is the only amino acid within Kaiso that is modified with SUMO. Kaiso is monoSUMOylated at lysine 42 in cell lines of kidney origin under normal physiological conditions. SUMOylated Kaiso can activate transcription from exogenous methylated promoters, wherein the deSUMOylated form of the protein kept the ability to be a repressor. Rapid Kaiso deSUMOylation occurs in response to hyperosmotic stress and is reversible upon return to an isotonic environment. DeSUMOylation occurs within minutes in HEK293 cells treated with 100 mM NaCl and relaxes in 3 h even in a salt-containing medium. Genomic editing of Kaiso by conversion of K42 into R42 (K42R) in HEK293 cells that resulted in fully deSUMOylated endogenous protein led to misregulation of genes associated with ion transport, blood pressure, and the immune response. TRIM25 was significantly repressed in two K42R HEK293 clones. By a series of rescue experiments with K42R and KO HEK293 cells, we show that TRIM25 is a direct transcriptional target for Kaiso. In the absence of Kaiso, the level of TRIM25 is insensitive to hyperosmotic stress. Extending our observations to animal models, we show that in response to a high salt diet, Kaiso knockout mice are characterized by significantly higher blood pressure increases when compared to wild-type animals. Thus, we propose a novel biological role for Kaiso in the regulation of homeostasis.

Organization and Regulation of Soybean SUMOylation System under Abiotic Stress Conditions

PubMed Central

Li, Yanjun; Wang, Guixin; Xu, Zeqian; Li, Jing; Sun, Mengwei; Guo, Jingsong; Ji, Wei

2017-01-01

Covalent attachment of the small ubiquitin-related modifier, SUMO, to substrate proteins plays a significant role in plants under stress conditions, which can alter target proteins' function, location, and protein-protein interactions. Despite this importance, information about SUMOylation in the major legume crop, soybean, remains obscure. In this study, we performed a bioinformatics analysis of the entire soybean genome and identified 40 genes belonged to six families involved in a cascade of enzymatic reactions in soybean SUMOylation system. The cis-acting elements analysis revealed that promoters of SUMO pathway genes contained different combinations of stress and development-related cis-regulatory elements. RNA-seq data analysis showed that SUMO pathway components exhibited versatile tissue-specific expression patterns, indicating coordinated functioning during plant growth and development. qRT-PCR analysis of 13 SUMO pathway members indicated that majority of the SUMO pathway members were transcriptionally up-regulated by NaCl, heat and ABA stimuli during the 24 h period of treatment. Furthermore, SUMOylation dynamics in soybean roots under abiotic stress treatment were analyzed by western blot, which were characterized by regulation of SUMOylated proteins. Collectively, this study defined the organization of the soybean SUMOylation system and implied an essential function for SUMOylation in soybean abiotic stress responses. PMID:28878795

Sumoylation of CCAAT/enhancer-binding protein α is implicated in hematopoietic stem/progenitor cell development through regulating runx1 in zebrafish

PubMed Central

Yuan, Hao; Zhang, Tao; Liu, Xiaohui; Deng, Min; Zhang, Wenqing; Wen, Zilong; Chen, Saijuan; Chen, Zhu; de The, Hugues; Zhou, Jun; Zhu, Jun

2015-01-01

The small ubiquitin-related modifier (SUMO) participates in various cellular processes, including maintenance of genome integrity, nuclear transport, transcription and signal transduction. However, the biological function of sumoylation in hematopoiesis has not been fully explored. We show here that definitive hematopoietic stem/progenitor cells (HSPCs) are depleted in SUMO-deficient zebrafish embryos. Impairment of sumoylation attenuates HSPC generation and proliferation. The hyposumoylation triggered HSPC defects are CCAAT/enhancer-binding protein α (C/ebpα) dependent. Critically, a SUMO-C/ebpα fusion rescues the defective hematopoiesis in SUMO-deficient embryos, at least in part through restored runx1 expression. While C/ebpα-dependent transcription is involved in myeloid differentiation, our studies here reveal that C/ebpα sumoylation is essential for HSPC development during definitive hematopoiesis. PMID:25757417

A Non-SUMOylated Tax Protein Is Still Functional for NF-κB Pathway Activation

PubMed Central

Pène, Sabrina; Waast, Laetitia; Bonnet, Amandine; Bénit, Laurence

2014-01-01

ABSTRACT Whether NF-κB promoter transactivation by the human T-cell leukemia virus type 1 (HTLV-1) Tax protein requires Tax SUMOylation is still a matter of debate. In this study, we revisited the role of Tax SUMOylation using a strategy based on the targeting of Ubc9, the unique E2 SUMO-conjugating enzyme. We show that either a catalytically inactive form of Ubc9 (Ubc9-C93S) or Ubc9 small interfering RNA (siRNA) dramatically reduces Tax conjugation to endogenous SUMO-1 or SUMO-2/3, demonstrating that as expected, Tax SUMOylation is under the control of the catalytic activity of Ubc9. We further report that a non-SUMOylated Tax protein produced in 293T cells is still able to activate either a transfected or an integrated NF-κB reporter promoter and to induce expression of an NF-κB-regulated endogenous gene. Importantly, blocking Ubc9 activity in T cells also results in the production of a non-SUMOylated Tax that is still fully functional for the activation of a NF-κB promoter. These results provide the definitive evidence that Tax SUMOylation is not required for NF-κB-driven gene induction. IMPORTANCE Human T-cell leukemia virus type 1 is able to transform CD4+ T lymphocytes. The viral oncoprotein Tax plays a key role in this process by promoting cell proliferation and survival, mainly through permanent activation of the NF-κB pathway. Elucidating the molecular mechanisms involved in NF-κB pathway activation by Tax is therefore a key issue to understand HTLV-1-mediated transformation. Tax SUMOylation was initially proposed to be critical for Tax-induced NF-κB promoter activation, which was challenged by our later observation that a low-level-SUMOylated Tax mutant was still functional for activation of NF-κB promoters. To clarify the role of Tax SUMOylation, we set up a new approach based on the inhibition of the SUMOylation machinery in Tax-expressing cells. We show that blocking the SUMO-conjugating enzyme Ubc9 abolishes Tax SUMOylation and that a non-SUMOylated

A non-SUMOylated tax protein is still functional for NF-κB pathway activation.

PubMed

Pène, Sabrina; Waast, Laetitia; Bonnet, Amandine; Bénit, Laurence; Pique, Claudine

2014-09-01

Whether NF-κB promoter transactivation by the human T-cell leukemia virus type 1 (HTLV-1) Tax protein requires Tax SUMOylation is still a matter of debate. In this study, we revisited the role of Tax SUMOylation using a strategy based on the targeting of Ubc9, the unique E2 SUMO-conjugating enzyme. We show that either a catalytically inactive form of Ubc9 (Ubc9-C93S) or Ubc9 small interfering RNA (siRNA) dramatically reduces Tax conjugation to endogenous SUMO-1 or SUMO-2/3, demonstrating that as expected, Tax SUMOylation is under the control of the catalytic activity of Ubc9. We further report that a non-SUMOylated Tax protein produced in 293T cells is still able to activate either a transfected or an integrated NF-κB reporter promoter and to induce expression of an NF-κB-regulated endogenous gene. Importantly, blocking Ubc9 activity in T cells also results in the production of a non-SUMOylated Tax that is still fully functional for the activation of a NF-κB promoter. These results provide the definitive evidence that Tax SUMOylation is not required for NF-κB-driven gene induction. Human T-cell leukemia virus type 1 is able to transform CD4(+) T lymphocytes. The viral oncoprotein Tax plays a key role in this process by promoting cell proliferation and survival, mainly through permanent activation of the NF-κB pathway. Elucidating the molecular mechanisms involved in NF-κB pathway activation by Tax is therefore a key issue to understand HTLV-1-mediated transformation. Tax SUMOylation was initially proposed to be critical for Tax-induced NF-κB promoter activation, which was challenged by our later observation that a low-level-SUMOylated Tax mutant was still functional for activation of NF-κB promoters. To clarify the role of Tax SUMOylation, we set up a new approach based on the inhibition of the SUMOylation machinery in Tax-expressing cells. We show that blocking the SUMO-conjugating enzyme Ubc9 abolishes Tax SUMOylation and that a non-SUMOylated Tax still

Sumoylation of FOXP2 regulates motor function and vocal communication through Purkinje cell development

PubMed Central

Usui, Noriyoshi; Co, Marissa; Harper, Matthew; Rieger, Michael A.; Dougherty, Joseph D.; Konopka, Genevieve

2016-01-01

Background Mutations in the gene encoding the transcription factor forkhead box P2, FOXP2, result in brain developmental abnormalities including reduced gray matter in both human patients and rodent models, and speech and language deficits. However, neither the region-specific function of FOXP2 in the brain, in particular the cerebellum, nor the effects of any post-translational modifications of FOXP2 in the brain and disorders have been explored. Methods We characterized sumoylation of FOXP2 biochemically, and analyzed the region-specific function and sumoylation of FOXP2 in the developing mouse cerebellum. Using in utero electroporation to manipulate the sumoylation-state of Foxp2 as well as Foxp2 expression levels in Purkinje cells (PCs) of the cerebellum in vivo, we reduced Foxp2 expression approximately 40% in the mouse cerebellum. Such a reduction approximates the haploinsufficiency observed in human patients who demonstrate speech and language impairments. Results We identified sumoylation of FOXP2 at K674 (K673 in mouse) in the cerebellum of neonates. In vitro co-immunoprecipitation and in vivo colocalization experiments suggest that PIAS3 acts as the SUMO E3 ligase for FOXP2 sumoylation. This sumoylation modifies transcriptional regulation by FOXP2. We demonstrate that Foxp2 sumoylation is required for regulation of cerebellar motor function and vocal communication, likely through dendritic outgrowth and arborization of PCs in the mouse cerebellum. Conclusions Sumoylation of Foxp2 in neonatal mouse cerebellum regulates PC development as well as motor functions and vocal communication, demonstrating evidence for sumoylation in regulating mammalian behaviors. PMID:27009683

Sumoylation of FOXP2 Regulates Motor Function and Vocal Communication Through Purkinje Cell Development.

PubMed

Usui, Noriyoshi; Co, Marissa; Harper, Matthew; Rieger, Michael A; Dougherty, Joseph D; Konopka, Genevieve

2017-02-01

Mutations in the gene encoding the transcription factor forkhead box P2 (FOXP2) result in brain developmental abnormalities, including reduced gray matter in both human patients and rodent models and speech and language deficits. However, neither the region-specific function of FOXP2 in the brain, in particular the cerebellum, nor the effects of any posttranslational modifications of FOXP2 in the brain and disorders have been explored. We characterized sumoylation of FOXP2 biochemically and analyzed the region-specific function and sumoylation of FOXP2 in the developing mouse cerebellum. Using in utero electroporation to manipulate the sumoylation state of FOXP2 as well as Foxp2 expression levels in Purkinje cells of the cerebellum in vivo, we reduced Foxp2 expression approximately 40% in the mouse cerebellum. Such a reduction approximates the haploinsufficiency observed in human patients who demonstrate speech and language impairments. We identified sumoylation of FOXP2 at K674 (K673 in mice) in the cerebellum of neonates. In vitro co-immunoprecipitation and in vivo colocalization experiments suggest that PIAS3 acts as the small ubiquitin-like modifier E3 ligase for FOXP2 sumoylation. This sumoylation modifies transcriptional regulation by FOXP2. We demonstrated that FOXP2 sumoylation is required for regulation of cerebellar motor function and vocal communication, likely through dendritic outgrowth and arborization of Purkinje cells in the mouse cerebellum. Sumoylation of FOXP2 in neonatal mouse cerebellum regulates Purkinje cell development and motor functions and vocal communication, demonstrating evidence for sumoylation in regulating mammalian behaviors. Copyright © 2016 Society of Biological Psychiatry. Published by Elsevier Inc. All rights reserved.

SUMOylation of TARBP2 regulates miRNA/siRNA efficiency

PubMed Central

Chen, Cheng; Zhu, Changhong; Huang, Jian; Zhao, Xian; Deng, Rong; Zhang, Hailong; Dou, Jinzhuo; Chen, Qin; Xu, Ming; Yuan, Haihua; Wang, Yanli; Yu, Jianxiu

2015-01-01

Small RNA-induced gene silencing is essential for post-transcriptional regulation of gene expression; however, it remains unclear how miRNA/siRNA efficiency is regulated. Here we show that TARBP2 is SUMOylated at K52, which can be enhanced by its phosphorylation. This modification can stabilize TARBP2 via repressing its K48-linked ubiquitination. We find that TARBP2 SUMOylation does not influence the overall production of mature miRNAs, but it regulates miRNA/siRNA efficiency. SUMOylated TARBP2 recruits Ago2 to constitute the RNA-induced silencing complex (RISC)-loading complex (RLC), and simultaneously promotes more pre-miRNAs to load into the RLC. Consequently, Ago2 is stabilized and miRNAs/siRNAs bound by TARBP2/Dicer is effectively transferred to Ago2. Thus, these processes lead to the formation of the effective RISC for RNA interference (RNAi). Collectively, our data suggest that SUMOylation of TARBP2 is required for regulating miRNA/siRNA efficiency, which is a general mechanism of miRNA/siRNA regulation. PMID:26582366

DeSUMOylation of MKK7 kinase by the SUMO2/3 protease SENP3 potentiates lipopolysaccharide-induced inflammatory signaling in macrophages

PubMed Central

Lao, Yimin; Yang, Kai; Wang, Zhaojun; Sun, Xueqing; Zou, Qiang; Yu, Xiaoyan; Cheng, Jinke; Tong, Xuemei; Yeh, Edward T. H.; Yang, Jie; Yi, Jing

2018-01-01

Protein SUMOylation has been reported to play a role in innate immune response, but the enzymes, substrates, and consequences of the specific inflammatory signaling events are largely unknown. Reactive oxygen species (ROS) are abundantly produced during macrophage activation and required for Toll-like receptor 4 (TLR4)–mediated inflammatory signaling. Previously, we demonstrated that SENP3 is a redox-sensitive SUMO2/3 protease. To explore any links between reversible SUMOylation and ROS-related inflammatory signaling in macrophage activation, we generated mice with Senp3 conditional knock-out in myeloid cells. In bacterial lipopolysaccharide (LPS)-induced in vitro and in vivo inflammation models, we found that SENP3 deficiency markedly compromises the activation of TLR4 inflammatory signaling and the production of proinflammatory cytokines in macrophages exposed to LPS. Moreover, Senp3 conditional knock-out mice were significantly less susceptible to septic shock. Of note, SENP3 deficiency was associated with impairment in JNK phosphorylation. We found that MKK7, which selectively phosphorylates JNK, is a SENP3 substrate and that SENP3-mediated deSUMOylation of MKK7 may favor its binding to JNK. Importantly, ROS-dependent SENP3 accumulation and MKK7 deSUMOylation rapidly occurred after LPS stimulation. In conclusion, our findings indicate that SENP3 potentiates LPS-induced TLR4 signaling via deSUMOylation of MKK7 leading to enhancement in JNK phosphorylation and the downstream events. Therefore this work provides novel mechanistic insights into redox regulation of innate immune responses. PMID:29352108

Human Adenovirus Core Protein V Is Targeted by the Host SUMOylation Machinery To Limit Essential Viral Functions.

PubMed

Freudenberger, Nora; Meyer, Tina; Groitl, Peter; Dobner, Thomas; Schreiner, Sabrina

2018-02-15

Human adenoviruses (HAdV) are nonenveloped viruses containing a linear, double-stranded DNA genome surrounded by an icosahedral capsid. To allow proper viral replication, the genome is imported through the nuclear pore complex associated with viral core proteins. Until now, the role of these incoming virion proteins during the early phase of infection was poorly understood. The core protein V is speculated to bridge the core and the surrounding capsid. It binds the genome in a sequence-independent manner and localizes in the nucleus of infected cells, accumulating at nucleoli. Here, we show that protein V contains conserved SUMO conjugation motifs (SCMs). Mutation of these consensus motifs resulted in reduced SUMOylation of the protein; thus, protein V represents a novel target of the host SUMOylation machinery. To understand the role of protein V SUMO posttranslational modification during productive HAdV infection, we generated a replication-competent HAdV with SCM mutations within the protein V coding sequence. Phenotypic analyses revealed that these SCM mutations are beneficial for adenoviral replication. Blocking protein V SUMOylation at specific sites shifts the onset of viral DNA replication to earlier time points during infection and promotes viral gene expression. Simultaneously, the altered kinetics within the viral life cycle are accompanied by more efficient proteasomal degradation of host determinants and increased virus progeny production than that observed during wild-type infection. Taken together, our studies show that protein V SUMOylation reduces virus growth; hence, protein V SUMOylation represents an important novel aspect of the host antiviral strategy to limit virus replication and thereby points to potential intervention strategies. IMPORTANCE Many decades of research have revealed that HAdV structural proteins promote viral entry and mainly physical stability of the viral genome in the capsid. Our work over the last years showed that this

The Key Regulator for Language and Speech Development, FOXP2, is a Novel Substrate for SUMOylation.

PubMed

Meredith, Leslie J; Wang, Chiung-Min; Nascimento, Leticia; Liu, Runhua; Wang, Lizhong; Yang, Wei-Hsiung

2016-02-01

Transcription factor forkhead box protein P2 (FOXP2) plays an essential role in the development of language and speech. However, the transcriptional activity of FOXP2 regulated by the post-translational modifications remains unknown. Here, we demonstrated that FOXP2 is clearly defined as a SUMO target protein at the cellular levels as FOXP2 is covalently modified by both SUMO1 and SUMO3. Furthermore, SUMOylation of FOXP2 was significantly decreased by SENP2 (a specific SUMOylation protease). We further showed that FOXP2 is selectively SUMOylated in vivo on a phylogenetically conserved lysine 674 but the SUMOylation does not alter subcellular localization and stability of FOXP2. Interestingly, we observed that human etiological FOXP2 R553H mutation robustly reduces its SUMOylation potential as compared to wild-type FOXP2. In addition, the acidic residues downstream the core SUMO motif on FOXP2 are required for its full SUMOylation capacity. Finally, our functional analysis using reporter gene assays showed that SUMOylation may modulate transcriptional activity of FOXP2 in regulating downstream target genes (DISC1, SRPX2, and MiR200c). Altogether, we provide the first evidence that FOXP2 is a substrate for SUMOylation and SUMOylation of FOXP2 plays a functional role in regulating its transcriptional activity. © 2015 Wiley Periodicals, Inc.

The Key Regulator for Language and Speech Development, FOXP2, is a Novel Substrate for SUMOylation

PubMed Central

Meredith, Leslie J.; Wang, Chiung-Min; Nascimento, Leticia; Liu, Runhua; Wang, Lizhong; Yang, Wei-Hsiung

2017-01-01

Transcription factor forkhead box protein P2 (FOXP2) plays an essential role in the development of language and speech. However, the transcriptional activity of FOXP2 regulated by the post-translational modifications remains unknown. Here we demonstrated that FOXP2 is clearly defined as a SUMO target protein at the cellular levels as FOXP2 is covalently modified by both SUMO1 and SUMO3. Furthermore, SUMOylation of FOXP2 was significantly decreased by SENP2 (a specific SUMOylation protease). We further showed that FOXP2 is selectively SUMOylated in vivo on a phylogenetically conserved lysine 674 but the SUMOylation does not alter subcellular localization and stability of FOXP2. Interestingly, we observed that human etiological FOXP2 R553H mutation robustly reduces its SUMOylation potential as compared to wild-type FOXP2. In addition, the acidic residues downstream the core SUMO motif on FOXP2 are required for its full SUMOylation capacity. Finally, our functional analysis using reporter gene assays showed that SUMOylation may modulate transcriptional activity of FOXP2 in regulating downstream target genes (DISC1, SRPX2 and MiR200c). Altogether, we provide the first evidence that FOXP2 is a substrate for SUMOylation and SUMOylation of FOXP2 plays a functional role in regulating its transcriptional activity. PMID:26212494

Sumoylation activates the transcriptional activity of Pax-6, an important transcription factor for eye and brain development

PubMed Central

Yan, Qin; Gong, Lili; Deng, Mi; Zhang, Lan; Sun, Shuming; Liu, Jiao; Ma, Haili; Yuan, Dan; Chen, Pei-Chao; Hu, Xiaohui; Liu, Jinping; Qin, Jichao; Xiao, Ling; Huang, Xiao-Qin; Zhang, Jian; Wan-Cheng Li, David

2010-01-01

Pax-6 is an evolutionarily conserved transcription factor regulating brain and eye development. Four Pax-6 isoforms have been reported previously. Although the longer Pax-6 isoforms (p46 and p48) bear two DNA-binding domains, the paired domain (PD) and the homeodomain (HD), the shorter Pax-6 isoform p32 contains only the HD for DNA binding. Although a third domain, the proline-, serine- and threonine-enriched activation (PST) domain, in the C termini of all Pax-6 isoforms mediates their transcriptional modulation via phosphorylation, how p32 Pax-6 could regulate target genes remains to be elucidated. In the present study, we show that sumoylation at K91 is required for p32 Pax-6 to bind to a HD-specific site and regulate expression of target genes. First, in vitro-synthesized p32 Pax-6 alone cannot bind the P3 sequence, which contains the HD recognition site, unless it is preincubated with nuclear extracts precleared by anti–Pax-6 but not by anti-small ubiquitin-related modifier 1 (anti-SUMO1) antibody. Second, in vitro-synthesized p32 Pax-6 can be sumoylated by SUMO1, and the sumoylated p32 Pax-6 then can bind to the P3 sequence. Third, Pax-6 and SUMO1 are colocalized in the embryonic optic and lens vesicles and can be coimmunoprecipitated. Finally, SUMO1-conjugated p32 Pax-6 exists in both the nucleus and cytoplasm, and sumoylation significantly enhances the DNA-binding ability of p32 Pax-6 and positively regulates gene expression. Together, our results demonstrate that sumoylation activates p32 Pax-6 in both DNA-binding and transcriptional activities. In addition, our studies demonstrate that p32 and p46 Pax-6 possess differential DNA-binding and regulatory activities. PMID:21084637

SUMOylation of phytochrome-B negatively regulates light-induced signaling in Arabidopsis thaliana

PubMed Central

Sadanandom, Ari; Ádám, Éva; Orosa, Beatriz; Viczián, András; Klose, Cornelia; Zhang, Cunjin; Josse, Eve-Marie; Kozma-Bognár, László; Nagy, Ferenc

2015-01-01

The red/far red light absorbing photoreceptor phytochrome-B (phyB) cycles between the biologically inactive (Pr, λmax, 660 nm) and active (Pfr; λmax, 730 nm) forms and functions as a light quality and quantity controlled switch to regulate photomorphogenesis in Arabidopsis. At the molecular level, phyB interacts in a conformation-dependent fashion with a battery of downstream regulatory proteins, including PHYTOCHROME INTERACTING FACTOR transcription factors, and by modulating their activity/abundance, it alters expression patterns of genes underlying photomorphogenesis. Here we report that the small ubiquitin-like modifier (SUMO) is conjugated (SUMOylation) to the C terminus of phyB; the accumulation of SUMOylated phyB is enhanced by red light and displays a diurnal pattern in plants grown under light/dark cycles. Our data demonstrate that (i) transgenic plants expressing the mutant phyBLys996Arg-YFP photoreceptor are hypersensitive to red light, (ii) light-induced SUMOylation of the mutant phyB is drastically decreased compared with phyB-YFP, and (iii) SUMOylation of phyB inhibits binding of PHYTOCHROME INTERACTING FACTOR 5 to phyB Pfr. In addition, we show that OVERLY TOLERANT TO SALT 1 (OTS1) de-SUMOylates phyB in vitro, it interacts with phyB in vivo, and the ots1/ots2 mutant is hyposensitive to red light. Taken together, we conclude that SUMOylation of phyB negatively regulates light signaling and it is mediated, at least partly, by the action of OTS SUMO proteases. PMID:26283376

PML, SUMOylation, and Senescence

PubMed Central

Ivanschitz, Lisa; De Thé, Hugues; Le Bras, Morgane

2013-01-01

Since its discovery, 25 years ago, promyelocytic leukemia (PML) has been an enigma. Implicated in the oncogenic PML/RARA fusion, forming elusive intranuclear domains, triggering cell death or senescence, controlled by and perhaps controlling SUMOylation… there are multiple PML-related issues. Here we review the reciprocal interactions between PML, senescence, and SUMOylation, notably in the context of cellular transformation. PMID:23847762

Dynamic Sumoylation of a Conserved Transcription Corepressor Prevents Persistent Inclusion Formation during Hyperosmotic Stress

PubMed Central

Oeser, Michelle L.; Amen, Triana; Nadel, Cory M.; Bradley, Amanda I.; Reed, Benjamin J.; Jones, Ramon D.; Gopalan, Janani; Kaganovich, Daniel; Gardner, Richard G.

2016-01-01

Cells are often exposed to physical or chemical stresses that can damage the structures of essential biomolecules. Stress-induced cellular damage can become deleterious if not managed appropriately. Rapid and adaptive responses to stresses are therefore crucial for cell survival. In eukaryotic cells, different stresses trigger post-translational modification of proteins with the small ubiquitin-like modifier SUMO. However, the specific regulatory roles of sumoylation in each stress response are not well understood. Here, we examined the sumoylation events that occur in budding yeast after exposure to hyperosmotic stress. We discovered by proteomic and biochemical analyses that hyperosmotic stress incurs the rapid and transient sumoylation of Cyc8 and Tup1, which together form a conserved transcription corepressor complex that regulates hundreds of genes. Gene expression and cell biological analyses revealed that sumoylation of each protein directs distinct outcomes. In particular, we discovered that Cyc8 sumoylation prevents the persistence of hyperosmotic stress-induced Cyc8-Tup1 inclusions, which involves a glutamine-rich prion domain in Cyc8. We propose that sumoylation protects against persistent inclusion formation during hyperosmotic stress, allowing optimal transcriptional function of the Cyc8-Tup1 complex. PMID:26800527

Human stanniocalcin-1 interacts with nuclear and cytoplasmic proteins and acts as a SUMO E3 ligase.

PubMed

dos Santos, Marcos Tadeu; Trindade, Daniel Maragno; Gonçalves, Kaliandra de Almeida; Bressan, Gustavo Costa; Anastassopoulos, Filipe; Yunes, José Andres; Kobarg, Jörg

2011-01-01

Human stanniocalcin-1 (STC1) is a glycoprotein that has been implicated in different physiological process, including angiogenesis, apoptosis and carcinogenesis. Here we identified STC1 as a putative molecular marker for the leukemic bone marrow microenvironment and identified new interacting protein partners for STC1. Seven selected interactions retrieved from yeast two-hybrid screens were confirmed by GST-pull down assays in vitro. The N-terminal region was mapped to be the region that mediates the interaction with cytoplasmic, mitochondrial and nuclear proteins. STC1 interacts with SUMO-1 and several proteins that have been shown to be SUMOylated and localized to SUMOylation related nuclear bodies. Although STC1 interacts with SUMO-1 and has a high theoretical prediction score for a SUMOylation site, endogenous co-immunoprecipitation and in vitro SUMOylation assays with the purified recombinant protein could not detect STC1 SUMOylation. However, when we tested STC1 for SUMO E3 ligase activity, we found in an in vitro assay, that it significantly increases the SUMOylation of two other proteins. Confocal microscopic subcellular localization studies using both transfected cells and specific antibodies for endogenous STC1 revealed a cytoplasmic and nuclear deposition, the latter in the form of some specific dot-like substructure resembling SUMOylation related nuclear bodies. Together, these findings suggest a new role for STC1 in SUMOylation pathways, in nuclear bodies.

SUMOylation regulates TGF-β1/Smad4 signalling in-resistant glioma cells.

PubMed

Wang, Zhengfeng; Wang, Kai; Wang, Ruihua; Liu, Xianzhi

2017-12-18

The aim of this study was to explore the role of TGF-β1/Smad4 signalling in the DNA damage-induced ionization radiation (IR) resistance of glioma cells. T98G cells were assigned to the IR group (treated with IR) or the Blank group (with no treatment). The IR-treated cells were also treated/transfected with the TGF-β receptor inhibitor SB431542, SUMO1-overexpressing plasmids (SUMO1 group), SUMO1-interfering plasmids (si-SUMO1 group) or negative control plasmids group. The wound-healing capacity, cell proliferation and cell apoptosis were evaluated by the scratch assay, flow cytometry and the CCK-8 assay, respectively, and protein interactions were investigated by coimmunoprecipitation and colocalization assays. IR-treated T98G cells had DNA damage, but the wound-healing capacity and cell apoptosis were not significantly suppressed. DNA damage also induced TGF-β1, Smad4, SUMO1, SUMO2/3 and Ubc9 expression. In IR-treated cells cultured with SB431542, the wound-healing capacity and proliferation were promoted. SUMO1 and Smad4 colocalized in the nucleus of T98G cells, and the IR-treated cells had a significantly higher expression of the SUMO1-Smad4 protein complex. Smad4 expression in the nucleus was significantly reduced in the si-SUMO1 group, but was markedly increased in the SUMO1 group; the SUMO1 group had significantly elevated apoptotic activity, whereas the si-SUMO1 group showed significantly suppressed apoptotic activity and the si-SUMO1+SB41542 group had the lowest levels of cell apoptosis. DNA damage may activate Smad4 SUMOylation and the SUMOylation of Smad4 participates in the activation of TGF-β/Smad4 signalling; therefore, enhanced Smad4 SUMOylation is critical for the damage-induced activation of IR resistance.

Senp2 regulates adipose lipid storage by de-SUMOylation of Setdb1.

PubMed

Zheng, Quan; Cao, Ying; Chen, Yalan; Wang, Jiqiu; Fan, Qiuju; Huang, Xian; Wang, Yiping; Wang, Tianshi; Wang, Xiuzhi; Ma, Jiao; Cheng, Jinke

2018-06-01

One major function of adipocytes is to store excess energy in the form of triglycerides. Insufficient adipose lipid storage is associated with many pathological conditions including hyperlipidemia, insulin resistance, and type 2 diabetes. In this study, we observed the overexpression of SUMO-specific protease 2 (Senp2) in adipose tissues during obesity. Adipocyte Senp2 deficiency resulted in less adipose lipid storage accompanied by an ectopic fat accumulation and insulin resistance under high-fat diet feeding. We further found that SET domain bifurcated 1 (Setdb1) was a SUMOylated protein and that SUMOylation promoted Setdb1 occupancy on the promoter locus of Pparg and Cebpa genes to suppress their expressions by H3K9me3. Senp2 could suppress Setdb1 function by de-SUMOylation. In adipocyte Senp2-deficiency mice, accumulation of the SUMOylated Setdb1 suppressed the expression of Pparg and Cebpa genes as well as lipid metabolism-related target genes, which would decrease the ability of lipid storage in adipocytes. These results revealed the crucial role of Senp2-Setdb1 axis in controlling adipose lipid storage.

RUNX family members are covalently modified and regulated by PIAS1-mediated sumoylation

PubMed Central

Kim, J-H; Jang, J-W; Lee, Y-S; Lee, J-W; Chi, X-Z; Li, Y-H; Kim, M-K; Kim, D-M; Choi, B-S; Kim, J; Kim, H-M; van Wijnen, A; Park, IlY; Bae, S-C

2014-01-01

Transcription factors of the RUNX family (RUNXs), which play pivotal roles in normal development and neoplasia, are regulated by various post-translational modifications. To understand the molecular mechanisms underlying the regulation of RUNXs, we performed a large-scale functional genetic screen of a fly mutant library. The screen identified dPias (the fly ortholog of mammalian PIASs), an E3 ligase for the SUMO (small ubiquitin-like modifier) modification, as a novel genetic modifier of lz (the fly ortholog of mammalian RUNX3). Molecular biological analysis revealed that lz/RUNXs are sumoylated by dPias/PIAS1 at an evolutionarily conserved lysine residue (K372 of lz, K144 of RUNX1, K181 of RUNX2 and K148 of RUNX3). PIAS1-mediated sumoylation inhibited RUNX3 transactivation activity, and this modification was promoted by the AKT1 kinase. Importantly, PIAS1 failed to sumoylate some RUNX1 mutants associated with breast cancer. In nude mice, tumorigenicity was promoted by RUNX3 bearing a mutation in the sumoylation site, but suppressed by wild-type RUNX3. Our results suggest that RUNXs are sumoylated by PIAS1, and that this modification could play a critical role in the regulation of the tumor-suppressive activity of these proteins. PMID:24777122

SUMOylation target sites at the C terminus protect Axin from ubiquitination and confer protein stability

PubMed Central

Kim, Min Jung; Chia, Ian V.; Costantini, Frank

2008-01-01

Axin is a scaffold protein for the β-catenin destruction complex, and a negative regulator of canonical Wnt signaling. Previous studies implicated the six C-terminal amino acids (C6 motif) in the ability of Axin to activate c-Jun N-terminal kinase, and identified them as a SUMOylation target. Deletion of the C6 motif of mouse Axin in vivo reduced the steady-state protein level, which caused embryonic lethality. Here, we report that this deletion (Axin-ΔC6) causes a reduced half-life in mouse embryonic fibroblasts and an increased susceptibility to ubiquitination in HEK 293T cells. We confirmed the C6 motif as a SUMOylation target in vitro, and found that mutating the C-terminal SUMOylation target residues increased the susceptibility of Axin to polyubiquitination and reduced its steady-state level. Heterologous SUMOylation target sites could replace C6 in providing this protective effect. These findings suggest that SUMOylation of the C6 motif may prevent polyubiquitination, thus increasing the stability of Axin. Although C6 deletion also caused increased association of Axin with Dvl-1, this interaction was not altered by mutating the lysine residues in C6, nor could heterologous SUMOylation motifs replace the C6 motif in this assay. Therefore, some other specific property of the C6 motif seems to reduce the interaction of Axin with Dvl-1.—Kim, M. J., Chia, I. V., Costantini, F. SUMOylation target sites at the C terminus protect Axin from ubiquitination and confer protein stability. PMID:18632848

Ubiquitination and sumoylation of the HTLV-2 Tax-2B protein regulate its NF-κB activity: a comparative study with the HTLV-1 Tax-1 protein

PubMed Central

2012-01-01

Background Retroviruses HTLV-1 and HTLV-2 have homologous genomic structures but differ significantly in pathogenicity. HTLV-1 is associated with Adult T cell Leukemia (ATL), whereas infection by HTLV-2 has no association with neoplasia. Transformation of T lymphocytes by HTLV-1 is linked to the capacity of its oncoprotein Tax-1 to alter cell survival and cell cycle control mechanisms. Among these functions, Tax-1-mediated activation of cellular gene expression via the NF-κB pathway depends on Tax-1 post-translational modifications by ubiquitination and sumoylation. The Tax-2 protein of HTLV-2B (Tax-2B) is also modified by ubiquitination and sumoylation and activates the NF-κB pathway to a level similar to that of Tax-1. The present study aims to understand whether ubiquitination and sumoylation modifications are involved in Tax-2B-mediated activation of the NF-κB pathway. Results The comparison of Tax-1 and Tax-2B lysine to arginine substitution mutants revealed conserved patterns and levels of ubiquitination with notable difference in the lysine usage for sumoylation. Neither Tax-1 nor Tax-2B ubiquitination and sumoylation deficient mutants could activate the NF-κB pathway and fusion of ubiquitin or SUMO-1 to the C-terminus of the ubiquitination and sumoylation deficient Tax-2B mutant strikingly restored transcriptional activity. In addition, ubiquitinated forms of Tax-2B colocalized with RelA and IKKγ in prominent cytoplasmic structures associated with the Golgi apparatus, whereas colocalization of Tax-2B with the RelA subunit of NF-κB and the transcriptional coactivator p300 in punctate nuclear structures was dependent on Tax-2B sumoylation, as previously observed for Tax-1. Conclusions Both Tax-1 and Tax-2 activate the NF-κB pathway via similar mechanisms involving ubiquitination and sumoylation. Therefore, the different transforming potential of HTLV-1 and HTLV-2 is unlikely to be related to different modes of activation of the canonical NF-κB pathway

Impairment of Human Immunodeficiency Virus Type-1 Integrase SUMOylation Correlates with an Early Replication Defect*

PubMed Central

Zamborlini, Alessia; Coiffic, Audrey; Beauclair, Guillaume; Delelis, Olivier; Paris, Joris; Koh, Yashuiro; Magne, Fabian; Giron, Marie-Lou; Tobaly-Tapiero, Joelle; Deprez, Eric; Emiliani, Stephane; Engelman, Alan; de Thé, Hugues; Saïb, Ali

2011-01-01

HIV-1 integrase (IN) orchestrates the integration of the reverse transcribed viral cDNA into the host cell genome and participates also in other steps of HIV-1 replication. Cellular and viral factors assist IN in performing its multiple functions, and post-translational modifications contribute to modulate its activities. Here, we show that HIV-1 IN is modified by SUMO proteins and that phylogenetically conserved SUMOylation consensus motifs represent major SUMO acceptor sites. Viruses harboring SUMOylation site IN mutants displayed a replication defect that was mapped during the early stages of infection, before integration but after reverse transcription. Because SUMOylation-defective IN mutants retained WT catalytic activity, we hypothesize that SUMOylation might regulate the affinity of IN for co-factors, contributing to efficient HIV-1 replication. PMID:21454548

Nuclear Receptors in Neurodegenerative Diseases

PubMed Central

Skerrett, Rebecca; Malm, Tarja; Landreth, Gary

2014-01-01

Nuclear receptors have generated substantial interest in the past decade as potential therapeutic targets for the treatment of neurodegenerative disorders. Despite years of effort, effective treatments for progressive neurodegenerative diseases such as Alzheimer’s disease, Parkinson’s disease, Huntington’s disease and ALS remain elusive, making non-classical drug targets such as nuclear receptors an attractive alternative. A substantial literature in mouse models of disease and several clinical trials have investigated the role of nuclear receptors in various neurodegenerative disorders, most prominently AD. These studies have met with mixed results, yet the majority of studies in mouse models report positive outcomes. The mechanisms by which nuclear receptor agonists affect disease pathology remain unclear. Deciphering the complex signaling underlying nuclear receptor action in neurodegenerative diseases is essential for understanding this variability in preclinical studies, and for the successful translation of nuclear receptor agonists into clinical therapies. PMID:24874548

JASSA: a comprehensive tool for prediction of SUMOylation sites and SIMs.

PubMed

Beauclair, Guillaume; Bridier-Nahmias, Antoine; Zagury, Jean-François; Saïb, Ali; Zamborlini, Alessia

2015-11-01

Post-translational modification by the Small Ubiquitin-like Modifier (SUMO) proteins, a process termed SUMOylation, is involved in many fundamental cellular processes. SUMO proteins are conjugated to a protein substrate, creating an interface for the recruitment of cofactors harboring SUMO-interacting motifs (SIMs). Mapping both SUMO-conjugation sites and SIMs is required to study the functional consequence of SUMOylation. To define the best candidate sites for experimental validation we designed JASSA, a Joint Analyzer of SUMOylation site and SIMs. JASSA is a predictor that uses a scoring system based on a Position Frequency Matrix derived from the alignment of experimental SUMOylation sites or SIMs. Compared with existing web-tools, JASSA displays on par or better performances. Novel features were implemented towards a better evaluation of the prediction, including identification of database hits matching the query sequence and representation of candidate sites within the secondary structural elements and/or the 3D fold of the protein of interest, retrievable from deposited PDB files. JASSA is freely accessible at http://www.jassa.fr/. Website is implemented in PHP and MySQL, with all major browsers supported. guillaume.beauclair@inserm.fr Supplementary data are available at Bioinformatics online. © The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

SUMOylation of ATRIP potentiates DNA damage signaling by boosting multiple protein interactions in the ATR pathway

PubMed Central

Wu, Ching-Shyi; Ouyang, Jian; Mori, Eiichiro; Nguyen, Hai Dang; Maréchal, Alexandre; Hallet, Alexander; Chen, David J.; Zou, Lee

2014-01-01

The ATR (ATM [ataxia telangiectasia-mutated]- and Rad3-related) checkpoint is a crucial DNA damage signaling pathway. While the ATR pathway is known to transmit DNA damage signals through the ATR–Chk1 kinase cascade, whether post-translational modifications other than phosphorylation are important for this pathway remains largely unknown. Here, we show that protein SUMOylation plays a key role in the ATR pathway. ATRIP, the regulatory partner of ATR, is modified by SUMO2/3 at K234 and K289. An ATRIP mutant lacking the SUMOylation sites fails to localize to DNA damage and support ATR activation efficiently. Surprisingly, the ATRIP SUMOylation mutant is compromised in the interaction with a protein group, rather than a single protein, in the ATR pathway. Multiple ATRIP-interacting proteins, including ATR, RPA70, TopBP1, and the MRE11–RAD50–NBS1 complex, exhibit reduced binding to the ATRIP SUMOylation mutant in cells and display affinity for SUMO2 chains in vitro, suggesting that they bind not only ATRIP but also SUMO. Fusion of a SUMO2 chain to the ATRIP SUMOylation mutant enhances its interaction with the protein group and partially suppresses its localization and functional defects, revealing that ATRIP SUMOylation promotes ATR activation by providing a unique type of protein glue that boosts multiple protein interactions along the ATR pathway. PMID:24990965

The gap junction channel protein connexin 43 is covalently modified and regulated by SUMOylation.

PubMed

Kjenseth, Ane; Fykerud, Tone A; Sirnes, Solveig; Bruun, Jarle; Yohannes, Zeremariam; Kolberg, Matthias; Omori, Yasufumi; Rivedal, Edgar; Leithe, Edward

2012-05-04

SUMOylation is a posttranslational modification in which a member of the small ubiquitin-like modifier (SUMO) family of proteins is conjugated to lysine residues in specific target proteins. Most known SUMOylation target proteins are located in the nucleus, but there is increasing evidence that SUMO may also be a key determinant of many extranuclear processes. Gap junctions consist of arrays of intercellular channels that provide direct transfer of ions and small molecules between adjacent cells. Gap junction channels are formed by integral membrane proteins called connexins, of which the best-studied isoform is connexin 43 (Cx43). Here we show that Cx43 is posttranslationally modified by SUMOylation. The data suggest that the SUMO system regulates the Cx43 protein level and the level of functional Cx43 gap junctions at the plasma membrane. Cx43 was found to be modified by SUMO-1, -2, and -3. Evidence is provided that the membrane-proximal lysines at positions 144 and 237, located in the Cx43 intracellular loop and C-terminal tail, respectively, act as SUMO conjugation sites. Mutations of lysine 144 or lysine 237 resulted in reduced Cx43 SUMOylation and reduced Cx43 protein and gap junction levels. Altogether, these data identify Cx43 as a SUMOylation target protein and represent the first evidence that gap junctions are regulated by the SUMO system.

System-wide Analysis of SUMOylation Dynamics in Response to Replication Stress Reveals Novel Small Ubiquitin-like Modified Target Proteins and Acceptor Lysines Relevant for Genome Stability*

PubMed Central

Xiao, Zhenyu; Chang, Jer-Gung; Hendriks, Ivo A.; Sigurðsson, Jón Otti; Olsen, Jesper V.; Vertegaal, Alfred C.O.

2015-01-01

Genotoxic agents can cause replication fork stalling in dividing cells because of DNA lesions, eventually leading to replication fork collapse when the damage is not repaired. Small Ubiquitin-like Modifiers (SUMOs) are known to counteract replication stress, nevertheless, only a small number of relevant SUMO target proteins are known. To address this, we have purified and identified SUMO-2 target proteins regulated by replication stress in human cells. The developed methodology enabled single step purification of His10-SUMO-2 conjugates under denaturing conditions with high yield and high purity. Following statistical analysis on five biological replicates, a total of 566 SUMO-2 targets were identified. After 2 h of hydroxyurea treatment, 10 proteins were up-regulated for SUMOylation and two proteins were down-regulated for SUMOylation, whereas after 24 h, 35 proteins were up-regulated for SUMOylation, and 13 proteins were down-regulated for SUMOylation. A site-specific approach was used to map over 1000 SUMO-2 acceptor lysines in target proteins. The methodology is generic and is widely applicable in the ubiquitin field. A large subset of these identified proteins function in one network that consists of interacting replication factors, transcriptional regulators, DNA damage response factors including MDC1, ATR-interacting protein ATRIP, the Bloom syndrome protein and the BLM-binding partner RMI1, the crossover junction endonuclease EME1, BRCA1, and CHAF1A. Furthermore, centromeric proteins and signal transducers were dynamically regulated by SUMOylation upon replication stress. Our results uncover a comprehensive network of SUMO target proteins dealing with replication damage and provide a framework for detailed understanding of the role of SUMOylation to counteract replication stress. Ultimately, our study reveals how a post-translational modification is able to orchestrate a large variety of different proteins to integrate different nuclear processes with the

High glucose induces activation of NF-κB inflammatory signaling through IκBα sumoylation in rat mesangial cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Huang, Wei; Department of Endocrinology, The People’s Hospital of Xindu, Xindu, Sichuang, 610500; Xu, Ling

Highlights: •The expression of SUMO1, SUMO2/3 under high glucose was obviously enhanced. •High glucose induced degradation of IκBα and activation of NF-κB pathway. •Sumoylation of IκBα in high glucose were significantly decreased. •The proteasome inhibitor MG132 could partially revert the degradation of IκBα. -- Abstract: The posttranslational modification of proteins by small ubiquitin-like modifiers (SUMOs) has emerged as an important regulatory mechanism for the alteration of protein activity, stability, and cellular localization. The latest research demonstrates that sumoylation is extensively involved in the regulation of the nuclear factor κB (NF-κB) pathway, which plays a critical role in the regulation ofmore » inflammation and contributes to fibrosis in diabetic nephropathy (DN). However, the role of sumoylation in the regulation of NF-κB signaling in DN is still unclear. In the present study, we cultured rat glomerular mesangial cells (GMCs) stimulated by high glucose and divided GMCs into six groups: normal glucose group (5.6 mmol/L), high glucose groups (10, 20, and 30 mmol/L), mannitol group (i.e., osmotic control group), and MG132 intervention group (30 mmol/L glucose with MG132, a proteasome inhibitor). The expression of SUMO1, SUMO2/3, IκBα, NF-κBp65, and monocyte chemotactic protein 1 (MCP-1) was measured by Western blot, reverse-transcription polymerase chain reaction, and indirect immunofluorescence laser scanning confocal microscopy. The interaction between SUMO1, SUMO2/3, and IκBα was observed by co-immunoprecipitation. The results showed that the expression of SUMO1 and SUMO2/3 was dose- and time-dependently enhanced by high glucose (p < 0.05). However, the expression of IκBα sumoylation in high glucose was significantly decreased compared with the normal glucose group (p < 0.05). The expression of IκBα was dose- and time-dependently decreased, and NF-κBp65 and MCP-1 were increased under high glucose conditions

MeCP2 SUMOylation rescues Mecp2-mutant-induced behavioural deficits in a mouse model of Rett syndrome

PubMed Central

Tai, Derek J. C.; Liu, Yen C.; Hsu, Wei L.; Ma, Yun L.; Cheng, Sin J.; Liu, Shau Y.; Lee, Eminy H. Y.

2016-01-01

The methyl-CpG-binding protein 2 (MeCP2) gene, MECP2, is an X-linked gene encoding the MeCP2 protein, and mutations of MECP2 cause Rett syndrome (RTT). However, the molecular mechanism of MECP2-mutation-caused RTT is less known. Here we find that MeCP2 could be SUMO-modified by the E3 ligase PIAS1 at Lys-412. MeCP2 phosphorylation (at Ser-421 and Thr-308) facilitates MeCP2 SUMOylation, and MeCP2 SUMOylation is induced by NMDA, IGF-1 and CRF in the rat brain. MeCP2 SUMOylation releases CREB from the repressor complex and enhances Bdnf mRNA expression. Several MECP2 mutations identified in RTT patients show decreased MeCP2 SUMOylation. Re-expression of wild-type MeCP2 or SUMO-modified MeCP2 in Mecp2-null neurons rescues the deficits of social interaction, fear memory and LTP observed in Mecp2 conditional knockout (cKO) mice. These results together reveal an important role of MeCP2 SUMOylation in social interaction, memory and synaptic plasticity, and that abnormal MeCP2 SUMOylation is implicated in RTT. PMID:26842955

Overexpression of SKI Oncoprotein Leads to p53 Degradation through Regulation of MDM2 Protein Sumoylation*

PubMed Central

Ding, Boxiao; Sun, Yin; Huang, Jiaoti

2012-01-01

Protooncogene Ski was identified based on its ability to transform avian fibroblasts in vitro. In support of its oncogenic activity, SKI was found to be overexpressed in a variety of human cancers, although the exact molecular mechanism(s) responsible for its oncogenic activity is not fully understood. We found that SKI can negatively regulate p53 by decreasing its level through up-regulation of MDM2 activity, which is mediated by the ability of SKI to enhance sumoylation of MDM2. This stimulation of MDM2 sumoylation is accomplished through a direct interaction of SKI with SUMO-conjugating enzyme E2, Ubc9, resulting in enhanced thioester bond formation and mono-sumoylation of Ubc9. A mutant SKI defective in transformation fails to increase p53 ubiquitination and is unable to increase MDM2 levels and to increase mono-sumoylation of Ubc9, suggesting that the ability of SKI to enhance Ubc9 activity is essential for its transforming function. These results established a detailed molecular mechanism that underlies the ability of SKI to cause cellular transformation while unraveling a novel connection between sumoylation and tumorigenesis, providing potential new therapeutic targets for cancer. PMID:22411991

The defective nuclear lamina in Hutchinson-gilford progeria syndrome disrupts the nucleocytoplasmic Ran gradient and inhibits nuclear localization of Ubc9.

PubMed

Kelley, Joshua B; Datta, Sutirtha; Snow, Chelsi J; Chatterjee, Mandovi; Ni, Li; Spencer, Adam; Yang, Chun-Song; Cubeñas-Potts, Caelin; Matunis, Michael J; Paschal, Bryce M

2011-08-01

The mutant form of lamin A responsible for the premature aging disease Hutchinson-Gilford progeria syndrome (termed progerin) acts as a dominant negative protein that changes the structure of the nuclear lamina. How the perturbation of the nuclear lamina in progeria is transduced into cellular changes is undefined. Using patient fibroblasts and a variety of cell-based assays, we determined that progerin expression in Hutchinson-Gilford progeria syndrome inhibits the nucleocytoplasmic transport of several factors with key roles in nuclear function. We found that progerin reduces the nuclear/cytoplasmic concentration of the Ran GTPase and inhibits the nuclear localization of Ubc9, the sole E2 for SUMOylation, and of TPR, the nucleoporin that forms the basket on the nuclear side of the nuclear pore complex. Forcing the nuclear localization of Ubc9 in progerin-expressing cells rescues the Ran gradient and TPR import, indicating that these pathways are linked. Reducing nuclear SUMOylation decreases the nuclear mobility of the Ran nucleotide exchange factor RCC1 in vivo, and the addition of SUMO E1 and E2 promotes the dissociation of RCC1 and Ran from chromatin in vitro. Our data suggest that the cellular effects of progerin are transduced, at least in part, through reduced function of the Ran GTPase and SUMOylation pathways.

LMP1-Induced Sumoylation Influences the Maintenance of Epstein-Barr Virus Latency through KAP1

PubMed Central

Moss, Charles Randall; Whitehurst, Christopher B.; Moody, Cary A.

2015-01-01

ABSTRACT As a herpesvirus, Epstein-Barr virus (EBV) establishes a latent infection that can periodically undergo reactivation, resulting in lytic replication and the production of new infectious virus. Latent membrane protein-1 (LMP1), the principal viral oncoprotein, is a latency-associated protein implicated in regulating viral reactivation and the maintenance of latency. We recently found that LMP1 hijacks the SUMO-conjugating enzyme Ubc9 via its C-terminal activating region-3 (CTAR3) and induces the sumoylation of cellular proteins. Because protein sumoylation can promote transcriptional repression, we hypothesized that LMP1-induced protein sumoylation induces the repression of EBV lytic promoters and helps maintain the viral genome in its latent state. We now show that with inhibition of LMP1-induced protein sumoylation, the latent state becomes less stable or leakier in EBV-transformed lymphoblastoid cell lines. The cells are also more sensitive to viral reactivation induced by irradiation, which results in the increased production and release of infectious virus, as well as increased susceptibility to ganciclovir treatment. We have identified a target of LMP1-mediated sumoylation that contributes to the maintenance of latency in this context: KRAB-associated protein-1 (KAP1). LMP1 CTAR3-mediated sumoylation regulates the function of KAP1. KAP1 also binds to EBV OriLyt and immediate early promoters in a CTAR3-dependent manner, and inhibition of sumoylation processes abrogates the binding of KAP1 to these promoters. These data provide an additional line of evidence that supports our findings that CTAR3 is a distinct functioning regulatory region of LMP1 and confirm that LMP1-induced sumoylation may help stabilize the maintenance of EBV latency. IMPORTANCE Epstein-Barr virus (EBV) latent membrane protein-1 (LMP1) plays an important role in the maintenance of viral latency. Previously, we documented that LMP1 targets cellular proteins to be modified by a

Global analysis of sumoylation function reveals novel insights into development and appressorium-mediated infection of the rice blast fungus.

PubMed

Liu, Caiyun; Li, Zhigang; Xing, Junjie; Yang, Jun; Wang, Zhao; Zhang, Hong; Chen, Deng; Peng, You-Liang; Chen, Xiao-Lin

2018-04-16

Protein post-translational modifications play critical roles in cellular processes, development and stress response. The small ubiquitin-like modifier (SUMO) to proteins is one of the essential modifications in eukaryotes, but its function remains largely unknown in plant pathogenic fungi. We present a comprehensive analysis combined with proteomic, molecular and cellular approaches to explore the roles of sumoylation in the model plant fungal pathogen, Magnaporthe oryzae. We found the SUMO pathway plays key roles in colony growth, conidia formation and virulence to the host, as well as cell-cycle-related phenotypes. Sumoylation is also involved in responding to different stresses. Affinity purification identified 940 putative SUMO substrates, many of which were reported to be involved in development, stress response and infection. Interestingly, four septins were also shown to be sumoylated. Mutation of consensus sumoylation sites in each septin all resulted in reduced virulence to the host and dislocation of septins in appressoria. Moreover, sumoylation is also involved in extracellular secretion of different effector proteins. Our study on the functions of sumoylation provides novel insight into development and infection of the rice blast fungus. © 2018 The Authors. New Phytologist © 2018 New Phytologist Trust.

Sumoylation promotes optimal APC/C Activation and Timely Anaphase.

PubMed

Lee, Christine C; Li, Bing; Yu, Hongtao; Matunis, Michael J

2018-03-08

The Anaphase Promoting Complex/Cyclosome (APC/C) is a ubiquitin E3 ligase that functions as the gatekeeper to mitotic exit. APC/C activity is controlled by an interplay of multiple pathways during mitosis, including the spindle assembly checkpoint (SAC), that are not yet fully understood. Here, we show that sumoylation of the APC4 subunit of the APC/C peaks during mitosis and is critical for timely APC/C activation and anaphase onset. We have also identified a functionally important SUMO interacting motif in the cullin-homology domain of APC2 located near the APC4 sumoylation sites and APC/C catalytic core. Our findings provide evidence of an important regulatory role for SUMO modification and binding in affecting APC/C activation and mitotic exit. © 2018, Lee et al.

Targeting Nuclear Receptors with Marine Natural Products

PubMed Central

Yang, Chunyan; Li, Qianrong; Li, Yong

2014-01-01

Nuclear receptors (NRs) are important pharmaceutical targets because they are key regulators of many metabolic and inflammatory diseases, including diabetes, dyslipidemia, cirrhosis, and fibrosis. As ligands play a pivotal role in modulating nuclear receptor activity, the discovery of novel ligands for nuclear receptors represents an interesting and promising therapeutic approach. The search for novel NR agonists and antagonists with enhanced selectivities prompted the exploration of the extraordinary chemical diversity associated with natural products. Recent studies involving nuclear receptors have disclosed a number of natural products as nuclear receptor ligands, serving to re-emphasize the translational possibilities of natural products in drug discovery. In this review, the natural ligands of nuclear receptors will be described with an emphasis on their mechanisms of action and their therapeutic potentials, as well as on strategies to determine potential marine natural products as nuclear receptor modulators. PMID:24473166

The Orphan Nuclear Receptor TR4 Is a Vitamin A-activated Nuclear Receptor

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zhou, X. Edward; Suino-Powell, Kelly M.; Xu, Yong

2015-11-30

Testicular receptors 2 and 4 (TR2/4) constitute a subgroup of orphan nuclear receptors that play important roles in spermatogenesis, lipid and lipoprotein regulation, and the development of the central nervous system. Currently, little is known about the structural features and the ligand regulation of these receptors. Here we report the crystal structure of the ligand-free TR4 ligand binding domain, which reveals an autorepressed conformation. The ligand binding pocket of TR4 is filled by the C-terminal half of helix 10, and the cofactor binding site is occupied by the AF-2 helix, thus preventing ligand-independent activation of the receptor. However, TR4 exhibitsmore » constitutive transcriptional activity on multiple promoters, which can be further potentiated by nuclear receptor coactivators. Mutations designed to disrupt cofactor binding, dimerization, or ligand binding substantially reduce the transcriptional activity of this receptor. Importantly, both retinol and retinoic acid are able to promote TR4 to recruit coactivators and to activate a TR4-regulated reporter. These findings demonstrate that TR4 is a ligand-regulated nuclear receptor and suggest that retinoids might have a much wider regulatory role via activation of orphan receptors such as TR4.« less

The Defective Nuclear Lamina in Hutchinson-Gilford Progeria Syndrome Disrupts the Nucleocytoplasmic Ran Gradient and Inhibits Nuclear Localization of Ubc9▿

PubMed Central

Kelley, Joshua B.; Datta, Sutirtha; Snow, Chelsi J.; Chatterjee, Mandovi; Ni, Li; Spencer, Adam; Yang, Chun-Song; Cubeñas-Potts, Caelin; Matunis, Michael J.; Paschal, Bryce M.

2011-01-01

The mutant form of lamin A responsible for the premature aging disease Hutchinson-Gilford progeria syndrome (termed progerin) acts as a dominant negative protein that changes the structure of the nuclear lamina. How the perturbation of the nuclear lamina in progeria is transduced into cellular changes is undefined. Using patient fibroblasts and a variety of cell-based assays, we determined that progerin expression in Hutchinson-Gilford progeria syndrome inhibits the nucleocytoplasmic transport of several factors with key roles in nuclear function. We found that progerin reduces the nuclear/cytoplasmic concentration of the Ran GTPase and inhibits the nuclear localization of Ubc9, the sole E2 for SUMOylation, and of TPR, the nucleoporin that forms the basket on the nuclear side of the nuclear pore complex. Forcing the nuclear localization of Ubc9 in progerin-expressing cells rescues the Ran gradient and TPR import, indicating that these pathways are linked. Reducing nuclear SUMOylation decreases the nuclear mobility of the Ran nucleotide exchange factor RCC1 in vivo, and the addition of SUMO E1 and E2 promotes the dissociation of RCC1 and Ran from chromatin in vitro. Our data suggest that the cellular effects of progerin are transduced, at least in part, through reduced function of the Ran GTPase and SUMOylation pathways. PMID:21670151

Smad4 SUMOylation is essential for memory formation through upregulation of the skeletal myopathy gene TPM2.

PubMed

Hsu, Wei L; Ma, Yun L; Liu, Yen C; Lee, Eminy H Y

2017-11-28

Smad4 is a critical effector of TGF-β signaling that regulates a variety of cellular functions. However, its role in the brain has rarely been studied. Here, we examined the molecular mechanisms underlying the post-translational regulation of Smad4 function by SUMOylation, and its role in spatial memory formation. In the hippocampus, Smad4 is SUMOylated by the E3 ligase PIAS1 at Lys-113 and Lys-159. Both spatial training and NMDA injection enhanced Smad4 SUMOylation. Inhibition of Smad4 SUMOylation impaired spatial learning and memory in rats by downregulating TPM2, a gene associated with skeletal myopathies. Similarly, knockdown of TPM2 expression impaired spatial learning and memory, while TPM2 mRNA and protein expression increased after spatial training. Among the TPM2 mutations associated with skeletal myopathies, the TPM2E122K mutation was found to reduce TPM2 expression and impair spatial learning and memory in rats. We have identified a novel role of Smad4 SUMOylation and TPM2 in learning and memory formation. These results suggest that patients with skeletal myopathies who carry the TPM2E122K mutation may also have deficits in learning and memory functions.

SUMOylation pathway alteration coupled with downregulation of SUMO E2 enzyme at mucosal epithelium modulates inflammation in inflammatory bowel disease

PubMed Central

Mustfa, Salman Ahmad; Singh, Mukesh; Suhail, Aamir; Mohapatra, Gayatree; Verma, Smriti; Chakravorty, Debangana; Rana, Sarika; Rampal, Ritika; Dhar, Atika; Saha, Sudipto; Ahuja, Vineet

2017-01-01

Post-translational modification pathways such as SUMOylation are integral to all cellular processes and tissue homeostasis. We investigated the possible involvement of SUMOylation in the epithelial signalling in Crohn's disease (CD) and ulcerative colitis (UC), the two major forms of inflammatory bowel disease (IBD). Initially in a murine model of IBD, induced by dextran–sulfate–sodium (DSS mice), we observed inflammation accompanied by a lowering of global SUMOylation of colonic epithelium. The observed SUMOylation alteration was due to a decrease in the sole SUMO E2 enzyme (Ubc9). Mass-spectrometric analysis revealed the existence of a distinct SUMOylome (SUMO-conjugated proteome) in DSS mice with alteration of key cellular regulators, including master kinase Akt1. Knocking-down of Ubc9 in epithelial cells resulted in dramatic activation of inflammatory gene expression, a phenomenon that acted via reduction in Akt1 and its SUMOylated form. Importantly, a strong decrease in Ubc9 and Akt1 was also seen in endoscopic biopsy samples (N = 66) of human CD and UC patients. Furthermore, patients with maximum disease indices were always accompanied by severely lowered Ubc9 or SUMOylated-Akt1. Mucosal tissues with severely compromised Ubc9 function displayed higher levels of pro-inflammatory cytokines and compromised wound-healing markers. Thus, our results reveal an important and previously undescribed role for the SUMOylation pathway involving Ubc9 and Akt1 in modulation of epithelial inflammatory signalling in IBD. PMID:28659381

Targeted Identification of SUMOylation Sites in Human Proteins Using Affinity Enrichment and Paralog-specific Reporter Ions*

PubMed Central

Lamoliatte, Frederic; Bonneil, Eric; Durette, Chantal; Caron-Lizotte, Olivier; Wildemann, Dirk; Zerweck, Johannes; Wenshuk, Holger; Thibault, Pierre

2013-01-01

Protein modification by small ubiquitin-like modifier (SUMO) modulates the activities of numerous proteins involved in different cellular functions such as gene transcription, cell cycle, and DNA repair. Comprehensive identification of SUMOylated sites is a prerequisite to determine how SUMOylation regulates protein function. However, mapping SUMOylated Lys residues by mass spectrometry (MS) is challenging because of the dynamic nature of this modification, the existence of three functionally distinct human SUMO paralogs, and the large SUMO chain remnant that remains attached to tryptic peptides. To overcome these problems, we created HEK293 cell lines that stably express functional SUMO paralogs with an N-terminal His6-tag and an Arg residue near the C terminus that leave a short five amino acid SUMO remnant upon tryptic digestion. We determined the fragmentation patterns of our short SUMO remnant peptides by collisional activation and electron transfer dissociation using synthetic peptide libraries. Activation using higher energy collisional dissociation on the LTQ-Orbitrap Elite identified SUMO paralog-specific fragment ions and neutral losses of the SUMO remnant with high mass accuracy (< 5 ppm). We exploited these features to detect SUMO modified tryptic peptides in complex cell extracts by correlating mass measurements of precursor and fragment ions using a data independent acquisition method. We also generated bioinformatics tools to retrieve MS/MS spectra containing characteristic fragment ions to the identification of SUMOylated peptide by conventional Mascot database searches. In HEK293 cell extracts, this MS approach uncovered low abundance SUMOylated peptides and 37 SUMO3-modified Lys residues in target proteins, most of which were previously unknown. Interestingly, we identified mixed SUMO-ubiquitin chains with ubiquitylated SUMO proteins (K20 and K32) and SUMOylated ubiquitin (K63), suggesting a complex crosstalk between these two modifications. PMID

The Spindle Positioning Protein Kar9p Interacts With the Sumoylation Machinery in Saccharomyces cerevisiae

PubMed Central

Meednu, Nida; Hoops, Harold; D'Silva, Sonia; Pogorzala, Leah; Wood, Schuyler; Farkas, David; Sorrentino, Mark; Sia, Elaine; Meluh, Pam; Miller, Rita K.

2008-01-01

Accurate positioning of the mitotic spindle is important for the genetic material to be distributed evenly in dividing cells, but little is known about the mechanisms that regulate this process. Here we report that two microtubule-associated proteins important for spindle positioning interact with several proteins in the sumoylation pathway. By two-hybrid analysis, Kar9p and Bim1p interact with the yeast SUMO Smt3p, the E2 enzyme Ubc9p, an E3 Nfi1p, as well as Wss1p, a weak suppressor of a temperature-sensitive smt3 allele. The physical interaction between Kar9p and Ubc9p was confirmed by in vitro binding assays. A single-amino-acid substitution in Kar9p, L304P disrupted its two-hybrid interaction with proteins in the sumoylation pathway, but retained its interactions with the spindle positioning proteins Bim1p, Stu2p, Bik1p, and Myo2p. The kar9-L304P mutant showed defects in positioning the mitotic spindle, with the spindle located more distally than normal. Whereas wild-type Kar9p-3GFP normally localizes to only the bud-directed spindle pole body (SPB), Kar9p-L304P-3GFP was mislocalized to both SPBs. Using a reconstitution assay, Kar9p was sumoylated in vitro. We propose a model in which sumoylation regulates spindle positioning by restricting Kar9p to one SPB. These findings raise the possibility that sumoylation could regulate other microtubule-dependent processes. PMID:18832349

Mediator-dependent Nuclear Receptor Functions

PubMed Central

Chen, Wei; Roeder, Robert

2011-01-01

As gene-specific transcription factors, nuclear hormone receptors are broadly involved in many important biological processes. Their function on target genes requires the stepwise assembly of different coactivator complexes that facilitate chromatin remodeling and subsequent preinitiation complex (PIC) formation and function. Mediator has proved to be a crucial, and general, nuclear receptor-interacting coactivator, with demonstrated functions in transcription steps ranging from chromatin remodeling to subsequent PIC formation and function. Here we discuss (i) our current understanding of pathways that nuclear receptors and other interacting cofactors employ to recruit Mediator to target gene enhancers and promoters, including conditional requirements for the strong NR-Mediator interactions mediated by the NR AF2 domain and the MED1 LXXLLL motifs and (ii) mechanisms by which Mediator acts to transmit signals from enhancer-bound nuclear receptors to the general transcription machinery at core promoters to effect PIC formation and function. PMID:21854863

Nuclear receptors in bile acid metabolism

PubMed Central

Li, Tiangang; Chiang, John Y. L.

2013-01-01

Bile acids are signaling molecules that activate nuclear receptors, such as farnesoid X receptor, pregnane X receptor, constitutive androstane receptor, and vitamin D receptor, and play a critical role in the regulation of lipid, glucose, energy, and drug metabolism. These xenobiotic/endobiotic-sensing nuclear receptors regulate phase I oxidation, phase II conjugation, and phase III transport in bile acid and drug metabolism in the digestive system. Integration of bile acid metabolism with drug metabolism controls absorption, transport, and metabolism of nutrients and drugs to maintain metabolic homeostasis and also protects against liver injury, inflammation, and related metabolic diseases, such as nonalcoholic fatty liver disease, diabetes, and obesity. Bile-acid–based drugs targeting nuclear receptors are in clinical trials for treating cholestatic liver diseases and fatty liver disease. PMID:23330546

SUMOylation of DRIL1 Directs Its Transcriptional Activity Towards Leukocyte Lineage-Specific Genes

PubMed Central

van Lohuizen, Maarten; Peeper, Daniel S.

2009-01-01

DRIL1 is an ARID family transcription factor that can immortalize primary mouse fibroblasts, bypass RASV12-induced cellular senescence and collaborate with RASV12 or MYC in mediating oncogenic transformation. It also activates immunoglobulin heavy chain transcription and engages in heterodimer formation with E2F to stimulate E2F-dependent transcription. Little, however, is known about the regulation of DRIL1 activity. Recently, DRIL1 was found to interact with the SUMO-conjugating enzyme Ubc9, but the functional relevance of this association has not been assessed. Here, we show that DRIL1 is sumoylated both in vitro and in vivo at lysine 398. Moreover, we provide evidence that PIASy functions as a specific SUMO E3-ligase for DRIL1 and promotes its sumoylation both in vitro and in vivo. Furthermore, consistent with the subnuclear localization of PIASy in the Matrix-Associated Region (MAR), SUMO-modified DRIL1 species are found exclusively in the MAR fraction. This post-translational modification interferes neither with the subcellular localization nor the DNA-binding activity of the protein. In contrast, DRIL1 sumoylation impairs its interaction with E2F1 in vitro and modifies its transcriptional activity in vivo, driving transcription of subset of genes regulating leukocyte fate. Taken together, these results identify sumoylation as a novel post-translational modification of DRIL1 that represents an important mechanism for targeting and modulating DRIL1 transcriptional activity. PMID:19436740

Nuclear pool of phosphatidylinositol 4 phosphate 5 kinase 1α is modified by polySUMO-2 during apoptosis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chakrabarti, Rajarshi; Bhowmick, Debajit; Bhargava, Varsha

2013-09-20

Highlights: •Nuclear pool of PIP5K is SUMOylated. •Enhancement of SUMOylated nuclear PIP5K during apoptosis. •Nuclear PIP5K is modified by polySUMO-1 during apoptosis. •Nuclear PIP5K is modified by polySUMO-2 chain during apoptosis. -- Abstract: Phosphatidylinositol 4 phosphate 5 kinase 1α (PIP5K) is mainly localized in the cytosol and plasma membrane. Studies have also indicated its prominent association with nuclear speckles. The exact nature of this nuclear pool of PIP5K is not clear. Using biochemical and microscopic techniques, we have demonstrated that the nuclear pool of PIP5K is modified by SUMO-1 in HEK-293 cells stably expressing PIP5K. Moreover, this SUMOylated pool ofmore » PIP5K increased during apoptosis. PolySUMO-2 chain conjugated PIP5K was detected by pull-down experiment using affinity-tagged RNF4, a polySUMO-2 binding protein, during late apoptosis.« less

Temporal and SUMO-specific SUMOylation contribute to the dynamics of Polo-like kinase 1 (PLK1) and spindle integrity during mouse oocyte meiosis.

PubMed

Feitosa, Weber Beringui; Hwang, KeumSil; Morris, Patricia L

2018-02-15

During mammalian meiosis, Polo-like kinase 1 (PLK1) is essential during cell cycle progression. In oocyte maturation, PLK1 expression is well characterized but timing of posttranslational modifications regulating its activity and subcellular localization are less clear. Small ubiquitin-related modifier (SUMO) posttranslational modifier proteins have been detected in mammalian gametes but their precise function during gametogenesis is largely unknown. In the present paper we report for mouse oocytes that both PLK1 and phosphorylated PLK1 undergo SUMOylation in meiosis II (MII) oocytes using immunocytochemistry, immunoprecipitation and in vitro SUMOylation assays. At MII, PLK1 is phosphorylated at threonine-210 and serine-137. MII oocyte PLK1 and phosphorylated PLK1 undergo SUMOylation by SUMO-1, -2 and -3 as shown by individual in vitro assays. Using these assays, forms of phosphorylated PLK1 normalized to PLK1 increased significantly and correlated with SUMOylated PLK1 levels. During meiotic progression and maturation, SUMO-1-SUMOylation of PLK1 is involved in spindle formation whereas SUMO-2/3-SUMOylation may regulate PLK1 activity at kinetochore-spindle attachment sites. Microtubule integrity is required for PLK1 localization with SUMO-1 but not with SUMO-2/3. Inhibition of SUMOylation disrupts proper meiotic bipolar spindle organization and spindle-kinetochore attachment. The data show that both temporal and SUMO-specific-SUMOylation play important roles in orchestrating functional dynamics of PLK1 during mouse oocyte meiosis, including subcellular compartmentalization. Copyright © 2018 Elsevier Inc. All rights reserved.

SUMOylation Promotes PML Degradation during Encephalomyocarditis Virus Infection▿

PubMed Central

El Mchichi, Bouchra; Regad, Tarik; Maroui, Mohamed-Ali; Rodriguez, Manuel S.; Aminev, Aleksey; Gerbaud, Sylvie; Escriou, Nicolas; Dianoux, Laurent; Chelbi-Alix, Mounira K.

2010-01-01

The promyelocytic leukemia (PML) protein is expressed in the diffuse nuclear fraction of the nucleoplasm and in matrix-associated structures, known as nuclear bodies (NBs). PML NB formation requires the covalent modification of PML to SUMO. The noncovalent interactions of SUMO with PML based on the identification of a SUMO-interacting motif within PML seem to be required for further recruitment within PML NBs of SUMOylated proteins. RNA viruses whose replication takes place in the cytoplasm and is inhibited by PML have developed various strategies to counteract the antiviral defense mediated by PML NBs. We show here that primary fibroblasts derived from PML knockout mice are more sensitive to infection with encephalomyocarditis virus (EMCV), suggesting that the absence of PML results in an increase in EMCV replication. Also, we found that EMCV induces a decrease in PML protein levels both in interferon-treated cells and in PMLIII-expressing cells. Reduction of PML was carried out by the EMCV 3C protease. Indeed, at early times postinfection, EMCV induced PML transfer from the nucleoplasm to the nuclear matrix and PML conjugation to SUMO-1, SUMO-2, and SUMO-3, leading to an increase in PML body size where the viral protease 3C and the proteasome component were found colocalizing with PML within the NBs. This process was followed by PML degradation occurring in a proteasome- and SUMO-dependent manner and did not involve the SUMO-interacting motif of PML. Together, these findings reveal a new mechanism evolved by EMCV to antagonize the PML pathway in the interferon-induced antiviral defense. PMID:20826694

Nuclear Receptors, RXR, and the Big Bang.

PubMed

Evans, Ronald M; Mangelsdorf, David J

2014-03-27

Isolation of genes encoding the receptors for steroids, retinoids, vitamin D, and thyroid hormone and their structural and functional analysis revealed an evolutionarily conserved template for nuclear hormone receptors. This discovery sparked identification of numerous genes encoding related proteins, termed orphan receptors. Characterization of these orphan receptors and, in particular, of the retinoid X receptor (RXR) positioned nuclear receptors at the epicenter of the "Big Bang" of molecular endocrinology. This Review provides a personal perspective on nuclear receptors and explores their integrated and coordinated signaling networks that are essential for multicellular life, highlighting the RXR heterodimer and its associated ligands and transcriptional mechanism. Copyright © 2014 Elsevier Inc. All rights reserved.

Sumoylation of Rap1 mediates the recruitment of TFIID to promote transcription of ribosomal protein genes

PubMed Central

Chymkowitch, Pierre; Nguéa P, Aurélie; Aanes, Håvard; Koehler, Christian J.; Thiede, Bernd; Lorenz, Susanne; Meza-Zepeda, Leonardo A.; Klungland, Arne; Enserink, Jorrit M.

2015-01-01

Transcription factors are abundant Sumo targets, yet the global distribution of Sumo along the chromatin and its physiological relevance in transcription are poorly understood. Using Saccharomyces cerevisiae, we determined the genome-wide localization of Sumo along the chromatin. We discovered that Sumo-enriched genes are almost exclusively involved in translation, such as tRNA genes and ribosomal protein genes (RPGs). Genome-wide expression analysis showed that Sumo positively regulates their transcription. We also discovered that the Sumo consensus motif at RPG promoters is identical to the DNA binding motif of the transcription factor Rap1. We demonstrate that Rap1 is a molecular target of Sumo and that sumoylation of Rap1 is important for cell viability. Furthermore, Rap1 sumoylation promotes recruitment of the basal transcription machinery, and sumoylation of Rap1 cooperates with the target of rapamycin kinase complex 1 (TORC1) pathway to promote RPG transcription. Strikingly, our data reveal that sumoylation of Rap1 functions in a homeostatic feedback loop that sustains RPG transcription during translational stress. Taken together, Sumo regulates the cellular translational capacity by promoting transcription of tRNA genes and RPGs. PMID:25800674

Nuclear receptors and nonalcoholic fatty liver disease1

PubMed Central

Cave, Matthew C.; Clair, Heather B.; Hardesty, Josiah E.; Falkner, K. Cameron; Feng, Wenke; Clark, Barbara J.; Sidey, Jennifer; Shi, Hongxue; Aqel, Bashar A.; McClain, Craig J.; Prough, Russell A.

2016-01-01

Nuclear receptors are transcription factors which sense changing environmental or hormonal signals and effect transcriptional changes to regulate core life functions including growth, development, and reproduction. To support this function, following ligand-activation by xenobiotics, members of subfamily 1 nuclear receptors (NR1s) may heterodimerize with the retinoid X receptor (RXR) to regulate transcription of genes involved in energy and xenobiotic metabolism and inflammation. Several of these receptors including the peroxisome proliferator-activated receptors (PPARs), the pregnane and xenobiotic receptor (PXR), the constitutive androstane receptor (CAR), the liver X receptor (LXR) and the farnesoid X receptor (FXR) are key regulators of the gut:liver:adipose axis and serve to coordinate metabolic responses across organ systems between the fed and fasting states. Non-alcoholic fatty liver disease (NAFLD) is the most common liver disease and may progress to cirrhosis and even hepatocellular carcinoma. NAFLD is associated with inappropriate nuclear receptor function and perturbations along the gut:liver:adipose axis including obesity, increased intestinal permeability with systemic inflammation, abnormal hepatic lipid metabolism, and insulin resistance. Environmental chemicals may compound the problem by directly interacting with nuclear receptors leading to metabolic confusion and the inability to differentiate fed from fasting conditions. This review focuses on the impact of nuclear receptors in the pathogenesis and treatment of NAFLD. Clinical trials including PIVENS and FLINT demonstrate that nuclear receptor targeted therapies may lead to the paradoxical dissociation of steatosis, inflammation, fibrosis, insulin resistance, dyslipidemia and obesity. Novel strategies currently under development (including tissue-specific ligands and dual receptor agonists) may be required to separate the beneficial effects of nuclear receptor activation from unwanted metabolic

PHD domain-mediated E3 ligase activity directs intramolecular sumoylation of an adjacent bromodomain required for gene silencing.

PubMed

Ivanov, Alexey V; Peng, Hongzhuang; Yurchenko, Vyacheslav; Yap, Kyoko L; Negorev, Dmitri G; Schultz, David C; Psulkowski, Elyse; Fredericks, William J; White, David E; Maul, Gerd G; Sadofsky, Moshe J; Zhou, Ming-Ming; Rauscher, Frank J

2007-12-14

Tandem PHD and bromodomains are often found in chromatin-associated proteins and have been shown to cooperate in gene silencing. Each domain can bind specifically modified histones: the mechanisms of cooperation between these domains are unknown. We show that the PHD domain of the KAP1 corepressor functions as an intramolecular E3 ligase for sumoylation of the adjacent bromodomain. The RING finger-like structure of the PHD domain is required for both Ubc9 binding and sumoylation and directs modification to specific lysine residues in the bromodomain. Sumoylation is required for KAP1-mediated gene silencing and functions by directly recruiting the SETDB1 histone methyltransferase and the CHD3/Mi2 component of the NuRD complex via SUMO-interacting motifs. Sumoylated KAP1 stimulates the histone methyltransferase activity of SETDB1. These data provide a mechanistic explanation for the cooperation of PHD and bromodomains in gene regulation and describe a function of the PHD domain as an intramolecular E3 SUMO ligase.

SUMOylation is developmentally regulated and required for cell pairing during conjugation in Tetrahymena thermophila.

PubMed

Nasir, Amjad M; Yang, Qianyi; Chalker, Douglas L; Forney, James D

2015-02-01

The covalent attachment of small ubiquitin-like modifier (SUMO) to target proteins regulates numerous nuclear events in eukaryotes, including transcription, mitosis and meiosis, and DNA repair. Despite extensive interest in nuclear pathways within the field of ciliate molecular biology, there have been no investigations of the SUMO pathway in Tetrahymena. The developmental program of sexual reproduction of this organism includes cell pairing, micronuclear meiosis, and the formation of a new somatic macronucleus. We identified the Tetrahymena thermophila SMT3 (SUMO) and UBA2 (SUMO-activating enzyme) genes and demonstrated that the corresponding green fluorescent protein (GFP) tagged gene products are found predominantly in the somatic macronucleus during vegetative growth. Use of an anti-Smt3p antibody to perform immunoblot assays with whole-cell lysates during conjugation revealed a large increase in SUMOylation that peaked during formation of the new macronucleus. Immunofluorescence using the same antibody showed that the increase was localized primarily within the new macronucleus. To initiate functional analysis of the SUMO pathway, we created germ line knockout cell lines for both the SMT3 and UBA2 genes and found both are essential for cell viability. Conditional Smt3p and Uba2p cell lines were constructed by incorporation of the cadmium-inducible metallothionein promoter. Withdrawal of cadmium resulted in reduced cell growth and increased sensitivity to DNA-damaging agents. Interestingly, Smt3p and Uba2p conditional cell lines were unable to pair during sexual reproduction in the absence of cadmium, consistent with a function early in conjugation. Our studies are consistent with multiple roles for SUMOylation in Tetrahymena, including a dynamic regulation associated with the sexual life cycle. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Phospholipid Regulation of the Nuclear Receptor Superfamily

PubMed Central

Crowder, Mark K.; Seacrist, Corey D.; Blind, Raymond D.

2016-01-01

Nuclear receptors are ligand-activated transcription factors whose diverse biological functions are classically regulated by cholesterol-based small molecules. Over the past few decades, a growing body of evidence has demonstrated that phospholipids and other similar amphipathic molecules can also specifically bind and functionally regulate the activity of certain nuclear receptors, suggesting a critical role for these non-cholesterol-based molecules in transcriptional regulation. Phosphatidylcholines, phosphoinositides and sphingolipids are a few of the many phospholipid like molecules shown to quite specifically regulate nuclear receptors in mouse models, cell lines and in vitro. More recent evidence has also shown that certain nuclear receptors can “present” a bound phospholipid headgroup to key lipid signaling enzymes, which can then modify the phospholipid headgroup with very unique kinetic properties. Here, we review the broad array of phospholipid / nuclear receptor interactions, from the perspective of the chemical nature of the phospholipid, and the cellular abundance of the phospholipid. We also view the data in the light of well established paradigms for phospholipid mediated transcriptional regulation, as well as newer models of how phospholipids might effect transcription in the acute regulation of complex nuclear signaling pathways. Thus, this review provides novel insight into the new, non-membrane associated roles nuclear phospholipids play in regulating complex nuclear events, centered on the nuclear receptor superfamily of transcription factors. PMID:27838257

Sumoylation of Rap1 mediates the recruitment of TFIID to promote transcription of ribosomal protein genes.

PubMed

Chymkowitch, Pierre; Nguéa, Aurélie P; Aanes, Håvard; Koehler, Christian J; Thiede, Bernd; Lorenz, Susanne; Meza-Zepeda, Leonardo A; Klungland, Arne; Enserink, Jorrit M

2015-06-01

Transcription factors are abundant Sumo targets, yet the global distribution of Sumo along the chromatin and its physiological relevance in transcription are poorly understood. Using Saccharomyces cerevisiae, we determined the genome-wide localization of Sumo along the chromatin. We discovered that Sumo-enriched genes are almost exclusively involved in translation, such as tRNA genes and ribosomal protein genes (RPGs). Genome-wide expression analysis showed that Sumo positively regulates their transcription. We also discovered that the Sumo consensus motif at RPG promoters is identical to the DNA binding motif of the transcription factor Rap1. We demonstrate that Rap1 is a molecular target of Sumo and that sumoylation of Rap1 is important for cell viability. Furthermore, Rap1 sumoylation promotes recruitment of the basal transcription machinery, and sumoylation of Rap1 cooperates with the target of rapamycin kinase complex 1 (TORC1) pathway to promote RPG transcription. Strikingly, our data reveal that sumoylation of Rap1 functions in a homeostatic feedback loop that sustains RPG transcription during translational stress. Taken together, Sumo regulates the cellular translational capacity by promoting transcription of tRNA genes and RPGs. © 2015 Chymkowitch et al.; Published by Cold Spring Harbor Laboratory Press.

Evolution of the nuclear receptor gene superfamily.

PubMed Central

Laudet, V; Hänni, C; Coll, J; Catzeflis, F; Stéhelin, D

1992-01-01

Nuclear receptor genes represent a large family of genes encoding receptors for various hydrophobic ligands such as steroids, vitamin D, retinoic acid and thyroid hormones. This family also contains genes encoding putative receptors for unknown ligands. Nuclear receptor gene products are composed of several domains important for transcriptional activation, DNA binding (C domain), hormone binding and dimerization (E domain). It is not known whether these genes have evolved through gene duplication from a common ancestor or if their different domains came from different independent sources. To test these possibilities we have constructed and compared the phylogenetic trees derived from two different domains of 30 nuclear receptor genes. The tree built from the DNA binding C domain clearly shows a common progeny of all nuclear receptors, which can be grouped into three subfamilies: (i) thyroid hormone and retinoic acid receptors, (ii) orphan receptors and (iii) steroid hormone receptors. The tree constructed from the central part of the E domain which is implicated in transcriptional regulation and dimerization shows the same distribution in three subfamilies but two groups of receptors are in a different position from that in the C domain tree: (i) the Drosophila knirps family genes have acquired very different E domains during evolution, and (ii) the vitamin D and ecdysone receptors, as well as the FTZ-F1 and the NGF1B genes, seem to have DNA binding and hormone binding domains belonging to different classes. These data suggest a complex evolutionary history for nuclear receptor genes in which gene duplication events and swapping between domains of different origins took place. PMID:1312460

IRAK2 directs stimulus-dependent nuclear export of inflammatory mRNAs

PubMed Central

Yu, Minjia; Qian, Wen; Wang, Han; Zhou, Gao; Chen, Xing; Yang, Hui; Hong, Lingzi; Zhao, Junjie; Qin, Luke; Fukuda, Koichi; Flotho, Annette; Gao, Ji; Dongre, Ashok; Carman, Julie A; Kang, Zizhen; Su, Bing; Kern, Timothy S; Smith, Jonathan D; Hamilton, Thomas A; Melchior, Frauke; Fox, Paul L

2017-01-01

Expression of inflammatory genes is determined in part by post-transcriptional regulation of mRNA metabolism but how stimulus- and transcript-dependent nuclear export influence is poorly understood. Here, we report a novel pathway in which LPS/TLR4 engagement promotes nuclear localization of IRAK2 to facilitate nuclear export of a specific subset of inflammation-related mRNAs for translation in murine macrophages. IRAK2 kinase activity is required for LPS-induced RanBP2-mediated IRAK2 sumoylation and subsequent nuclear translocation. Array analysis showed that an SRSF1-binding motif is enriched in mRNAs dependent on IRAK2 for nuclear export. Nuclear IRAK2 phosphorylates SRSF1 to reduce its binding to target mRNAs, which promotes the RNA binding of the nuclear export adaptor ALYREF and nuclear export receptor Nxf1 loading for the export of the mRNAs. In summary, LPS activates a nuclear function of IRAK2 that facilitates the assembly of nuclear export machinery to export selected inflammatory mRNAs to the cytoplasm for translation. PMID:28990926

IRAK2 directs stimulus-dependent nuclear export of inflammatory mRNAs.

PubMed

Zhou, Hao; Bulek, Katarzyna; Li, Xiao; Herjan, Tomasz; Yu, Minjia; Qian, Wen; Wang, Han; Zhou, Gao; Chen, Xing; Yang, Hui; Hong, Lingzi; Zhao, Junjie; Qin, Luke; Fukuda, Koichi; Flotho, Annette; Gao, Ji; Dongre, Ashok; Carman, Julie A; Kang, Zizhen; Su, Bing; Kern, Timothy S; Smith, Jonathan D; Hamilton, Thomas A; Melchior, Frauke; Fox, Paul L; Li, Xiaoxia

2017-10-09

Expression of inflammatory genes is determined in part by post-transcriptional regulation of mRNA metabolism but how stimulus- and transcript-dependent nuclear export influence is poorly understood. Here, we report a novel pathway in which LPS/TLR4 engagement promotes nuclear localization of IRAK2 to facilitate nuclear export of a specific subset of inflammation-related mRNAs for translation in murine macrophages. IRAK2 kinase activity is required for LPS-induced RanBP2-mediated IRAK2 sumoylation and subsequent nuclear translocation. Array analysis showed that an SRSF1-binding motif is enriched in mRNAs dependent on IRAK2 for nuclear export. Nuclear IRAK2 phosphorylates SRSF1 to reduce its binding to target mRNAs, which promotes the RNA binding of the nuclear export adaptor ALYREF and nuclear export receptor Nxf1 loading for the export of the mRNAs. In summary, LPS activates a nuclear function of IRAK2 that facilitates the assembly of nuclear export machinery to export selected inflammatory mRNAs to the cytoplasm for translation.

A New Family of Nuclear Receptor Coregulators That Integrate Nuclear Receptor Signaling through CREB-Binding Protein

PubMed Central

Mahajan, Muktar A.; Samuels, Herbert H.

2000-01-01

We describe the cloning and characterization of a new family of nuclear receptor coregulators (NRCs) which modulate the function of nuclear hormone receptors in a ligand-dependent manner. NRCs are expressed as alternatively spliced isoforms which may exhibit different intrinsic activities and receptor specificities. The NRCs are organized into several modular structures and contain a single functional LXXLL motif which associates with members of the steroid hormone and thyroid hormone/retinoid receptor subfamilies with high affinity. Human NRC (hNRC) harbors a potent N-terminal activation domain (AD1), which is as active as the herpesvirus VP16 activation domain, and a second activation domain (AD2) which overlaps with the receptor-interacting LXXLL region. The C-terminal region of hNRC appears to function as an inhibitory domain which influences the overall transcriptional activity of the protein. Our results suggest that NRC binds to liganded receptors as a dimer and this association leads to a structural change in NRC resulting in activation. hNRC binds CREB-binding protein (CBP) with high affinity in vivo, suggesting that hNRC may be an important functional component of a CBP complex involved in mediating the transcriptional effects of nuclear hormone receptors. PMID:10866662

Phenobarbital Meets Phosphorylation of Nuclear Receptors

PubMed Central

2017-01-01

Phenobarbital was the first therapeutic drug to be characterized for its induction of hepatic drug metabolism. Essentially at the same time, cytochrome P450, an enzyme that metabolizes drugs, was discovered. After nearly 50 years of investigation, the molecular target of phenobarbital induction has now been delineated to phosphorylation at threonine 38 of the constitutive androstane receptor (NR1I3), a member of the nuclear receptor superfamily. Determining this mechanism has provided us with the molecular basis to understand drug induction of drug metabolism and disposition. Threonine 38 is conserved as a phosphorylation motif in the majority of both mouse and human nuclear receptors, providing us with an opportunity to integrate diverse functions of nuclear receptors. Here, I review the works and accomplishments of my laboratory at the National Institutes of Health National Institute of Environmental Health Sciences and the future research directions of where our study of the constitutive androstane receptor might take us. PMID:28356313

Phenobarbital Meets Phosphorylation of Nuclear Receptors.

PubMed

Negishi, Masahiko

2017-05-01

Phenobarbital was the first therapeutic drug to be characterized for its induction of hepatic drug metabolism. Essentially at the same time, cytochrome P450, an enzyme that metabolizes drugs, was discovered. After nearly 50 years of investigation, the molecular target of phenobarbital induction has now been delineated to phosphorylation at threonine 38 of the constitutive androstane receptor (NR1I3), a member of the nuclear receptor superfamily. Determining this mechanism has provided us with the molecular basis to understand drug induction of drug metabolism and disposition. Threonine 38 is conserved as a phosphorylation motif in the majority of both mouse and human nuclear receptors, providing us with an opportunity to integrate diverse functions of nuclear receptors. Here, I review the works and accomplishments of my laboratory at the National Institutes of Health National Institute of Environmental Health Sciences and the future research directions of where our study of the constitutive androstane receptor might take us. U.S. Government work not protected by U.S. copyright.

Sumoylation of the Basic Helix-Loop-Helix Transcription Factor Sharp-1 Regulates Recruitment of the Histone Methyltransferase G9a and Function in Myogenesis*

PubMed Central

Wang, Yaju; Shankar, Shilpa Rani; Kher, Devaki; Ling, Belinda Mei Tze; Taneja, Reshma

2013-01-01

Sumoylation is an important post-translational modification that alters the activity of many transcription factors. However, the mechanisms that link sumoylation to alterations in chromatin structure, which culminate in tissue specific gene expression, are not fully understood. In this study, we demonstrate that SUMO modification of the transcription factor Sharp-1 is required for its full transcriptional repression activity and function as an inhibitor of skeletal muscle differentiation. Sharp-1 is modified by sumoylation at two conserved lysine residues 240 and 255. Mutation of these SUMO acceptor sites in Sharp-1 does not impact its subcellular localization but attenuates its ability to act as a transcriptional repressor and inhibit myogenic differentiation. Consistently, co-expression of the SUMO protease SENP1 with wild type Sharp-1 abrogates Sharp-1-dependent inhibition of myogenesis. Interestingly, sumoylation acts as a signal for recruitment of the co-repressor G9a. Thus, enrichment of G9a, and histone H3 lysine 9 dimethylation (H3K9me2), a signature of G9a activity, is dramatically reduced at muscle promoters in cells expressing sumoylation-defective Sharp-1. Our findings demonstrate how sumoylation of Sharp-1 exerts an impact on chromatin structure and transcriptional repression of muscle gene expression through recruitment of G9a. PMID:23637228

Nuclear Receptor Coactivator Function in Reproductive Physiology and Behavior

PubMed Central

Molenda, Heather A.; Kilts, Caitlin P.; Allen, Rachel L.; Tetel, Marc J.

2009-01-01

Gonadal steroid hormones act throughout the body to elicit changes in gene expression that result in profound effects on reproductive physiology and behavior. Steroid hormones exert many of these effects by binding to their respective intracellular receptors, which are members of a nuclear receptor superfamily of transcriptional activators. A variety of in vitro studies indicate that nuclear receptor coactivators are required for efficient transcriptional activity of steroid receptors. Many of these coactivators are found in a variety of steroid hormone-responsive reproductive tissues, including the reproductive tract, mammary gland, and brain. While many nuclear receptor coactivators have been investigated in vitro, we are only now beginning to understand their function in reproductive physiology and behavior. In this review, we discuss the general mechanisms of action of nuclear receptor coactivators in steroid-dependent gene transcription. We then review some recent and exciting findings on the function of nuclear receptor coactivators in steroid-dependent brain development and reproductive physiology and behavior. PMID:12855594

Identification of sumoylation activating enzyme 1 inhibitors by structure-based virtual screening.

PubMed

Kumar, Ashutosh; Ito, Akihiro; Hirohama, Mikako; Yoshida, Minoru; Zhang, Kam Y J

2013-04-22

SUMO activating enzyme 1 (SUMO E1) is responsible for the activation of SUMO in the first step of the sumoylation cascade. SUMO E1 is linked to many human diseases including cancer, thus making it a potential therapeutic target. There are few reported SUMO E1 inhibitors including several natural products. To identify small molecule inhibitors of SUMO E1 with better drug-like properties for potential therapeutic studies, we have used structure-based virtual screening to identify hits from the Maybridge small molecule library for biological assay. Our virtual screening protocol involves fast docking of the entire small molecule library with rigid protein and ligands followed by redocking of top hits using a method that incorporates both ligand and protein flexibility. Subsequently, the top-ranking compounds were prioritized using the molecular dynamics simulation-based binding free energy calculation. Out of 24 compounds that were acquired and tested using in vitro sumoylation assay, four of them showed more than 85% inhibition of sumoylation with the most active compound showing an IC50 of 14.4 μM. A similarity search with the most active compound in the ZINC database has identified three more compounds with improved potency. These compounds share a common phenyl urea scaffold and have been confirmed to inhibit SUMO E1 by in vitro SUMO-1 thioester bond formation assay. Our study suggests that these phenyl urea compounds could be used as a starting point for the development of novel therapeutic agents.

PHD Domain-Mediated E3 Ligase Activity Directs Intramolecular Sumoylation of an Adjacent Bromodomain which is Required for Gene Silencing

PubMed Central

Ivanov, Alexey V.; Peng, Hongzhuang; Yurchenko, Vyacheslav; Yap, Kyoko L.; Negorev, Dmitri G.; Schultz, David C.; Psulkowski, Elyse; Fredericks, William J.; White, David E.; Maul, Gerd G.; Sadofsky, Moshe J.; Zhou, Ming-Ming; Rauscher, Frank J.

2015-01-01

SUMMARY Tandem PHD and bromodomains are often found in chromatin-associated proteins and have been shown to cooperate in gene silencing. Each domain can bind specifically modified histones: the mechanisms of cooperation between these domains are unknown. We show that the PHD domain of the KAP1 corepressor functions as an intramolecular E3 ligase for sumoylation of the adjacent bromodomain. The RING finger-like structure of the PHD domain is required for both Ubc9 binding and sumoylation and directs modification to specific lysine residues in the bromodomain. Sumoylation is required for KAP1-mediated gene silencing and functions by directly recruiting the SETDB1 histone methyltransferase and the CHD3/Mi2 component of the NuRD complex via SUMO interacting motifs. Sumoylated KAP1 stimulates the histone methyltransferase activity of SETDB1. These data provide a mechanistic explanation for the cooperation of PHD and bromodomains in gene regulation and describe a new function of the PHD domain as an intramolecular E3 SUMO ligase. PMID:18082607

Studying Nuclear Receptor Complexes in the Cellular Environment.

PubMed

Schaufele, Fred

2016-01-01

The ligand-regulated structure and biochemistry of nuclear receptor complexes are commonly determined by in vitro studies of isolated receptors, cofactors, and their fragments. However, in the living cell, the complexes that form are governed not just by the relative affinities of isolated cofactors for the receptor but also by the cell-specific sequestration or concentration of subsets of competing or cooperating cofactors, receptors, and other effectors into distinct subcellular domains and/or their temporary diversion into other cellular activities. Most methods developed to understand nuclear receptor function in the cellular environment involve the direct tagging of the nuclear receptor or its cofactors with fluorescent proteins (FPs) and the tracking of those FP-tagged factors by fluorescence microscopy. One of those approaches, Förster resonance energy transfer (FRET) microscopy, quantifies the transfer of energy from a higher energy "donor" FP to a lower energy "acceptor" FP attached to a single protein or to interacting proteins. The amount of FRET is influenced by the ligand-induced changes in the proximities and orientations of the FPs within the tagged nuclear receptor complexes, which is an indicator of the structure of the complexes, and by the kinetics of the interaction between FP-tagged factors. Here, we provide a guide for parsing information about the structure and biochemistry of nuclear receptor complexes from FRET measurements in living cells.

Improved Dual-Luciferase Reporter Assays for Nuclear Receptors

PubMed Central

Paguio, Aileen; Stecha, Pete; Wood, Keith V; Fan, Frank

2010-01-01

Nuclear receptors play important roles in many cellular functions through control of gene transcription. It is also a large target class for drug discovery. Luciferase reporter assays are frequently used to study nuclear receptor function because of their wide dynamic range, low endogenous activity, and ease of use. Recent improvements of luciferase genes and vectors have further enhanced their utilities. Here we applied these improvements to two reporter formats for studying nuclear receptors. The first assay contains a Murine Mammary Tumor Virus promoter upstream of a destabilized luciferase. The presence of response elements for nuclear hormone receptor in this promoter allows the studies of endogenous and/or exogenous full length receptors. The second assay contains a ligand binding domain (LBD) of a nuclear receptor fused to the GAL4 DNA binding domain (DBD) on one vector and multiple Gal4 Upstream Activator Sequences (UAS) upstream of luciferase reporter on another vector. We showed that codon optimization of luciferase reporter genes increased expression levels in conjunction with the incorporation of protein destabilizing sequences into luciferase led to a larger assay dynamic range in both formats. The optimum number of UAS to generate the best response was determined. The expression vector for nuclear receptor LBD/GAL4 DBD fusion also constitutively expresses a Renilla luciferase-neoR fusion protein, which provides selection capability (G418 resistance, neoR) as well as an internal control (Renilla luciferase). This dual-luciferase format allowed detecting compound cytotoxicity or off-target change in expression during drug screening, therefore improved data quality. These luciferase reporter assays provided better research and drug discovery tools for studying the functions of full length nuclear receptors and ligand binding domains. PMID:21687560

An emerging link between LIM domain proteins and nuclear receptors.

PubMed

Sala, Stefano; Ampe, Christophe

2018-06-01

Nuclear receptors are ligand-activated transcription factors that partake in several biological processes including development, reproduction and metabolism. Over the last decade, evidence has accumulated that group 2, 3 and 4 LIM domain proteins, primarily known for their roles in actin cytoskeleton organization, also partake in gene transcription regulation. They shuttle between the cytoplasm and the nucleus, amongst other as a consequence of triggering cells with ligands of nuclear receptors. LIM domain proteins act as important coregulators of nuclear receptor-mediated gene transcription, in which they can either function as coactivators or corepressors. In establishing interactions with nuclear receptors, the LIM domains are important, yet pleiotropy of LIM domain proteins and nuclear receptors frequently occurs. LIM domain protein-nuclear receptor complexes function in diverse physiological processes. Their association is, however, often linked to diseases including cancer.

The adenovirus E4-ORF3 protein functions as a SUMO E3 ligase for TIF-1γ sumoylation and poly-SUMO chain elongation.

PubMed

Sohn, Sook-Young; Hearing, Patrick

2016-06-14

The adenovirus (Ad) early region 4 (E4)-ORF3 protein regulates diverse cellular processes to optimize the host environment for the establishment of Ad replication. E4-ORF3 self-assembles into multimers to form a nuclear scaffold in infected cells and creates distinct binding interfaces for different cellular target proteins. Previous studies have shown that the Ad5 E4-ORF3 protein induces sumoylation of multiple cellular proteins and subsequent proteasomal degradation of some of them, but the detailed mechanism of E4-ORF3 function remained unknown. Here, we investigate the role of E4-ORF3 in the sumoylation process by using transcription intermediary factor (TIF)-1γ as a substrate. Remarkably, we discovered that purified E4-ORF3 protein stimulates TIF-1γ sumoylation in vitro, demonstrating that E4-ORF3 acts as a small ubiquitin-like modifier (SUMO) E3 ligase. Furthermore, E4-ORF3 significantly increases poly-SUMO3 chain formation in vitro in the absence of substrate, showing that E4-ORF3 has SUMO E4 elongase activity. An E4-ORF3 mutant, which is defective in protein multimerization, exhibited severely decreased activity, demonstrating that E4-ORF3 self-assembly is required for these activities. Using a SUMO3 mutant, K11R, we found that E4-ORF3 facilitates the initial acceptor SUMO3 conjugation to TIF-1γ as well as poly-SUMO chain elongation. The E4-ORF3 protein displays no SUMO-targeted ubiquitin ligase activity in our assay system. These studies reveal the mechanism by which E4-ORF3 targets specific cellular proteins for sumoylation and proteasomal degradation and provide significant insight into how a small viral protein can play a role as a SUMO E3 ligase and E4-like SUMO elongase to impact a variety of cellular responses.

Identification of Gene Markers for Activation of the Nuclear Receptor Pregnane X Receptor

EPA Science Inventory

Many environmentally-relevant chemicals and drugs activate the nuclear receptor pregnane X receptor (PXR). Activation of PXR in the mouse liver can lead to increases in liver weight in part through increased hepatocyte replication similar to chemicals that activate other nuclear ...

Systematic Localization and Identification of SUMOylation Substrates in Knock-In Mice Expressing Affinity-Tagged SUMO1.

PubMed

Tirard, Marilyn; Brose, Nils

2016-01-01

Protein SUMOylation is a posttranslational protein modification that is emerging as a key regulatory process in neurobiology. To date, however, SUMOylation in vivo has only been studied cursorily. Knock-in mice expressing His6-HA-SUMO1 from the Sumo1 locus allow for the highly specific localization and identification of endogenous SUMO1 substrates under physiological and pathophysiological conditions. By making use of the HA-tag and using wild-type mice for highly stringent negative control samples, SUMO1 targets can be specifically localized in and purified from cultured mouse nerve cells and mouse tissues.

NUREBASE: database of nuclear hormone receptors.

PubMed

Duarte, Jorge; Perrière, Guy; Laudet, Vincent; Robinson-Rechavi, Marc

2002-01-01

Nuclear hormone receptors are an abundant class of ligand activated transcriptional regulators, found in varying numbers in all animals. Based on our experience of managing the official nomenclature of nuclear receptors, we have developed NUREBASE, a database containing protein and DNA sequences, reviewed protein alignments and phylogenies, taxonomy and annotations for all nuclear receptors. The reviewed NUREBASE is completed by NUREBASE_DAILY, automatically updated every 24 h. Both databases are organized under a client/server architecture, with a client written in Java which runs on any platform. This client, named FamFetch, integrates a graphical interface allowing selection of families, and manipulation of phylogenies and alignments. NUREBASE sequence data is also accessible through a World Wide Web server, allowing complex queries. All information on accessing and installing NUREBASE may be found at http://www.ens-lyon.fr/LBMC/laudet/nurebase.html.

REV-ERB and ROR nuclear receptors as drug targets

PubMed Central

Kojetin, Douglas J.; Burris, Thomas P.

2016-01-01

The nuclear receptors REV-ERB (consisting of REV-ERBα and REV-ERBβ) and retinoic acid receptor-related orphan receptors (RORs; consisting of RORα, RORβ and RORγ) are involved in many physiological processes, including regulation of metabolism, development and immunity as well as the circadian rhythm. The recent characterization of endogenous ligands for these former orphan nuclear receptors has stimulated the development of synthetic ligands and opened up the possibility of targeting these receptors to treat several diseases, including diabetes, atherosclerosis, autoimmunity and cancer. This Review focuses on the latest developments in ROR and REV-ERB pharmacology indicating that these nuclear receptors are druggable targets and that ligands targeting these receptors may be useful in the treatment of several disorders. PMID:24577401

Nuclear Receptor Regulation of Aquaglyceroporins in Metabolic Organs.

PubMed

Tardelli, Matteo; Claudel, Thierry; Bruschi, Francesca Virginia; Trauner, Michael

2018-06-15

Nuclear receptors, such as the farnesoid X receptor (FXR) and the peroxisome proliferator-activated receptors gamma and alpha (PPAR-γ, -α), are major metabolic regulators in adipose tissue and the liver, where they govern lipid, glucose, and bile acid homeostasis, as well as inflammatory cascades. Glycerol and free fatty acids are the end products of lipid droplet catabolism driven by PPARs. Aquaporins (AQPs), a family of 13 small transmembrane proteins, facilitate the shuttling of water, urea, and/or glycerol. The peculiar role of AQPs in glycerol transport makes them pivotal targets in lipid metabolism, especially considering their tissue-specific regulation by the nuclear receptors PPARγ and PPARα. Here, we review the role of nuclear receptors in the regulation of glycerol shuttling in liver and adipose tissue through the function and expression of AQPs.

Hes-1 SUMOylation by protein inhibitor of activated STAT1 enhances the suppressing effect of Hes-1 on GADD45α expression to increase cell survival

PubMed Central

2014-01-01

Background Hairy and Enhancer of split 1 (Hes-1) is a transcriptional repressor that plays an important role in neuronal differentiation and development, but post-translational modifications of Hes-1 are much less known. In the present study, we aimed to investigate whether Hes-1 could be SUMO-modified and identify the candidate SUMO acceptors on Hes-1. We also wished to examine the role of the SUMO E3 ligase protein inhibitor of activated STAT1 (PIAS1) in SUMOylation of Hes-1 and the molecular mechanism of Hes-1 SUMOylation. Further, we aimed to identify the molecular target of Hes-1 and examine how Hes-1 SUMOylation affects its molecular target to affect cell survival. Results In this study, by using HEK293T cells, we have found that Hes-1 could be SUMO-modified and Hes-1 SUMOylation was greatly enhanced by the SUMO E3 ligase PIAS1 at Lys8, Lys27 and Lys39. Furthermore, Hes-1 SUMOylation stabilized the Hes-1 protein and increased the transcriptional suppressing activity of Hes-1 on growth arrest and DNA damage-inducible protein alpha (GADD45α) expression. Overexpression of GADD45α increased, whereas knockdown of GADD45αα expression decreased cell apoptosis. In addition, H2O2 treatment increased the association between PIAS1 and Hes-1 and enhanced the SUMOylation of Hes-1 for endogenous protection. Overexpression of Hes-1 decreased H2O2-induced cell death, but this effect was blocked by transfection of the Hes-1 triple sumo-mutant (Hes-1 3KR). Overexpression of PIAS1 further facilitated the anti-apoptotic effect of Hes-1. Moreover, Hes-1 SUMOylation was independent of Hes-1 phosphorylation and vice versa. Conclusions The present results revealed, for the first time, that Hes-1 could be SUMO-modified by PIAS1 and GADD45α is a novel target of Hes-1. Further, Hes-1 SUMOylation mediates cell survival through enhanced suppression of GADD45α expression. These results revealed a novel role of Hes-1 in addition to its involvement in Notch signaling. They also

The story so far: post-translational regulation of peroxisome proliferator-activated receptors by ubiquitination and SUMOylation

PubMed Central

Wadosky, Kristine M.

2012-01-01

Many studies have implicated the peroxisome proliferator-activated receptor (PPAR) family of nuclear receptor transcription factors in regulating cardiac substrate metabolism and ATP generation. Recently, evidence from a variety of cell culture and organ systems has implicated ubiquitin and small ubiquitin-like modifier (SUMO) conjugation as post-translational modifications that regulate the activity of PPAR transcription factors and their coreceptors/coactivators. Here we introduce the ubiquitin and SUMO conjugation systems and extensively review how they have been shown to regulate all three PPAR isoforms (PPARα, PPARβ/δ, and PPARγ) in addition to the retinoid X receptor and PPARγ coactivator-1α subunits of the larger PPAR transcription factor complex. We then present how the specific ubiquitin (E3) ligases have been implicated and review emerging evidence that post-translational modifications of PPARs with ubiquitin and/or SUMO may play a role in cardiac disease. Because PPAR activity is perturbed in a variety of forms of heart disease and specific proteins regulate this process (E3 ligases), this may be a fruitful area of investigation with respect to finding new therapeutic targets. PMID:22037188

Atypical nuclear localization of VIP receptors in glioma cell lines and patients

DOE Office of Scientific and Technical Information (OSTI.GOV)

Barbarin, Alice; Séité, Paule; Godet, Julie

Highlights: • The VIP receptor VPAC1 contains a putative NLS signal. • VPAC1 is predominantly nuclear in GBM cell lines but not VPAC2. • Non-nuclear VPAC1/2 protein expression is correlated with glioma grade. • Nuclear VPAC1 is observed in 50% of stage IV glioma (GBM). - Abstract: An increasing number of G protein-coupled receptors, like receptors for vasoactive intestinal peptide (VIP), are found in cell nucleus. As VIP receptors are involved in the regulation of glioma cell proliferation and migration, we investigated the expression and the nuclear localization of the VIP receptors VPAC1 and VPAC2 in this cancer. First, bymore » applying Western blot and immunofluorescence detection in three human glioblastoma (GBM) cell lines, we observed a strong nuclear staining for the VPAC1 receptor and a weak nuclear VPAC2 receptor staining. Second, immunohistochemical staining of VPAC1 and VPAC2 on tissue microarrays (TMA) showed that the two receptors were expressed in normal brain and glioma tissues. Expression in the non-nuclear compartment of the two receptors significantly increased with the grade of the tumors. Analysis of nuclear staining revealed a significant increase of VPAC1 staining with glioma grade, with up to 50% of GBM displaying strong VPAC1 nuclear staining, whereas nuclear VPAC2 staining remained marginal. The increase in VPAC receptor expression with glioma grades and the enhanced nuclear localization of the VPAC1 receptors in GBM might be of importance for glioma progression.« less

Steroid receptor coupling becomes nuclear.

PubMed

Galigniana, Mario D

2012-06-22

In this issue of Chemistry & Biology, Grossman et al. report a study on aldosterone-dependent nuclear translocation of the mineralocorticoid receptor (MR). They analyze the dependency of MR retrotransport, DNA-binding, and transcriptional activity on Hsp90 and demonstrate that MR dimerization is a nuclear event. Copyright © 2012 Elsevier Ltd. All rights reserved.

Targeting nuclear receptors for the treatment of fatty liver disease.

PubMed

Tanaka, Naoki; Aoyama, Toshifumi; Kimura, Shioko; Gonzalez, Frank J

2017-11-01

Ligand-activated nuclear receptors, including peroxisome proliferator-activated receptor alpha (PPARα), pregnane X receptor, and constitutive androstane receptor, were first identified as key regulators of the responses against chemical toxicants. However, numerous studies using mouse disease models and human samples have revealed critical roles for these receptors and others, such as PPARβ/δ, PPARγ, farnesoid X receptor (FXR), and liver X receptor (LXR), in maintaining nutrient/energy homeostasis in part through modulation of the gut-liver-adipose axis. Recently, disorders associated with disrupted nutrient/energy homeostasis, e.g., obesity, metabolic syndrome, and non-alcoholic fatty liver disease (NAFLD), are increasing worldwide. Notably, in NAFLD, a progressive subtype exists, designated as non-alcoholic steatohepatitis (NASH) that is characterized by typical histological features resembling alcoholic steatohepatitis (ASH), and NASH/ASH are recognized as major causes of hepatitis virus-unrelated liver cirrhosis and hepatocellular carcinoma. Since hepatic steatosis is basically caused by an imbalance between fat/energy influx and utilization, abnormal signaling of these nuclear receptors contribute to the pathogenesis of fatty liver disease. Standard therapeutic interventions have not been fully established for fatty liver disease, but some new agents that activate or inhibit nuclear receptor signaling have shown promise as possible therapeutic targets. In this review, we summarize recent findings on the roles of nuclear receptors in fatty liver disease and discuss future perspectives to develop promising pharmacological strategies targeting nuclear receptors for NAFLD/NASH. Copyright © 2017 Elsevier Inc. All rights reserved.

HTLV-2B Tax oncoprotein is modified by ubiquitination and sumoylation and displays intracellular localization similar to its homologue HTLV-1 Tax

DOE Office of Scientific and Technical Information (OSTI.GOV)

Turci, Marco; Lodewick, Julie; Righi, Paola

HTLV-1 is more pathogenic than HTLV-2B. The difference is generally attributed to the properties of their individual transactivating Tax proteins. By using internal Flag-6His tagged Tax-1 and Tax-2B, which display transcriptional activities comparable to the untagged proteins and can be recognized by a single anti-Flag antibody, we demonstrate that Tax-2B is modified by ubiquitination and sumoylation. In addition, Tax2B is distributed in punctuate nuclear structures that include the RelA subunit of NF-{kappa}B, as has been previously demonstrated for Tax-1.

SUMOylation Regulates the Homologous to E6-AP Carboxyl Terminus (HECT) Ubiquitin Ligase Rsp5p*

PubMed Central

Novoselova, Tatiana Vladislavovna; Rose, Ruth-Sarah; Marks, Helen Margaret; Sullivan, James Andrew

2013-01-01

The post-translational modifiers ubiquitin and small ubiquitin-related modifier (SUMO) regulate numerous critical signaling pathways and are key to controlling the cellular fate of proteins in eukaryotes. The attachment of ubiquitin and SUMO involves distinct, but related, machinery. However, it is now apparent that many substrates can be modified by both ubiquitin and SUMO and that some regulatory interaction takes place between the respective attachment machinery. Here, we demonstrate that the Saccharomyces cerevisiae ubiquitin ligase Rsp5p, a member of the highly conserved Nedd4 family of ubiquitin ligases, is SUMOylated in vivo. We further show that Rsp5p SUMOylation is mediated by the SUMO ligases Siz1p and Siz2p, members of the conserved family of PIAS SUMO ligases that are, in turn, substrates for Rsp5p-mediated ubiquitylation. Our experiments show that SUMOylated Rsp5p has reduced ubiquitin ligase activity, and similarly, ubiquitylated Siz1p demonstrates reduced SUMO ligase activity leading to respective changes in both ubiquitin-mediated sorting of the manganese transporter Smf1p and polySUMO chain formation. This reciprocal regulation of these highly conserved ligases represents an exciting and previously unidentified system of cross talk between the ubiquitin and SUMO systems. PMID:23443663

A nuclear-receptor-dependent phosphatidylcholine pathway with antidiabetic effects

USDA-ARS?s Scientific Manuscript database

Nuclear hormone receptors regulate diverse metabolic pathways and the orphan nuclear receptor LRH-1 (also known as NR5A2) regulates bile acid biosynthesis. Structural studies have identified phospholipids as potential LRH-1 ligands, but their functional relevance is unclear. Here we show that an unu...

SUMO-Specific Cysteine Protease 1 Promotes Epithelial Mesenchymal Transition of Prostate Cancer Cells via Regulating SMAD4 deSUMOylation.

PubMed

Zhang, Xiaoyan; Wang, Hao; Wang, Hua; Xiao, Fengjun; Seth, Prem; Xu, Weidong; Jia, Qinghua; Wu, Chutse; Yang, Yuefeng; Wang, Lisheng

2017-04-12

In advanced prostate cancer, small ubiquitin-like modifier (SUMO)-specific cysteine protease 1 (SENP1) is up-regulated. However, the role of SENP1 in regulating deSUMOylation of TGF-β/SMADs signaling is unknown. In this study, we developed a lentiviral vector, PLKO.1-shSENP1, to silence SENP1 in prostate cancer cells with high metastatic characteristics (PC3M). Likewise, we also created an adenovirus vector, Ad5/F11p-SENP1 to over-express SENP1 in prostate cancer cells with low metastatic potential (LNCaP). We showed that silencing of SENP1 promoted cellular apoptosis, and inhibited proliferation and migration of PC3M cells. Moreover, SENP1 silencing increased the SMAD4 expression at protein level, up-regulated E-cadherin and down-regulated Vimentin expression, indicating the inhibition of epithelial mesenchymal transition (EMT). Furthermore, SMAD4 interference abolished SENP1-mediated up-regulation of E-cadherin, suggesting that SENP1 regulated E-cadherin expression via SMAD4. SENP1 over-expression in LNCaP cells reduced SMAD4 protein, and promoted EMT via decreasing E-cadherin and increasing Vimentin. Moreover, down-regulation of SMAD4 and E-cadherin were blocked, after transfection with two SUMOylation sites mutated SMAD4, suggesting that SENP1 might reduce SMAD4 levels to regulate E-cadherin expression via deSUMOylation of SMAD4. In conclusion, SENP1 deSUMOylated SMAD4 to promote EMT via up-regulating E-cadherin in prostate cancer cells. Therefore, SENP1 is a potential target for treatment of advanced prostate cancer.

SUMO-Specific Cysteine Protease 1 Promotes Epithelial Mesenchymal Transition of Prostate Cancer Cells via Regulating SMAD4 deSUMOylation

PubMed Central

Zhang, Xiaoyan; Wang, Hao; Wang, Hua; Xiao, Fengjun; Seth, Prem; Xu, Weidong; Jia, Qinghua; Wu, Chutse; Yang, Yuefeng; Wang, Lisheng

2017-01-01

In advanced prostate cancer, small ubiquitin-like modifier (SUMO)-specific cysteine protease 1 (SENP1) is up-regulated. However, the role of SENP1 in regulating deSUMOylation of TGF-β/SMADs signaling is unknown. In this study, we developed a lentiviral vector, PLKO.1-shSENP1, to silence SENP1 in prostate cancer cells with high metastatic characteristics (PC3M). Likewise, we also created an adenovirus vector, Ad5/F11p-SENP1 to over-express SENP1 in prostate cancer cells with low metastatic potential (LNCaP). We showed that silencing of SENP1 promoted cellular apoptosis, and inhibited proliferation and migration of PC3M cells. Moreover, SENP1 silencing increased the SMAD4 expression at protein level, up-regulated E-cadherin and down-regulated Vimentin expression, indicating the inhibition of epithelial mesenchymal transition (EMT). Furthermore, SMAD4 interference abolished SENP1-mediated up-regulation of E-cadherin, suggesting that SENP1 regulated E-cadherin expression via SMAD4. SENP1 over-expression in LNCaP cells reduced SMAD4 protein, and promoted EMT via decreasing E-cadherin and increasing Vimentin. Moreover, down-regulation of SMAD4 and E-cadherin were blocked, after transfection with two SUMOylation sites mutated SMAD4, suggesting that SENP1 might reduce SMAD4 levels to regulate E-cadherin expression via deSUMOylation of SMAD4. In conclusion, SENP1 deSUMOylated SMAD4 to promote EMT via up-regulating E-cadherin in prostate cancer cells. Therefore, SENP1 is a potential target for treatment of advanced prostate cancer. PMID:28417919

Identification of a molecular recognition feature in the E1A oncoprotein that binds the SUMO conjugase UBC9 and likely interferes with polySUMOylation.

PubMed

Yousef, A F; Fonseca, G J; Pelka, P; Ablack, J N G; Walsh, C; Dick, F A; Bazett-Jones, D P; Shaw, G S; Mymryk, J S

2010-08-19

Hub proteins have central roles in regulating cellular processes. By targeting a single cellular hub, a viral oncogene may gain control over an entire module in the cellular interaction network that is potentially comprised of hundreds of proteins. The adenovirus E1A oncoprotein is a viral hub that interacts with many cellular hub proteins by short linear motifs/molecular recognition features (MoRFs). These interactions transform the architecture of the cellular protein interaction network and virtually reprogram the cell. To identify additional MoRFs within E1A, we screened portions of E1A for their ability to activate yeast pseudohyphal growth or differentiation. This identified a novel functional region within E1A conserved region 2 comprised of the sequence EVIDLT. This MoRF is necessary and sufficient to bind the N-terminal region of the SUMO conjugase UBC9, which also interacts with SUMO noncovalently and is involved in polySUMOylation. Our results suggest that E1A interferes with polySUMOylation, but not with monoSUMOylation. These data provide the first insight into the consequences of the interaction of E1A with UBC9, which was initially described in 1996. We further demonstrate that polySUMOylation regulates pseudohyphal growth and promyelocytic leukemia body reorganization by E1A. In conclusion, the interaction of the E1A oncogene with UBC9 mimics the normal binding between SUMO and UBC9 and represents a novel mechanism to modulate polySUMOylation.

Nuclear Receptor Signaling Atlas: Opening Access to the Biology of Nuclear Receptor Signaling Pathways.

PubMed

Becnel, Lauren B; Darlington, Yolanda F; Ochsner, Scott A; Easton-Marks, Jeremy R; Watkins, Christopher M; McOwiti, Apollo; Kankanamge, Wasula H; Wise, Michael W; DeHart, Michael; Margolis, Ronald N; McKenna, Neil J

2015-01-01

Signaling pathways involving nuclear receptors (NRs), their ligands and coregulators, regulate tissue-specific transcriptomes in diverse processes, including development, metabolism, reproduction, the immune response and neuronal function, as well as in their associated pathologies. The Nuclear Receptor Signaling Atlas (NURSA) is a Consortium focused around a Hub website (www.nursa.org) that annotates and integrates diverse 'omics datasets originating from the published literature and NURSA-funded Data Source Projects (NDSPs). These datasets are then exposed to the scientific community on an Open Access basis through user-friendly data browsing and search interfaces. Here, we describe the redesign of the Hub, version 3.0, to deploy "Web 2.0" technologies and add richer, more diverse content. The Molecule Pages, which aggregate information relevant to NR signaling pathways from myriad external databases, have been enhanced to include resources for basic scientists, such as post-translational modification sites and targeting miRNAs, and for clinicians, such as clinical trials. A portal to NURSA's Open Access, PubMed-indexed journal Nuclear Receptor Signaling has been added to facilitate manuscript submissions. Datasets and information on reagents generated by NDSPs are available, as is information concerning periodic new NDSP funding solicitations. Finally, the new website integrates the Transcriptomine analysis tool, which allows for mining of millions of richly annotated public transcriptomic data points in the field, providing an environment for dataset re-use and citation, bench data validation and hypothesis generation. We anticipate that this new release of the NURSA database will have tangible, long term benefits for both basic and clinical research in this field.

Nuclear receptors in pancreatic tumor cells.

PubMed

Damaskos, Christos; Garmpis, Nikolaos; Karatzas, Theodore; Kostakis, Ioannis D; Nikolidakis, Lampros; Kostakis, Alkiviadis; Kouraklis, Gregory

2014-12-01

This review focuses on nuclear receptors expressed in pancreatic cancer. An extensive search of articles published up to March 2013 was conducted using the MEDLINE database. The key words used were "pancreatic cancer", "molecular receptors" and "growth factors". A total of 112 articles referred to pancreatic cancer, molecular receptors and/or growth factors were included. Receptors of growth factors, such as the epithelial growth factor receptor, insulin-like growth factor-1 receptor, vascular endothelial growth factor receptor and others, such as integrin α5β1, somatostatin receptors, the death receptor 5, claudin, notch receptors, mesothelin receptors, follicle-stimulating hormone receptors, the MUC1 receptor, the adrenomedullin receptor, the farnesoid X receptor, the transferrin receptor, sigma-2 receptors, the chemokine receptor CXCR4, the urokinase plasminogen activator receptor, the ephrine A2 receptor, the GRIA3 receptor, the RON receptor and the angiotensin II receptor AT-1 are expressed in pancreatic tumor cells. These molecules are implicated in tumor growth, apoptosis, angiogenesis, metastasis etc. After identifying the molecular receptors associated with the pancreatic cancer, many more target molecules playing important roles in tumor pathophysiology and senescence-associated signal transduction in cancer cells will be identified. This may have a significant influence on diagnosis, therapy and prognosis of pancreatic cancer. Copyright© 2014 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.

Rethinking Nuclear Receptors as Potential Therapeutic Targets for Retinal Diseases.

PubMed

Choudhary, Mayur; Malek, Goldis

2016-12-01

Collectively, retinal diseases, including age-related macular degeneration, retinitis pigmentosa, and diabetic retinopathy, result in severe vision impairment worldwide. The absence and/or limited availability of successful drug therapies for these blinding disorders necessitates further understanding their pathobiology and identifying new targetable signaling pathways. Nuclear receptors are transcription regulators of many key aspects of human physiology, as well as pathophysiology, with reported roles in development, aging, and disease. Some of the pathways regulated by nuclear receptors include, but are not limited to, angiogenesis, inflammation, and lipid metabolic dysregulation, mechanisms also important in the initiation and development of several retinal diseases. Herein, we present an overview of the biology of three diseases affecting the posterior eye, summarize a growing body of evidence that suggests direct or indirect involvement of nuclear receptors in disease progression, and discuss the therapeutic potential of targeting nuclear receptors for treatment.

Rethinking Nuclear Receptors as Potential Therapeutic Targets for Retinal Diseases

PubMed Central

Choudhary, Mayur; Malek, Goldis

2017-01-01

Collectively, retinal diseases, including age-related macular degeneration, retinitis pigmentosa, and diabetic retinopathy, result in severe vision impairment worldwide. The absence and/or limited availability of successful drug therapies for these blinding disorders necessitates further understanding their pathobiology and identifying new targetable signaling pathways. Nuclear receptors are transcription regulators of many key aspects of human physiology, as well as pathophysiology, with reported roles in development, aging, and disease. Some of the pathways regulated by nuclear receptors include, but are not limited to, angiogenesis, inflammation, and lipid metabolic dysregulation, mechanisms also important in the initiation and development of several retinal diseases. Herein, we present an overview of the biology of three diseases affecting the posterior eye, summarize a growing body of evidence that suggests direct or indirect involvement of nuclear receptors in disease progression, and discuss the therapeutic potential of targeting nuclear receptors for treatment. PMID:27455994

Computational identification of post-translational modification-based nuclear import regulations by characterizing nuclear localization signal-import receptor interaction.

PubMed

Lin, Jhih-Rong; Liu, Zhonghao; Hu, Jianjun

2014-10-01

The binding affinity between a nuclear localization signal (NLS) and its import receptor is closely related to corresponding nuclear import activity. PTM-based modulation of the NLS binding affinity to the import receptor is one of the most understood mechanisms to regulate nuclear import of proteins. However, identification of such regulation mechanisms is challenging due to the difficulty of assessing the impact of PTM on corresponding nuclear import activities. In this study we proposed NIpredict, an effective algorithm to predict nuclear import activity given its NLS, in which molecular interaction energy components (MIECs) were used to characterize the NLS-import receptor interaction, and the support vector regression machine (SVR) was used to learn the relationship between the characterized NLS-import receptor interaction and the corresponding nuclear import activity. Our experiments showed that nuclear import activity change due to NLS change could be accurately predicted by the NIpredict algorithm. Based on NIpredict, we developed a systematic framework to identify potential PTM-based nuclear import regulations for human and yeast nuclear proteins. Application of this approach has identified the potential nuclear import regulation mechanisms by phosphorylation of two nuclear proteins including SF1 and ORC6. © 2014 Wiley Periodicals, Inc.

Identification of COUP-TFII Orphan Nuclear Receptor as a Retinoic Acid-Activated Receptor

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kruse, Schoen W; Suino-Powell, Kelly; Zhou, X Edward

2010-01-12

The chicken ovalbumin upstream promoter-transcription factors (COUP-TFI and II) make up the most conserved subfamily of nuclear receptors that play key roles in angiogenesis, neuronal development, organogenesis, cell fate determination, and metabolic homeostasis. Although the biological functions of COUP-TFs have been studied extensively, little is known of their structural features or aspects of ligand regulation. Here we report the ligand-free 1.48 {angstrom} crystal structure of the human COUP-TFII ligand-binding domain. The structure reveals an autorepressed conformation of the receptor, where helix {alpha}10 is bent into the ligand-binding pocket and the activation function-2 helix is folded into the cofactor binding site,more » thus preventing the recruitment of coactivators. In contrast, in multiple cell lines, COUP-TFII exhibits constitutive transcriptional activity, which can be further potentiated by nuclear receptor coactivators. Mutations designed to disrupt cofactor binding, dimerization, and ligand binding, substantially reduce the COUP-TFII transcriptional activity. Importantly, retinoid acids are able to promote COUP-TFII to recruit coactivators and activate a COUP-TF reporter construct. Although the concentration needed is higher than the physiological levels of retinoic acids, these findings demonstrate that COUP-TFII is a ligand-regulated nuclear receptor, in which ligands activate the receptor by releasing it from the autorepressed conformation.« less

Nuclear Receptor Signaling Atlas: Opening Access to the Biology of Nuclear Receptor Signaling Pathways

PubMed Central

Becnel, Lauren B.; Darlington, Yolanda F.; Ochsner, Scott A.; Easton-Marks, Jeremy R.; Watkins, Christopher M.; McOwiti, Apollo; Kankanamge, Wasula H.; Wise, Michael W.; DeHart, Michael; Margolis, Ronald N.; McKenna, Neil J.

2015-01-01

Signaling pathways involving nuclear receptors (NRs), their ligands and coregulators, regulate tissue-specific transcriptomes in diverse processes, including development, metabolism, reproduction, the immune response and neuronal function, as well as in their associated pathologies. The Nuclear Receptor Signaling Atlas (NURSA) is a Consortium focused around a Hub website (www.nursa.org) that annotates and integrates diverse ‘omics datasets originating from the published literature and NURSA-funded Data Source Projects (NDSPs). These datasets are then exposed to the scientific community on an Open Access basis through user-friendly data browsing and search interfaces. Here, we describe the redesign of the Hub, version 3.0, to deploy “Web 2.0” technologies and add richer, more diverse content. The Molecule Pages, which aggregate information relevant to NR signaling pathways from myriad external databases, have been enhanced to include resources for basic scientists, such as post-translational modification sites and targeting miRNAs, and for clinicians, such as clinical trials. A portal to NURSA’s Open Access, PubMed-indexed journal Nuclear Receptor Signaling has been added to facilitate manuscript submissions. Datasets and information on reagents generated by NDSPs are available, as is information concerning periodic new NDSP funding solicitations. Finally, the new website integrates the Transcriptomine analysis tool, which allows for mining of millions of richly annotated public transcriptomic data points in the field, providing an environment for dataset re-use and citation, bench data validation and hypothesis generation. We anticipate that this new release of the NURSA database will have tangible, long term benefits for both basic and clinical research in this field. PMID:26325041

SUMOylation of the mitochondrial fission protein Drp1 occurs at multiple nonconsensus sites within the B domain and is linked to its activity cycle

PubMed Central

Figueroa-Romero, Claudia; Iñiguez-Lluhí, Jorge A.; Stadler, Julia; Chang, Chuang-Rung; Arnoult, Damien; Keller, Peter J.; Hong, Yu; Blackstone, Craig; Feldman, Eva L.

2009-01-01

Dynamin-related protein (Drp) 1 is a key regulator of mitochondrial fission and is composed of GTP-binding, Middle, insert B, and C-terminal GTPase effector (GED) domains. Drp1 associates with mitochondrial fission sites and promotes membrane constriction through its intrinsic GTPase activity. The mechanisms that regulate Drp1 activity remain poorly understood but are likely to involve reversible post-translational modifications, such as conjugation of small ubiquitin-like modifier (SUMO) proteins. Through a detailed analysis, we find that Drp1 interacts with the SUMO-conjugating enzyme Ubc9 via multiple regions and demonstrate that Drp1 is a direct target of SUMO modification by all three SUMO isoforms. While Drp1 does not harbor consensus SUMOylation sequences, our analysis identified2 clusters of lysine residues within the B domain that serve as noncanonical conjugation sites. Although initial analysis indicates that mitochondrial recruitment of ectopically expressed Drp1 in response to staurosporine is unaffected by loss of SUMOylation, we find that Drp1 SUMOylation is enhanced in the context of the K38A mutation. This dominant-negative mutant, which is deficient in GTP binding and hydrolysis, does not associate with mitochondria and prevents normal mitochondrial fission. This finding suggests that SUMOylation of Drp1 is linked to its activity cycle and is influenced by Drp1 localization.—Figueroa-Romero, C., Iñiguez-Lluhí, J. A., Stadler, J., Chang, C.-R., Arnoult, D., Keller, P. J., Hong, Y., Blackstone, C., Feldman, E. L. SUMOylation of the mitochondrial fission protein Drp1 occurs at multiple nonconsensus sites within the B domain and is linked to its activity cycle. PMID:19638400

NR4A nuclear receptors support memory enhancement by histone deacetylase inhibitors

PubMed Central

Hawk, Joshua D.; Bookout, Angie L.; Poplawski, Shane G.; Bridi, Morgan; Rao, Allison J.; Sulewski, Michael E.; Kroener, Brian T.; Manglesdorf, David J.; Abel, Ted

2012-01-01

The formation of a long-lasting memory requires a transcription-dependent consolidation period that converts a short-term memory into a long-term memory. Nuclear receptors compose a class of transcription factors that regulate diverse biological processes, and several nuclear receptors have been implicated in memory formation. Here, we examined the potential contribution of nuclear receptors to memory consolidation by measuring the expression of all 49 murine nuclear receptors after learning. We identified 13 nuclear receptors with increased expression after learning, including all 3 members of the Nr4a subfamily. These CREB-regulated Nr4a genes encode ligand-independent “orphan” nuclear receptors. We found that blocking NR4A activity in memory-supporting brain regions impaired long-term memory but did not impact short-term memory in mice. Further, expression of Nr4a genes increased following the memory-enhancing effects of histone deacetylase (HDAC) inhibitors. Blocking NR4A signaling interfered with the ability of HDAC inhibitors to enhance memory. These results demonstrate that the Nr4a gene family contributes to memory formation and is a promising target for improving cognitive function. PMID:22996661

The peroxisome proliferator-activated receptor: A family of nuclear receptors role in various diseases

PubMed Central

Tyagi, Sandeep; Gupta, Paras; Saini, Arminder Singh; Kaushal, Chaitnya; Sharma, Saurabh

2011-01-01

Peroxisome proliferator-activated receptors (PPARs) are ligand-activated transcription factors of nuclear hormone receptor superfamily comprising of the following three subtypes: PPARα, PPARγ, and PPARβ/δ. Activation of PPAR-α reduces triglyceride level and is involved in regulation of energy homeostasis. Activation of PPAR-γ causes insulin sensitization and enhances glucose metabolism, whereas activation of PPAR-β/δ enhances fatty acids metabolism. Thus, PPAR family of nuclear receptors plays a major regulatory role in energy homeostasis and metabolic function. The present review critically analyzes the protective and detrimental effect of PPAR agonists in dyslipidemia, diabetes, adipocyte differentiation, inflammation, cancer, lung diseases, neurodegenerative disorders, fertility or reproduction, pain, and obesity. PMID:22247890

Luteolin Modulates SERCA2a Leading to Attenuation of Myocardial Ischemia/ Reperfusion Injury via Sumoylation at Lysine 585 in Mice.

PubMed

Du, Yinping; Liu, Ping; Xu, Tongda; Pan, Defeng; Zhu, Hong; Zhai, Nana; Zhang, Yanbin; Li, Dongye

2018-01-01

The myocardial sarcoplasmic reticulum calcium ATPase (SERCA2a) is a pivotal pump responsible for calcium cycling in cardiomyocytes. The present study investigated the effect of luteolin (Lut) on restoring SERCA2a protein level and stability reduced by myocardial ischemia/reperfusion (I/R) injury. We verified a hypothesis that Lut protected against myocardial I/R injury by regulating SERCA2a SUMOylation. The hemodynamic data, myocardial infarct size of intact hearts, apoptotic analysis, mitochondrial membrane potential (ΔΨm), the level of SERCA2a SUMOylation, and the activity and expression of SERCA2a were examined in vivo and in vitro to clarify the cardioprotective effects of Lut after SUMO1 was knocked down or over-expressed. The putative SUMO conjugation sites in mouse SERCA2a were investigated as the possible regulatory mechanism of Lut. Initially, we found that Lut reversed the SUMOylation and stability of SERCA2a as well as the expression of SUMO1, which were reduced by I/R injury in vitro. Furthermore, Lut increased the expression and activity of SERCA2a partly through SUMO1, thus improving ΔΨm and reducing apoptotic cells in vitro and promoting the recovery of heart function and reducing infarct size in vivo. We also demonstrated that SUMO acceptor sites in mouse SERCA2a involving lysine 585, 480 and 571. Among the three acceptor sites, Lut enhanced SERCA2a stability via lysine 585. Our results suggest that Lut regulates SERCA2a through SUMOylation at lysine 585 to attenuate myocardial I/R injury. © 2018 The Author(s). Published by S. Karger AG, Basel.

The Therapeutic Role of Xenobiotic Nuclear Receptors against Metabolic Syndrome.

PubMed

Pu, Shuqi; Wu, Xiaojie; Yang, Xiaoying; Zhang, Yunzhan; Dai, Yunkai; Zhang, Yueling; Wu, Xiaoting; Liu, Yan; Cui, Xiaona; Jin, Haiyong; Cao, Jianhong; Li, Ruliu; Cai, Jiazhong; Cao, Qizhi; Hu, Ling; Gao, Yong

2018-06-10

Xenobiotic nuclear receptors (XNRs) are nuclear receptors that characterized by coordinately regulating the expression of genes encoding drug-metabolizing enzymes and transporters to essentially eliminate and detoxify xenobiotics and endobiotics from the body, including the peroxisome proliferator-activated receptor (PPAR), the farnesoid X receptor (FXR), the liver X receptor (LXR), the pregnane X receptor (PXR) and the constitutive androstane receptor (CAR). Heretofore, increasing evidences have suggested that these five XNRs are not only involved in the regulation of xeno-/endo-biotics detoxication but also the development of human diseases, such as cancer, obesity and diabetes. PPAR, FXR, LXR, PXR and CAR, as the receptors for numerous natural or synthetic compounds may be the most effective therapeutic targets in the treatment of metabolic diseases. In this review, we will focus on these five XNRs and their recently discovered functions in diabetes and its complications. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

Minireview: The Role of Nuclear Receptors in Photoreceptor Differentiation and Disease

PubMed Central

Swaroop, Anand

2012-01-01

Rod and cone photoreceptors are specialized sensory cells that mediate vision. Transcriptional controls are critical for the development and long-term survival of photoreceptors; when these controls become ineffective, retinal dysfunction or degenerative disease may result. This review discusses the role of nuclear receptors, a class of ligand-regulated transcription factors, at key stages of photoreceptor life in the mammalian retina. Nuclear receptors with known ligands, such as retinoids or thyroid hormone, together with several orphan receptors without identified physiological ligands, complement other classes of transcription factors in directing the differentiation and functional maintenance of photoreceptors. The potential of nuclear receptors to respond to ligands introduces versatility into the control of photoreceptor development and function and may suggest new opportunities for treatments of photoreceptor disease. PMID:22556342

NR4A nuclear receptors are orphans but not lonesome.

PubMed

Kurakula, Kondababu; Koenis, Duco S; van Tiel, Claudia M; de Vries, Carlie J M

2014-11-01

The NR4A subfamily of nuclear receptors consists of three mammalian members: Nur77, Nurr1, and NOR-1. The NR4A receptors are involved in essential physiological processes such as adaptive and innate immune cell differentiation, metabolism and brain function. They act as transcription factors that directly modulate gene expression, but can also form trans-repressive complexes with other transcription factors. In contrast to steroid hormone nuclear receptors such as the estrogen receptor or the glucocorticoid receptor, no ligands have been described for the NR4A receptors. This lack of known ligands might be explained by the structure of the ligand-binding domain of NR4A receptors, which shows an active conformation and a ligand-binding pocket that is filled with bulky amino acid side-chains. Other mechanisms, such as transcriptional control, post-translational modifications and protein-protein interactions therefore seem to be more important in regulating the activity of the NR4A receptors. For Nur77, over 80 interacting proteins (the interactome) have been identified so far, and roughly half of these interactions has been studied in more detail. Although the NR4As show some overlap in interacting proteins, less information is available on the interactome of Nurr1 and NOR-1. Therefore, the present review will describe the current knowledge on the interactomes of all three NR4A nuclear receptors with emphasis on Nur77. Copyright © 2014 Elsevier B.V. All rights reserved.

Characterization and expression analysis of genes involved in SUMOylation during embryogenesis in rainbow trout (Oncorhynchus mykiss)

USDA-ARS?s Scientific Manuscript database

SUMOylation is the post-translational modification of proteins by the addition of the small ubiquitin-like modifier (SUMO), which plays an important role in various cellular processes. It has been reported that SUMO and its related proteins are important in diverse reproductive functions such as ovu...

Endothelial nuclear lamina is not required for glucocorticoid receptor nuclear import but does affect receptor-mediated transcription activation.

PubMed

Nayebosadri, Arman; Ji, Julie Y

2013-08-01

The lamina serves to maintain the nuclear structure and stiffness while acting as a scaffold for heterochromatin and many transcriptional proteins. Its role in endothelial mechanotransduction, specifically how nuclear mechanics impact gene regulation under shear stress, is not fully understood. In this study, we successfully silenced lamin A/C in bovine aortic endothelial cells to determine its role in both glucocorticoid receptor (GR) nuclear translocation and glucocorticoid response element (GRE) transcriptional activation in response to dexamethasone and shear stress. Nuclear translocation of GR, an anti-inflammatory nuclear receptor, in response to dexamethasone or shear stress (5, 10, and 25 dyn/cm(2)) was observed via time-lapse cell imaging and quantified using a Bayesian image analysis algorithm. Transcriptional activity of the GRE promoter was assessed using a dual-luciferase reporter plasmid. We found no dependence on nuclear lamina for GR translocation from the cytoplasm into the nucleus. However, the absence of lamin A/C led to significantly increased expression of luciferase under dexamethasone and shear stress induction as well as changes in histone protein function. PCR results for NF-κB inhibitor alpha (NF-κBIA) and dual specificity phosphatase 1 (DUSP1) genes further supported our luciferase data with increased expression in the absence of lamin. Our results suggest that absence of lamin A/C does not hinder passage of GR into the nucleus, but nuclear lamina is important to properly regulate GRE transcription. Nuclear lamina, rather than histone deacetylase (HDAC), is a more significant mediator of shear stress-induced transcriptional activity, while dexamethasone-initiated transcription is more HDAC dependent. Our findings provide more insights into the molecular pathways involved in nuclear mechanotransduction.

Endothelial nuclear lamina is not required for glucocorticoid receptor nuclear import but does affect receptor-mediated transcription activation

PubMed Central

Nayebosadri, Arman

2013-01-01

The lamina serves to maintain the nuclear structure and stiffness while acting as a scaffold for heterochromatin and many transcriptional proteins. Its role in endothelial mechanotransduction, specifically how nuclear mechanics impact gene regulation under shear stress, is not fully understood. In this study, we successfully silenced lamin A/C in bovine aortic endothelial cells to determine its role in both glucocorticoid receptor (GR) nuclear translocation and glucocorticoid response element (GRE) transcriptional activation in response to dexamethasone and shear stress. Nuclear translocation of GR, an anti-inflammatory nuclear receptor, in response to dexamethasone or shear stress (5, 10, and 25 dyn/cm2) was observed via time-lapse cell imaging and quantified using a Bayesian image analysis algorithm. Transcriptional activity of the GRE promoter was assessed using a dual-luciferase reporter plasmid. We found no dependence on nuclear lamina for GR translocation from the cytoplasm into the nucleus. However, the absence of lamin A/C led to significantly increased expression of luciferase under dexamethasone and shear stress induction as well as changes in histone protein function. PCR results for NF-κB inhibitor alpha (NF-κBIA) and dual specificity phosphatase 1 (DUSP1) genes further supported our luciferase data with increased expression in the absence of lamin. Our results suggest that absence of lamin A/C does not hinder passage of GR into the nucleus, but nuclear lamina is important to properly regulate GRE transcription. Nuclear lamina, rather than histone deacetylase (HDAC), is a more significant mediator of shear stress-induced transcriptional activity, while dexamethasone-initiated transcription is more HDAC dependent. Our findings provide more insights into the molecular pathways involved in nuclear mechanotransduction. PMID:23703529

The yeast homologue of the microtubule-associated protein Lis1 interacts with the sumoylation machinery and a SUMO-targeted ubiquitin ligase

PubMed Central

Alonso, Annabel; D'Silva, Sonia; Rahman, Maliha; Meluh, Pam B.; Keeling, Jacob; Meednu, Nida; Hoops, Harold J.; Miller, Rita K.

2012-01-01

Microtubules and microtubule-associated proteins are fundamental for multiple cellular processes, including mitosis and intracellular motility, but the factors that control microtubule-associated proteins (MAPs) are poorly understood. Here we show that two MAPs—the CLIP-170 homologue Bik1p and the Lis1 homologue Pac1p—interact with several proteins in the sumoylation pathway. Bik1p and Pac1p interact with Smt3p, the yeast SUMO; Ubc9p, an E2; and Nfi1p, an E3. Bik1p interacts directly with SUMO in vitro, and overexpression of Smt3p and Bik1p results in its in vivo sumoylation. Modified Pac1p is observed when the SUMO protease Ulp1p is inactivated. Both ubiquitin and Smt3p copurify with Pac1p. In contrast to ubiquitination, sumoylation does not directly tag the substrate for degradation. However, SUMO-targeted ubiquitin ligases (STUbLs) can recognize a sumoylated substrate and promote its degradation via ubiquitination and the proteasome. Both Pac1p and Bik1p interact with the STUbL Nis1p-Ris1p and the protease Wss1p. Strains deleted for RIS1 or WSS1 accumulate Pac1p conjugates. This suggests a novel model in which the abundance of these MAPs may be regulated via STUbLs. Pac1p modification is also altered by Kar9p and the dynein regulator She1p. This work has implications for the regulation of dynein's interaction with various cargoes, including its off-loading to the cortex. PMID:23034179

Mapping the Dynamics of the Glucocorticoid Receptor within the Nuclear Landscape.

PubMed

Stortz, Martin; Presman, Diego M; Bruno, Luciana; Annibale, Paolo; Dansey, Maria V; Burton, Gerardo; Gratton, Enrico; Pecci, Adali; Levi, Valeria

2017-07-24

The distribution of the transcription machinery among different sub-nuclear domains raises the question on how the architecture of the nucleus modulates the transcriptional response. Here, we used fluorescence fluctuation analyses to quantitatively explore the organization of the glucocorticoid receptor (GR) in the interphase nucleus of living cells. We found that this ligand-activated transcription factor diffuses within the nucleus and dynamically interacts with bodies enriched in the coregulator NCoA-2, DNA-dependent foci and chromatin targets. The distribution of the receptor among the nuclear compartments depends on NCoA-2 and the conformation of the receptor as assessed with synthetic ligands and GR mutants with impaired transcriptional abilities. Our results suggest that the partition of the receptor in different nuclear reservoirs ultimately regulates the concentration of receptor available for the interaction with specific targets, and thus has an impact on transcription regulation.

Dose-response approaches for nuclear receptor-mediated ...

EPA Pesticide Factsheets

A public workshop, organized by a Steering Committee of scientists from government, industry, universities, and research organizations, was held at the National Institute of Environmental Health Sciences (NIEHS) in September, 2010. The workshop explored the dose-response implications of toxicant modes of action (MOA) mediated by nuclear receptors. The dominant paradigm in human health risk assessment has been linear extrapolation without a threshold for cancer, and estimation of sub-threshold doses for non-cancer and (in appropriate cases) cancer endpoints. However, recent publications question the application of dose-response modeling approaches with a threshold. The growing body of molecular toxicology information and computational toxicology tools has allowed for exploration of the presence or absence of subthreshold doses for a number of receptor-mediated MOPs. The workshop explored the development of dose-response approaches for nuclear receptor-mediated liver cancer, within a MOA Human Relevance framework (HRF). Case studies addressed activation of the AHR; the CAR/PXR, and the PPARa. This paper describes the workshop process, key issues discussed, and conclusions. The value of an interactive workshop approach to apply current MOA/HRF frameworks was demonstrated. The results may help direct research on the MOA and dose-response of receptor-based toxicity, since there are commonalities for many receptors in the basic pathways involved for late steps in the

SUMO5, a Novel Poly-SUMO Isoform, Regulates PML Nuclear Bodies

PubMed Central

Liang, Ya-Chen; Lee, Chia-Chin; Yao, Ya-Li; Lai, Chien-Chen; Schmitz, M. Lienhard; Yang, Wen-Ming

2016-01-01

Promyelocytic leukemia nuclear bodies (PML-NBs) are PML-based nuclear structures that regulate various cellular processes. SUMOylation, the process of covalently conjugating small ubiquitin-like modifiers (SUMOs), is required for both the formation and the disruption of PML-NBs. However, detailed mechanisms of how SUMOylation regulates these processes remain unknown. Here we report that SUMO5, a novel SUMO variant, mediates the growth and disruption of PML-NBs. PolySUMO5 conjugation of PML at lysine 160 facilitates recruitment of PML-NB components, which enlarges PML-NBs. SUMO5 also increases polySUMO2/3 conjugation of PML, resulting in RNF4-mediated disruption of PML-NBs. The acute promyelocytic leukemia oncoprotein PML-RARα blocks SUMO5 conjugation of PML, causing cytoplasmic displacement of PML and disruption of PML-NBs. Our work not only identifies a new member of the SUMO family but also reveals the mechanistic basis of the PML-NB life cycle in human cells. PMID:27211601

Cross-talk between an activator of nuclear receptors-mediated transcription and the D1 dopamine receptor signaling pathway.

PubMed

Schmidt, Azriel; Vogel, Robert; Rutledge, Su Jane; Opas, Evan E; Rodan, Gideon A; Friedman, Eitan

2005-03-01

Nuclear receptors are transcription factors that usually interact, in a ligand-dependent manner, with specific DNA sequences located within promoters of target genes. The nuclear receptors can also be controlled in a ligand-independent manner via the action of membrane receptors and cellular signaling pathways. 5-Tetradecyloxy-2-furancarboxylic acid (TOFA) was shown to stimulate transcription from the MMTV promoter via chimeric receptors that consist of the DNA binding domain of GR and the ligand binding regions of the PPARbeta or LXRbeta nuclear receptors (GR/PPARbeta and GR/LXRbeta). TOFA and hydroxycholesterols also modulate transcription from NF-kappaB- and AP-1-controlled reporter genes and induce neurite differentiation in PC12 cells. In CV-1 cells that express D(1) dopamine receptors, D(1) dopamine receptor stimulation was found to inhibit TOFA-stimulated transcription from the MMTV promoter that is under the control of chimeric GR/PPARbeta and GR/LXRbeta receptors. Treatment with the D(1) dopamine receptor antagonist, SCH23390, prevented dopamine-mediated suppression of transcription, and by itself increased transcription controlled by GR/LXRbeta. Furthermore, combined treatment of CV-1 cells with TOFA and SCH23390 increased transcription controlled by the GR/LXRbeta chimeric receptor synergistically. The significance of this in vitro synergy was demonstrated in vivo, by the observation that SCH23390 (but not haloperidol)-mediated catalepsy in rats was potentiated by TOFA, thus showing that an agent that mimics the in vitro activities of compounds that activate members of the LXR and PPAR receptor families can influence D1 dopamine receptor elicited responses.

Lipid-sensors, enigmatic-orphan and orphan nuclear receptors as therapeutic targets in breast-cancer.

PubMed

Garattini, Enrico; Bolis, Marco; Gianni', Maurizio; Paroni, Gabriela; Fratelli, Maddalena; Terao, Mineko

2016-07-05

Breast-cancer is heterogeneous and consists of various groups with different biological characteristics. Innovative pharmacological approaches accounting for this heterogeneity are needed. The forty eight human Nuclear-Hormone-Receptors are ligand-dependent transcription-factors and are classified into Endocrine-Receptors, Adopted-Orphan-Receptors (Lipid-sensors and Enigmatic-Orphans) and Orphan-receptors. Nuclear-Receptors represent ideal targets for the design/synthesis of pharmacological ligands. We provide an overview of the literature available on the expression and potential role played by Lipid-sensors, Enigmatic-Orphans and Orphan-Receptors in breast-cancer. The data are complemented by an analysis of the expression levels of each selected Nuclear-Receptor in the PAM50 breast-cancer groups, following re-elaboration of the data publicly available. The major aim is to support the idea that some of the Nuclear-Receptors represent largely unexploited therapeutic-targets in breast-cancer treatment/chemo-prevention. On the basis of our analysis, we conclude that the Lipid-Sensors, NR1C3, NR1H2 and NR1H3 are likely to be onco-suppressors in breast-cancer. The Enigmatic-Orphans, NR1F1 NR2A1 and NR3B3 as well as the Orphan-Receptors, NR0B1, NR0B2, NR1D1, NR2F1, NR2F2 and NR4A3 exert a similar action. These Nuclear-Receptors represent candidates for the development of therapeutic strategies aimed at increasing their expression or activating them in tumor cells. The group of Nuclear-Receptors endowed with potential oncogenic properties consists of the Lipid-Sensors, NR1C2 and NR1I2, the Enigmatic-Orphans, NR1F3, NR3B1 and NR5A2, as well as the Orphan-Receptors, NR2E1, NR2E3 and NR6A1. These oncogenic Nuclear-Receptors should be targeted with selective antagonists, reverse-agonists or agents/strategies capable of reducing their expression in breast-cancer cells.

Sumoylated α-skeletal muscle actin in the skeletal muscle of adult rats.

PubMed

Uda, Munehiro; Kawasaki, Hiroaki; Iizumi, Kyoichi; Shigenaga, Ayako; Baba, Takeshi; Naito, Hisashi; Yoshioka, Toshitada; Yamakura, Fumiyuki

2015-11-01

Skeletal muscles are composed of two major muscle fiber types: slow-twitch oxidative fibers and fast-twitch glycolytic fibers. The proteins in these muscle fibers are known to differ in their expression, relative abundance, and post-translational modifications. In this study, we report a previously unreported post-translational modification of α-skeletal muscle actin in the skeletal muscles of adult male F344 rats in vivo. Using two-dimensional electrophoresis (2D-PAGE), we first examined the differences in the protein expression profiles between the soleus and plantaris muscles. We found higher intensity protein spots at approximately 60 kDa and pH 9 on 2D-PAGE for the soleus muscle compared with the plantaris muscle. These spots were identified as α-skeletal muscle actin by liquid chromatography-nanoelectrospray ionization-tandem mass spectrometry and western blot analyses. In addition, we found that the 60 kDa α-skeletal muscle actin is modified by small ubiquitin-like modifier (SUMO) 1, using 2D-PAGE and western blot analyses. Furthermore, we found that α-skeletal muscle actin with larger molecular weight was localized in the nuclear and cytosol of the skeletal muscle, but not in the myofibrillar fraction by the combination of subcellular fractionation and western blot analyses. These results suggest that α-skeletal muscle actin is modified by SUMO-1 in the skeletal muscles, localized in nuclear and cytosolic fractions, and the extent of this modification is much higher in the slow muscles than in the fast muscles. This is the first study to show the presence of SUMOylated actin in animal tissues.

The GTP-bound and Sumoylated Form of the rab17 Small Molecular Weight GTPase Selectively Binds Syntaxin 2 in Polarized Hepatic WIF-B Cells*

PubMed Central

Striz, Anneliese C.; Tuma, Pamela L.

2016-01-01

A major focus for our laboratory is identifying the molecules and mechanisms that regulate polarized apical protein sorting in hepatocytes, the major epithelial cells of the liver. These trafficking pathways are regulated, in part, by small molecular weight rab GTPases. We chose to investigate rab17, whose expression is restricted to polarized epithelial cells, is enriched in liver, and has been implicated in regulating basolateral to apical transcytosis. To initiate our studies, we generated three recombinant adenoviruses expressing wild type, constitutively active (GTP bound), or dominant-negative (GDP bound) rab17. Immunoblotting revealed rab17 immunoreactive species at 25 kDa (the predicted rab17 molecular mass) and 40 kDa. We determined that mono-sumoylation of the 25-kDa rab17 is responsible for the shift in molecular mass, and that rab17 prenylation is required for sumoylation. We further determined that sumoylation selectively promotes interactions with syntaxin 2 (but not syntaxins 3 or 4) and that these interactions are nucleotide dependent. Furthermore, a K68R-mutated rab17 led to the redistribution of syntaxin 2 and 5′ nucleotidase from the apical membrane to subapical puncta, whereas multidrug resistance protein 2 distributions were not changed. Together these data are consistent with the proposed role of rab17 in vesicle fusion with the apical plasma membrane and further implicate sumoylation as an important mediator of protein-protein interactions. The selectivity in syntaxin binding and apical protein redistribution further suggests that rab17 and syntaxin 2 mediate fusion of transcytotic vesicles at the apical surface. PMID:26957544

Exclusive nuclear location of estrogen receptors in Squalus testis.

PubMed Central

Callard, G V; Mak, P

1985-01-01

An estrogen (E)-binding molecule having both occupied and unoccupied sites is restricted to nuclear subfractions in the testis of the spiny dogfish (Squalus acanthias). We investigated the hypothesis that a species characterized by high body-fluid osmolarity (1010 mosM) has an estrogen receptor (ER) that binds to chromatin with high affinity and consequently resists redistribution during tissue processing. Although the steroid binding and sedimentation properties of the Squalus nuclear ER conformed to those of classical ER, its elution maximum from DNA-cellulose was unusually high (0.55 M NaCl). A tendency to adhere tightly to cell nuclei was reflected in the high salt concentration (0.43 M KCl) required to extract 50% of the receptors from the nuclear compartment during homogenization and in the stability of the nuclear ER population in the presence of high concentrations of a nonionic solute (urea) or increased buffer volume. Mixing and redistribution experiments showed that nuclear ER could be quantitatively and qualitatively measured in cytosolic extracts, ruling out the possibility that soluble receptors were being masked. Although Squalus oviduct ER was similar to that of testis, ER in the testis and liver of a related elasmobranch (Potamotrygon) that maintains osmotic equilibrium at 300 mosM more closely resembled mammalian ER in its elution maximum from DNA-cellulose (0.22 M NaCl) and cytosolic/nuclear ratios in low-salt buffers. We conclude that Squalus testis has a single ER pool located exclusively in the nuclear compartment. These observations support a revised concept of steroid action and further indicate that the chromatin affinity of the hormone-ER complex is an important factor in determining subfractional distribution during tissue processing. PMID:3856265

Orphan Nuclear Receptors as Targets for Drug Development

PubMed Central

Mukherjee, Subhajit

2012-01-01

Orphan nuclear receptors regulate diverse biological processes. These important molecules are ligand-activated transcription factors that act as natural sensors for a wide range of steroid hormones and xenobiotic ligands. Because of their importance in regulating various novel signaling pathways, recent research has focused on identifying xenobiotics targeting these receptors for the treatment of multiple human diseases. In this review, we will highlight these receptors in several physiologic and pathophysiologic actions and demonstrate how their functions can be exploited for the successful development of newer drugs. PMID:20372994

Specific regulation of thermosensitive lipid droplet fusion by a nuclear hormone receptor pathway

PubMed Central

Li, Shiwei; Li, Qi; Kong, Yuanyuan; Wu, Shuang; Cui, Qingpo; Zhang, Mingming; Zhang, Shaobing O.

2017-01-01

Nuclear receptors play important roles in regulating fat metabolism and energy production in humans. The regulatory functions and endogenous ligands of many nuclear receptors are still unidentified, however. Here, we report that CYP-37A1 (ortholog of human cytochrome P450 CYP4V2), EMB-8 (ortholog of human P450 oxidoreductase POR), and DAF-12 (homolog of human nuclear receptors VDR/LXR) constitute a hormone synthesis and nuclear receptor pathway in Caenorhabditis elegans. This pathway specifically regulates the thermosensitive fusion of fat-storing lipid droplets. CYP-37A1, together with EMB-8, synthesizes a lipophilic hormone not identical to Δ7-dafachronic acid, which represses the fusion-promoting function of DAF-12. CYP-37A1 also negatively regulates thermotolerance and lifespan at high temperature in a DAF-12–dependent manner. Human CYP4V2 can substitute for CYP-37A1 in C. elegans. This finding suggests the existence of a conserved CYP4V2-POR–nuclear receptor pathway that functions in converting multilocular lipid droplets to unilocular ones in human cells; misregulation of this pathway may lead to pathogenic fat storage. PMID:28760992

Specific regulation of thermosensitive lipid droplet fusion by a nuclear hormone receptor pathway.

PubMed

Li, Shiwei; Li, Qi; Kong, Yuanyuan; Wu, Shuang; Cui, Qingpo; Zhang, Mingming; Zhang, Shaobing O

2017-08-15

Nuclear receptors play important roles in regulating fat metabolism and energy production in humans. The regulatory functions and endogenous ligands of many nuclear receptors are still unidentified, however. Here, we report that CYP-37A1 (ortholog of human cytochrome P450 CYP4V2), EMB-8 (ortholog of human P450 oxidoreductase POR), and DAF-12 (homolog of human nuclear receptors VDR/LXR) constitute a hormone synthesis and nuclear receptor pathway in Caenorhabditis elegans This pathway specifically regulates the thermosensitive fusion of fat-storing lipid droplets. CYP-37A1, together with EMB-8, synthesizes a lipophilic hormone not identical to Δ7-dafachronic acid, which represses the fusion-promoting function of DAF-12. CYP-37A1 also negatively regulates thermotolerance and lifespan at high temperature in a DAF-12-dependent manner. Human CYP4V2 can substitute for CYP-37A1 in C. elegans This finding suggests the existence of a conserved CYP4V2-POR-nuclear receptor pathway that functions in converting multilocular lipid droplets to unilocular ones in human cells; misregulation of this pathway may lead to pathogenic fat storage.

Nuclear Receptors, Mitochondria, and Lipid Metabolism

PubMed Central

Alaynick, William A.

2009-01-01

Lipid metabolism is a continuum from emulsification and uptake of lipids in the intestine to cellular uptake and transport to compartments such as mitochondria. Whether fats are shuttled into lipid droplets in adipose tissue or oxidized in mitochondria and peroxisomes depends on metabolic substrate availability, energy balance and endocrine signaling of the organism. Several members of the nuclear hormone receptor superfamily are lipid-sensing factors that affect all aspects of lipid metabolism. The physiologic actions of glandular hormones (e.g. thyroid, mineralocorticoid and glucocorticoid), vitamins (e.g. vitamins A and D) and reproductive hormones (e.g. progesterone, estrogen and testosterone) and their cognate receptors are well established. The peroxisome proliferator activated receptors (PPARs) and Liver X receptors (LXRs), acting in concert with PPARγ Coactivator 1α (PGC-1α), have been shown to regulate insulin sensitivity and lipid handling. These receptors are the focus of intense pharmacologic studies to expand the armamentarium of small molecule ligands to treat diabetes and the metabolic syndrome (hypertension, insulin resistance, hyperglycemia, dyslipidemia, and obesity). Recently, additional partners of PGC-1α have moved to the forefront of metabolic research, the Estrogen-related Receptors (ERRs). Although no endogenous ligands for these receptors have been identified, phenotypic analyses of knockout mouse models demonstrate an important role for these molecules in substrate sensing and handling as well as mitochondrial function. PMID:18375192

Tracking Progesterone Receptor-Mediated Actions in Breast Cancer

PubMed Central

Knutson, Todd P.; Lange, Carol A.

2014-01-01

Ovarian steroid hormones contribute to breast cancer initiation and progression primarily through the actions of their nuclear transcription factors, the estrogen receptor alpha (ERα) and progesterone receptors (PRs). These receptors are important drivers of the luminal A and B subtypes of breast cancer, where estrogen-blocking drugs have been effective endocrine therapies for patients with these tumors. However, many patients do not respond, or become resistant to treatment. When endocrine therapies fail, the luminal subtypes of breast cancer are more difficult to treat because these subtypes are among the most heterogeneous in terms of mutation diversity and gene expression profiles. Recent evidence suggests that progestin and PR actions may be important drivers of luminal breast cancers. Clinical trial data has demonstrated that hormone replacement therapy with progestins drives invasive breast cancer and results in greater mortality. PR transcriptional activity is dependent upon cross-talk with growth factor signaling pathways that alter PR phosphorylation, acetylation, or SUMOylation as mechanisms for regulating PR target gene selection required for increased cell proliferation and survival. Site-specific PR phosphorylation is the primary driver of gene-selective PR transcriptional activity. However, PR phosphorylation and heightened transcriptional activity is coupled to rapid PR protein degradation; the range of active PR detected in tumors is likely to be dynamic. Thus, PR target gene signatures may provide a more accurate means of tracking PR’s contribution to tumor progression rather than standard clinical protein-based (IHC) assays. Further development of antiprogestin therapies should be considered along side antiestrogens and aromatase inhibitors. PMID:24291072

Brain nuclear receptors and body weight regulation

USDA-ARS?s Scientific Manuscript database

Neural pathways, especially those in the hypothalamus, integrate multiple nutritional, hormonal, and neural signals, resulting in the coordinated control of body weight balance and glucose homeostasis. Nuclear receptors (NRs) sense changing levels of nutrients and hormones, and therefore play essent...

The Orphan Nuclear Receptors at Their 25th Year Reunion

PubMed Central

Mullican, Shannon E.; DiSpirito, Joanna R.; Lazar, Mitchell A.

2013-01-01

The Nuclear Receptor superfamily includes many receptors identified based on their similarity to steroid hormone receptors but without a known ligand. The study of how these receptors are diversely regulated to interact with genomic regions to control a plethora of biological processes has provided critical insight into development, physiology and the molecular pathology of disease. Here we provide a compendium of these so-called Orphan Receptors, and focus on what has been learned about their modes of action, physiological functions, and therapeutic promise. PMID:24096517

Quantitative SUMO proteomics reveals the modulation of several PML nuclear body associated proteins and an anti-senescence function of UBC9.

PubMed

McManus, Francis P; Bourdeau, Véronique; Acevedo, Mariana; Lopes-Paciencia, Stéphane; Mignacca, Lian; Lamoliatte, Frédéric; Rojas Pino, John W; Ferbeyre, Gerardo; Thibault, Pierre

2018-05-17

Several regulators of SUMOylation have been previously linked to senescence but most targets of this modification in senescent cells remain unidentified. Using a two-step purification of a modified SUMO3, we profiled the SUMO proteome of senescent cells in a site-specific manner. We identified 25 SUMO sites on 23 proteins that were significantly regulated during senescence. Of note, most of these proteins were PML nuclear body (PML-NB) associated, which correlates with the increased number and size of PML-NBs observed in senescent cells. Interestingly, the sole SUMO E2 enzyme, UBC9, was more SUMOylated during senescence on its Lys-49. Functional studies of a UBC9 mutant at Lys-49 showed a decreased association to PML-NBs and the loss of UBC9's ability to delay senescence. We thus propose both pro- and anti-senescence functions of protein SUMOylation.

Purification and characterization of rat liver nuclear thyroid hormone receptors.

PubMed Central

Ichikawa, K; DeGroot, L J

1987-01-01

Nuclear thyroid hormone receptor was purified to 904 pmol of L-3,5,3'-triiodothyronine (T3) binding capacity per mg of protein with 2.5-5.2% recovery by sequentially using hydroxylapatite column chromatography, ammonium sulfate precipitation, Sephadex G-150 gel filtration, DNA-cellulose column chromatography, DEAE-Sephadex column chromatography, and heparin-Sepharose column chromatography. Assuming that one T3 molecule binds to the 49,000-Da unit of the receptor, we reproducibly obtained 6.4-14.7 micrograms of receptor protein with 4.2-4.9% purity from 4-5 kg of rat liver. Elution of receptor from the heparin-Sepharose column was performed using 10 mM pyridoxal 5'-phosphate, which was observed to diminish binding of receptor to heparin-Sepharose or DNA-cellulose. This effect was specific for pyridoxal 5'-phosphate, since related compounds were not effective. Purified receptor bound T3 with high affinity (6.0 X 10(9) liter/mol), and the order of affinity of iodothyronine analogues to purified receptor was identical to that observed with crude receptor preparations [3,5,3'-triiodothyroacetic acid greater than L-T3 greater than D-3,5,3'-triiodothyronine (D-T3) greater than L-thyroxine greater than D-thyroxine]. Purified receptor had a sedimentation coefficient of 3.4 S, Stokes radius of 34 A, and calculated molecular mass of 49,000. Among several bands identified by silver staining after electrophoresis in NaDodSO4/polyacrylamide gels, one 49,000-Da protein showed photoaffinity labeling with [125I]thyroxine that was displaceable with excess unlabeled T3. The tryptic fragment and endogenous proteinase-digested fragment of the affinity-labeled receptor showed saturable binding in 27,000-Da and 36,000-Da peptides, respectively. These molecular masses are in agreement with estimates from gel filtration and gradient sedimentation, indicating that affinity labeling occurred at the hormone binding domain of nuclear thyroid hormone receptor. This procedure reproducibly

The Nuclear Receptor HIZR-1 Uses Zinc as a Ligand to Mediate Homeostasis in Response to High Zinc

PubMed Central

Warnhoff, Kurt; Roh, Hyun C.; Kocsisova, Zuzana; Tan, Chieh-Hsiang; Morrison, Andrew; Croswell, Damari; Schneider, Daniel L.; Kornfeld, Kerry

2017-01-01

Nuclear receptors were originally defined as endocrine sensors in humans, leading to the identification of the nuclear receptor superfamily. Despite intensive efforts, most nuclear receptors have no known ligand, suggesting new ligand classes remain to be discovered. Furthermore, nuclear receptors are encoded in the genomes of primitive organisms that lack endocrine signaling, suggesting the primordial function may have been environmental sensing. Here we describe a novel Caenorhabditis elegans nuclear receptor, HIZR-1, that is a high zinc sensor in an animal and the master regulator of high zinc homeostasis. The essential micronutrient zinc acts as a HIZR-1 ligand, and activated HIZR-1 increases transcription of genes that promote zinc efflux and storage. The results identify zinc as the first inorganic molecule to function as a physiological ligand for a nuclear receptor and direct environmental sensing as a novel function of nuclear receptors. PMID:28095401

Using Nuclear Receptor Activity to Stratify Hepatocarcinogens

EPA Science Inventory

Nuclear receptors (NR) are a superfamily of ligand-activated transcription factors that control a range of cellular processes. Persistent stimulation of some NR is a non-genotoxic mechanism of rodent liver cancer with unclear relevance to humans. Here we report on a systematic an...

Sumoylation of the net inhibitory domain (NID) is stimulated by PIAS1 and has a negative effect on the transcriptional activity of Net.

PubMed

Wasylyk, Christine; Criqui-Filipe, Paola; Wasylyk, Bohdan

2005-01-27

Net (Elk-3, Sap-2, Erp) and the related ternary complex factors Elk-1 and Sap-1 are effectors of multiple signalling pathways at the transcriptional level and play a key role in the dynamic regulation of gene expression. Net is distinct from Elk-1 and Sap-1, in that it is a strong repressor of transcription that is converted to an activator by the Ras/Erk signalling pathway. Two autonomous repression domains of Net, the NID and the CID, mediate repression. We have previously shown that the co-repressor CtBP is implicated in repression by the CID. In this report we show that repression by the NID involves a different pathway, sumoylation by Ubc9 and PIAS1. PIAS1 interacts with the NID in the two-hybrid assay and in vitro. Ubc9 and PIAS1 stimulate sumoylation in vivo of lysine 162 in the NID. Sumoylation of lysine 162 increases repression by Net and decreases the positive activity of Net. These results increase our understanding of how one of the ternary complex factors regulates transcription, and contribute to the understanding of how different domains of a transcription factor participate in the complexity of regulation of gene expression.

Natural compounds regulate energy metabolism by the modulating the activity of lipid-sensing nuclear receptors.

PubMed

Goto, Tsuyoshi; Kim, Young-Il; Takahashi, Nobuyuki; Kawada, Teruo

2013-01-01

Obesity causes excess fat accumulation in various tissues, most notoriously in the adipose tissue, along with other insulin-responsive organs such as skeletal muscle and the liver, which predisposes an individual to the development of metabolic abnormalities. The molecular mechanisms underlying obesity-induced metabolic abnormalities have not been completely elucidated; however, in recent years, the search for therapies to prevent the development of obesity and obesity-associated metabolic disorders has increased. It is known that several nuclear receptors, when activated by specific ligands, regulate carbohydrate and lipid metabolism at the transcriptional level. The expression of lipid metabolism-related enzymes is directly regulated by the activity of various nuclear receptors via their interaction with specific response elements in promoters of those genes. Many natural compounds act as ligands of nuclear receptors and regulate carbohydrate and lipid metabolism by regulating the activities of these nuclear receptors. In this review, we describe our current knowledge of obesity, the role of lipid-sensing nuclear receptors in energy metabolism, and several examples of food factors that act as agonists or antagonists of nuclear receptors, which may be useful for the management of obesity and the accompanying energy metabolism abnormalities. © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

ROR nuclear receptors: structures, related diseases, and drug discovery

PubMed Central

Zhang, Yan; Luo, Xiao-yu; Wu, Dong-hai; Xu, Yong

2015-01-01

Nuclear receptors (NRs) are ligand-regulated transcription factors that regulate metabolism, development and immunity. The NR superfamily is one of the major classes of drug targets for human diseases. Retinoic acid receptor-related orphan receptor (ROR) α, β and γ belong to the NR superfamily, and these receptors are still considered as 'orphan' receptors because the identification of their endogenous ligands has been controversial. Recent studies have demonstrated that these receptors are regulated by synthetic ligands, thus emerge as important drug targets for the treatment of multiple sclerosis, rheumatoid arthritis, psoriasis, etc. Studying the structural basis and ligand development of RORs will pave the way for a better understanding of the roles of these receptors in human diseases. Here, we review the structural basis, disease relevance, strategies for ligand identification, and current status of development of therapeutic ligands for RORs. PMID:25500868

Daphnia HR96 is a Promiscuous Xenobiotic and Endobiotic Nuclear Receptor

PubMed Central

Karimullina, Elina; Li, Yangchun; Ginjupalli, Gautam; Baldwin, William S.

2012-01-01

Daphnia pulex is the first crustacean to have its genome sequenced. The genome project provides new insight and data into how an aquatic crustacean may respond to environmental stressors, including toxicants. We cloned Daphnia pulex HR96 (DappuHR96), a nuclear receptor orthologous to the CAR/PXR/VDR group of nuclear receptors. In Drosophila melanogaster, (hormone receptor 96) HR96 responds to phenobarbital exposure and has been hypothesized as a toxicant receptor. Therefore, we set up a transactivation assay to test whether DappuHR96 is a promiscuous receptor activated by xenobiotics and endobiotics similar to the constitutive androstane receptor (CAR) and the pregnane X-receptor (PXR). Transactivation assays performed with a GAL4-HR96 chimera demonstrate that HR96 is a promiscuous toxicant receptor activated by a diverse set of chemicals such as pesticides, hormones, and fatty acids. Several environmental toxicants activate HR96 including estradiol, pyriproxyfen, chlorpyrifos, atrazine, and methane arsonate. We also observed repression of HR96 activity by chemicals such as triclosan, androstanol, and fluoxetine. Nearly 50% of the chemicals tested activated or inhibited HR96. Interestingly, unsaturated fatty acids were common activators or inhibitors of HR96 activity, indicating a link between diet and toxicant response. The omega-6 and omega-9 unsaturated fatty acids linoleic and oleic acid activated HR96, but the omega-3 unsaturated fatty acids alpha-linolenic acid and docosahexaenoic acid inhibited HR96, suggesting that these two distinct sets of lipids perform opposing roles in Daphnia physiology. This also provides a putative mechanism by which the ratio of dietary unsaturated fats may affect the ability of an organism to respond to a toxic insult. In summary, HR96 is a promiscuous nuclear receptor activated by numerous endo- and xenobiotics. PMID:22466357

Plant nuclear hormone receptors: a role for small molecules in protein-protein interactions.

PubMed

Lumba, Shelley; Cutler, Sean; McCourt, Peter

2010-01-01

Plant hormones are a group of chemically diverse small molecules that direct processes ranging from growth and development to biotic and abiotic stress responses. Surprisingly, genome analyses suggest that classic animal nuclear hormone receptor homologs do not exist in plants. It now appears that plants have co-opted several protein families to perceive hormones within the nucleus. In one solution to the problem, the hormones auxin and jasmonate (JA) act as “molecular glue” that promotes protein-protein interactions between receptor F-boxes and downstream corepressor targets. In another solution, gibberellins (GAs) bind and elicit a conformational change in a novel soluble receptor family related to hormone-sensitive lipases. Abscisic acid (ABA), like GA, also acts through an allosteric mechanism involving a START-domain protein. The molecular identification of plant nuclear hormone receptors will allow comparisons with animal nuclear receptors and testing of fundamental questions about hormone function in plant development and evolution.

Retinoids induce integrin-independent lymphocyte adhesion through RAR-α nuclear receptor activity

DOE Office of Scientific and Technical Information (OSTI.GOV)

Whelan, Jarrett T.; Wang, Lei; Chen, Jianming

2014-11-28

Highlights: • Transcription and translation are required for retinoid-induced lymphocyte adhesion. • RAR activation is sufficient to induced lymphocyte cell adhesion. • Vitamin D derivatives inhibit RAR-prompted lymphocyte adhesion. • Adhesion occurs through a novel binding site within ADAM disintegrin domains. • RARα is a key nuclear receptor for retinoid-dependent lymphocyte cell adhesion. - Abstract: Oxidative metabolites of vitamin A, in particular all-trans-retinoic acid (atRA), have emerged as key factors in immunity by specifying the localization of immune cells to the gut. Although it is appreciated that isomers of retinoic acid activate the retinoic acid receptor (RAR) and retinoid Xmore » receptor (RXR) family of nuclear receptors to elicit cellular changes, the molecular details of retinoic acid action remain poorly defined in immune processes. Here we employ a battery of agonists and antagonists to delineate the specific nuclear receptors utilized by retinoids to evoke lymphocyte cell adhesion to ADAM (adisintegrin and metalloprotease) protein family members. We report that RAR agonism is sufficient to promote immune cell adhesion in both immortal and primary immune cells. Interestingly, adhesion occurs independent of integrin function, and mutant studies demonstrate that atRA-induced adhesion to ADAM members required a distinct binding interface(s) as compared to integrin recognition. Anti-inflammatory corticosteroids as well as 1,25-(OH){sub 2}D{sub 3}, a vitamin D metabolite that prompts immune cell trafficking to the skin, potently inhibited the observed adhesion. Finally, our data establish that induced adhesion was specifically attributable to the RAR-α receptor isotype. The current study provides novel molecular resolution as to which nuclear receptors transduce retinoid exposure into immune cell adhesion.« less

Nuclear receptor/microRNA circuitry links muscle fiber type to energy metabolism.

PubMed

Gan, Zhenji; Rumsey, John; Hazen, Bethany C; Lai, Ling; Leone, Teresa C; Vega, Rick B; Xie, Hui; Conley, Kevin E; Auwerx, Johan; Smith, Steven R; Olson, Eric N; Kralli, Anastasia; Kelly, Daniel P

2013-06-01

The mechanisms involved in the coordinate regulation of the metabolic and structural programs controlling muscle fitness and endurance are unknown. Recently, the nuclear receptor PPARβ/δ was shown to activate muscle endurance programs in transgenic mice. In contrast, muscle-specific transgenic overexpression of the related nuclear receptor, PPARα, results in reduced capacity for endurance exercise. We took advantage of the divergent actions of PPARβ/δ and PPARα to explore the downstream regulatory circuitry that orchestrates the programs linking muscle fiber type with energy metabolism. Our results indicate that, in addition to the well-established role in transcriptional control of muscle metabolic genes, PPARβ/δ and PPARα participate in programs that exert opposing actions upon the type I fiber program through a distinct muscle microRNA (miRNA) network, dependent on the actions of another nuclear receptor, estrogen-related receptor γ (ERRγ). Gain-of-function and loss-of-function strategies in mice, together with assessment of muscle biopsies from humans, demonstrated that type I muscle fiber proportion is increased via the stimulatory actions of ERRγ on the expression of miR-499 and miR-208b. This nuclear receptor/miRNA regulatory circuit shows promise for the identification of therapeutic targets aimed at maintaining muscle fitness in a variety of chronic disease states, such as obesity, skeletal myopathies, and heart failure.

Discovering relationships between nuclear receptor signaling pathways, genes, and tissues in Transcriptomine.

PubMed

Becnel, Lauren B; Ochsner, Scott A; Darlington, Yolanda F; McOwiti, Apollo; Kankanamge, Wasula H; Dehart, Michael; Naumov, Alexey; McKenna, Neil J

2017-04-25

We previously developed a web tool, Transcriptomine, to explore expression profiling data sets involving small-molecule or genetic manipulations of nuclear receptor signaling pathways. We describe advances in biocuration, query interface design, and data visualization that enhance the discovery of uncharacterized biology in these pathways using this tool. Transcriptomine currently contains about 45 million data points encompassing more than 2000 experiments in a reference library of nearly 550 data sets retrieved from public archives and systematically curated. To make the underlying data points more accessible to bench biologists, we classified experimental small molecules and gene manipulations into signaling pathways and experimental tissues and cell lines into physiological systems and organs. Incorporation of these mappings into Transcriptomine enables the user to readily evaluate tissue-specific regulation of gene expression by nuclear receptor signaling pathways. Data points from animal and cell model experiments and from clinical data sets elucidate the roles of nuclear receptor pathways in gene expression events accompanying various normal and pathological cellular processes. In addition, data sets targeting non-nuclear receptor signaling pathways highlight transcriptional cross-talk between nuclear receptors and other signaling pathways. We demonstrate with specific examples how data points that exist in isolation in individual data sets validate each other when connected and made accessible to the user in a single interface. In summary, Transcriptomine allows bench biologists to routinely develop research hypotheses, validate experimental data, or model relationships between signaling pathways, genes, and tissues. Copyright © 2017, American Association for the Advancement of Science.

Microsomal receptor for steroid hormones: functional implications for nuclear activity.

PubMed

Muldoon, T G; Watson, G H; Evans, A C; Steinsapir, J

1988-01-01

microsomal binding sites extracted. These observations suggest three possible roles for the microsomal receptor-like proteins: (a) modulation of estrogen access to nuclear binding sites; (b) formation of functional complexes which diffuse to other extranuclear sites to alter non-genomic cellular processes; (c) regulation of nuclear concentration of estrogen-receptor complexes by virtue of producing microsomal acceptor sites for uptake of free or loosely associated nuclear complexes, previously thought to exist in the cytoplasm.

Nuclear Receptors in Bone Physiology and Diseases

PubMed Central

Youn, Min-Young; Inoue, Kazuki; Takada, Ichiro; Kouzmenko, Alexander; Kato, Shigeaki

2013-01-01

During the last decade, our view on the skeleton as a mere solid physical support structure has been transformed, as bone emerged as a dynamic, constantly remodeling tissue with systemic regulatory functions including those of an endocrine organ. Reflecting this remarkable functional complexity, distinct classes of humoral and intracellular regulatory factors have been shown to control vital processes in the bone. Among these regulators, nuclear receptors (NRs) play fundamental roles in bone development, growth, and maintenance. NRs are DNA-binding transcription factors that act as intracellular transducers of the respective ligand signaling pathways through modulation of expression of specific sets of cognate target genes. Aberrant NR signaling caused by receptor or ligand deficiency may profoundly affect bone health and compromise skeletal functions. Ligand dependency of NR action underlies a major strategy of therapeutic intervention to correct aberrant NR signaling, and significant efforts have been made to design novel synthetic NR ligands with enhanced beneficial properties and reduced potential negative side effects. As an example, estrogen deficiency causes bone loss and leads to development of osteoporosis, the most prevalent skeletal disorder in postmenopausal women. Since administration of natural estrogens for the treatment of osteoporosis often associates with undesirable side effects, several synthetic estrogen receptor ligands have been developed with higher therapeutic efficacy and specificity. This review presents current progress in our understanding of the roles of various nuclear receptor-mediated signaling pathways in bone physiology and disease, and in development of advanced NR ligands for treatment of common skeletal disorders. PMID:23589826

Nuclear hormone receptor coregulator: role in hormone action, metabolism, growth, and development.

PubMed

Mahajan, Muktar A; Samuels, Herbert H

2005-06-01

Nuclear hormone receptor coregulator (NRC) (also referred to as activating signal cointegrator-2, thyroid hormone receptor-binding protein, peroxisome proliferator activating receptor-interacting protein, and 250-kDa receptor associated protein) belongs to a growing class of nuclear cofactors widely known as coregulators or coactivators that are necessary for transcriptional activation of target genes. The NRC gene is also amplified and overexpressed in breast, colon, and lung cancers. NRC is a 2063-amino acid protein that harbors a potent N-terminal activation domain (AD1) and a second more centrally located activation domain (AD2) that is rich in Glu and Pro. Near AD2 is a receptor-interacting domain containing an LxxLL motif (LxxLL-1), which interacts with a wide variety of ligand-bound nuclear hormone receptors with high affinity. A second LxxLL motif (LxxLL-2) located in the C-terminal region of NRC is more restricted in its nuclear hormone receptor specificity. The intrinsic activation potential of NRC is regulated by a C-terminal serine, threonine, leucine-regulatory domain. The potential role of NRC as a cointegrator 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 transcriptional regulators including CBP/p300. Recent studies in mice indicate that deletion of both NRC alleles leads to embryonic lethality resulting from general growth retardation coupled with developmental defects in the heart, liver, brain, and placenta. NRC(-/-) mouse embryo fibroblasts spontaneously undergo apoptosis, indicating the importance of NRC as a prosurvival and antiapoptotic gene. Studies with 129S6 NRC(+/-) mice indicate that NRC is a pleiotropic regulator that is involved in growth, development, reproduction, metabolism, and wound healing.

Nuclear receptors and pathogenesis of pancreatic cancer

PubMed Central

Polvani, Simone; Tarocchi, Mirko; Tempesti, Sara; Galli, Andrea

2014-01-01

Pancreatic ductal adenocarcinoma (PDAC) is a devastating disease with a median overall survival time of 5 mo and the five years survival less than 5%, a rate essentially unchanged over the course of the years. A well defined progression model of accumulation of genetic alterations ranging from single point mutations to gross chromosomal abnormalities has been introduced to describe the origin of this disease. However, due to the its subtle nature and concurring events PDAC cure remains elusive. Nuclear receptors (NR) are members of a large superfamily of evolutionarily conserved ligand-regulated DNA-binding transcription factors functionally involved in important cellular functions ranging from regulation of metabolism, to growth and development. Given the nature of their ligands, NR are very tempting drug targets and their pharmacological modulation has been widely exploited for the treatment of metabolic and inflammatory diseases. There are now clear evidences that both classical ligand-activated and orphan NR are involved in the pathogenesis of PDAC from its very early stages; nonetheless many aspects of their role are not fully understood. The purpose of this review is to highlight the striking connections that link peroxisome proliferator activated receptors, retinoic acid receptors, retinoid X receptor, androgen receptor, estrogen receptors and the orphan NR Nur, chicken ovalbumin upstream promoter transcription factor II and the liver receptor homologue-1 receptor to PDAC development, connections that could lead to the identification of novel therapies for this disease. PMID:25232244

Genome-wide identification of nuclear receptor (NR) superfamily genes in the copepod Tigriopus japonicus.

PubMed

Hwang, Dae-Sik; Lee, Bo-Young; Kim, Hui-Su; Lee, Min Chul; Kyung, Do-Hyun; Om, Ae-Son; Rhee, Jae-Sung; Lee, Jae-Seong

2014-11-18

Nuclear receptors (NRs) are a large superfamily of proteins defined by a DNA-binding domain (DBD) and a ligand-binding domain (LBD). They function as transcriptional regulators to control expression of genes involved in development, homeostasis, and metabolism. The number of NRs differs from species to species, because of gene duplications and/or lineage-specific gene losses during metazoan evolution. Many NRs in arthropods interact with the ecdysteroid hormone and are involved in ecdysone-mediated signaling in arthropods. The nuclear receptor superfamily complement has been reported in several arthropods, including crustaceans, but not in copepods. We identified the entire NR repertoire of the copepod Tigriopus japonicus, which is an important marine model species for ecotoxicology and environmental genomics. Using whole genome and transcriptome sequences, we identified a total of 31 nuclear receptors in the genome of T. japonicus. Nomenclature of the nuclear receptors was determined based on the sequence similarities of the DNA-binding domain (DBD) and ligand-binding domain (LBD). The 7 subfamilies of NRs separate into five major clades (subfamilies NR1, NR2, NR3, NR4, and NR5/6). Although the repertoire of NR members in, T. japonicus was similar to that reported for other arthropods, there was an expansion of the NR1 subfamily in Tigriopus japonicus. The twelve unique nuclear receptors identified in T. japonicus are members of NR1L. This expansion may be a unique lineage-specific feature of crustaceans. Interestingly, E78 and HR83, which are present in other arthropods, were absent from the genomes of T. japonicus and two congeneric copepod species (T. japonicus and Tigriopus californicus), suggesting copepod lineage-specific gene loss. We identified all NR receptors present in the copepod, T. japonicus. Knowledge of the copepod nuclear receptor repertoire will contribute to a better understanding of copepod- and crustacean-specific NR evolution.

BXR, an embryonic orphan nuclear receptor activated by a novel class of endogenous benzoate metabolites

PubMed Central

Blumberg, Bruce; Kang, Heonjoong; Bolado, Jack; Chen, Hongwu; Craig, A. Grey; Moreno, Tanya A.; Umesono, Kazuhiko; Perlmann, Thomas; De Robertis, Eddy M.; Evans, Ronald M.

1998-01-01

Nuclear receptors are ligand-modulated transcription factors that respond to steroids, retinoids, and thyroid hormones to control development and body physiology. Orphan nuclear receptors, which lack identified ligands, provide a unique, and largely untapped, resource to discover new principles of physiologic homeostasis. We describe the isolation and characterization of the vertebrate orphan receptor, BXR, which heterodimerizes with RXR and binds high-affinity DNA sites composed of a variant thyroid hormone response element. A bioactivity-guided screen of embryonic extracts revealed that BXR is activatable by low-molecular-weight molecules with spectral patterns distinct from known nuclear receptor ligands. Mass spectrometry and 1H NMR analysis identified alkyl esters of amino and hydroxy benzoic acids as potent, stereoselective activators. In vitro cofactor association studies, along with competable binding of radiolabeled compounds, establish these molecules as bona fide ligands. Benzoates comprise a new molecular class of nuclear receptor ligand and their activity suggests that BXR may control a previously unsuspected vertebrate signaling pathway. PMID:9573044

Nuclear Receptor Activity and Liver Cancer Lesion Progression

EPA Science Inventory

Nuclear receptors (NRs) are ligand-activated transcription factors that control diverse cellular processes. Chronic stimulation of some NRs is a non-genotoxic mechanism of rodent liver cancer with unclear relevance to humans. We explored this question using human CAR, PXR, PPARα,...

Design of selective nuclear receptor modulators: RAR and RXR as a case study.

PubMed

de Lera, Angel R; Bourguet, William; Altucci, Lucia; Gronemeyer, Hinrich

2007-10-01

Retinoic acid receptors (RARs) and retinoid X receptors (RXRs) are members of the nuclear receptor superfamily whose effects on cell growth and survival can be modulated therapeutically by small-molecule ligands. Although compounds that target these receptors are powerful anticancer drugs, their use is limited by toxicity. An improved understanding of the structural biology of RXRs and RARs and recent advances in the chemical synthesis of modified retinoid and rexinoid ligands should enable the rational design of more selective agents that might overcome such problems. Here, we review structural data for RXRs and RARs, discuss strategies in the design of selective RXR and RAR modulators, and consider lessons that can be learned for the design of selective nuclear-receptor modulators in general.

PML nuclear bodies.

PubMed

Lallemand-Breitenbach, Valérie; de Thé, Hugues

2010-05-01

PML nuclear bodies are matrix-associated domains that recruit an astonishing variety of seemingly unrelated proteins. Since their discovery in the early 1960s, PML bodies have fascinated cell biologists because of their beauty and their tight association with cellular disorders. The identification of PML, a gene involved in an oncogenic chromosomal translocation, as the key organizer of these domains drew instant interest onto them. The multiple levels of PML body regulation by a specific posttranslational modification, sumoylation, have raised several unsolved issues. Functionally, PML bodies may sequester, modify or degrade partner proteins, but in many ways, PML bodies still constitute an enigma.

Nuclear Import and Export of the Thyroid Hormone Receptor.

PubMed

Zhang, Jibo; Roggero, Vincent R; Allison, Lizabeth A

2018-01-01

The thyroid hormone receptors, TRα1 and TRβ1, are members of the nuclear receptor superfamily that forms one of the most abundant classes of transcription factors in multicellular organisms. Although primarily localized to the nucleus, TRα1 and TRβ1 shuttle rapidly between the nucleus and cytoplasm. The fine balance between nuclear import and export of TRs has emerged as a critical control point for modulating thyroid hormone-responsive gene expression. Mutagenesis studies have defined two nuclear localization signal (NLS) motifs that direct nuclear import of TRα1: NLS-1 in the hinge domain and NLS-2 in the N-terminal A/B domain. Three nuclear export signal (NES) motifs reside in the ligand-binding domain. A combined approach of shRNA-mediated knockdown and coimmunoprecipitation assays revealed that nuclear entry of TRα1 is facilitated by importin 7, likely through interactions with NLS-2, and importin β1 and the adapter importin α1 interacting with both NLS-1 and NLS-2. Interestingly, TRβ1 lacks NLS-2 and nuclear import depends solely on the importin α1/β1 heterodimer. Heterokaryon and fluorescence recovery after photobleaching shuttling assays identified multiple exportins that play a role in nuclear export of TRα1, including CRM1 (exportin 1), and exportins 4, 5, and 7. Even single amino acid changes in TRs dramatically alter their intracellular distribution patterns. We conclude that mutations within NLS and NES motifs affect nuclear shuttling activity, and propose that TR mislocalization contributes to the development of some types of cancer and Resistance to Thyroid Hormone syndrome. © 2018 Elsevier Inc. All rights reserved.

Design principles of nuclear receptor signaling: how complex networking improves signal transduction

PubMed Central

Kolodkin, Alexey N; Bruggeman, Frank J; Plant, Nick; Moné, Martijn J; Bakker, Barbara M; Campbell, Moray J; van Leeuwen, Johannes P T M; Carlberg, Carsten; Snoep, Jacky L; Westerhoff, Hans V

2010-01-01

The topology of nuclear receptor (NR) signaling is captured in a systems biological graphical notation. This enables us to identify a number of ‘design' aspects of the topology of these networks that might appear unnecessarily complex or even functionally paradoxical. In realistic kinetic models of increasing complexity, calculations show how these features correspond to potentially important design principles, e.g.: (i) cytosolic ‘nuclear' receptor may shuttle signal molecules to the nucleus, (ii) the active export of NRs may ensure that there is sufficient receptor protein to capture ligand at the cytoplasmic membrane, (iii) a three conveyor belts design dissipating GTP-free energy, greatly aids response, (iv) the active export of importins may prevent sequestration of NRs by importins in the nucleus and (v) the unspecific nature of the nuclear pore may ensure signal-flux robustness. In addition, the models developed are suitable for implementation in specific cases of NR-mediated signaling, to predict individual receptor functions and differential sensitivity toward physiological and pharmacological ligands. PMID:21179018

Protein kinase A is part of a mechanism that regulates nuclear reimport of the nuclear tRNA export receptors Los1p and Msn5p.

PubMed

Pierce, Jacqueline B; van der Merwe, George; Mangroo, Dev

2014-02-01

The two main signal transduction mechanisms that allow eukaryotes to sense and respond to changes in glucose availability in the environment are the cyclic AMP (cAMP)/protein kinase A (PKA) and AMP-activated protein kinase (AMPK)/Snf1 kinase-dependent pathways. Previous studies have shown that the nuclear tRNA export process is inhibited in Saccharomyces cerevisiae deprived of glucose. However, the signal transduction pathway involved and the mechanism by which glucose availability regulates nuclear-cytoplasmic tRNA trafficking are not understood. Here, we show that inhibition of nuclear tRNA export is caused by a block in nuclear reimport of the tRNA export receptors during glucose deprivation. Cytoplasmic accumulation of the tRNA export receptors during glucose deprivation is not caused by activation of Snf1p. Evidence obtained suggests that PKA is part of the mechanism that regulates nuclear reimport of the tRNA export receptors in response to glucose availability. This mechanism does not appear to involve phosphorylation of the nuclear tRNA export receptors by PKA. The block in nuclear reimport of the tRNA export receptors appears to be caused by activation of an unidentified mechanism when PKA is turned off during glucose deprivation. Taken together, the data suggest that PKA facilitates return of the tRNA export receptors to the nucleus by inhibiting an unidentified activity that facilitates cytoplasmic accumulation of the tRNA export receptors when glucose in the environment is limiting. A PKA-independent mechanism was also found to regulate nuclear tRNA export in response to glucose availability. This mechanism, however, does not regulate nuclear reimport of the tRNA export receptors.

Protein Kinase A Is Part of a Mechanism That Regulates Nuclear Reimport of the Nuclear tRNA Export Receptors Los1p and Msn5p

PubMed Central

Pierce, Jacqueline B.; van der Merwe, George

2014-01-01

The two main signal transduction mechanisms that allow eukaryotes to sense and respond to changes in glucose availability in the environment are the cyclic AMP (cAMP)/protein kinase A (PKA) and AMP-activated protein kinase (AMPK)/Snf1 kinase-dependent pathways. Previous studies have shown that the nuclear tRNA export process is inhibited in Saccharomyces cerevisiae deprived of glucose. However, the signal transduction pathway involved and the mechanism by which glucose availability regulates nuclear-cytoplasmic tRNA trafficking are not understood. Here, we show that inhibition of nuclear tRNA export is caused by a block in nuclear reimport of the tRNA export receptors during glucose deprivation. Cytoplasmic accumulation of the tRNA export receptors during glucose deprivation is not caused by activation of Snf1p. Evidence obtained suggests that PKA is part of the mechanism that regulates nuclear reimport of the tRNA export receptors in response to glucose availability. This mechanism does not appear to involve phosphorylation of the nuclear tRNA export receptors by PKA. The block in nuclear reimport of the tRNA export receptors appears to be caused by activation of an unidentified mechanism when PKA is turned off during glucose deprivation. Taken together, the data suggest that PKA facilitates return of the tRNA export receptors to the nucleus by inhibiting an unidentified activity that facilitates cytoplasmic accumulation of the tRNA export receptors when glucose in the environment is limiting. A PKA-independent mechanism was also found to regulate nuclear tRNA export in response to glucose availability. This mechanism, however, does not regulate nuclear reimport of the tRNA export receptors. PMID:24297441

Post-Translational Modification Biology of Glutamate Receptors and Drug Addiction

PubMed Central

Mao, Li-Min; Guo, Ming-Lei; Jin, Dao-Zhong; Fibuch, Eugene E.; Choe, Eun Sang; Wang, John Q.

2011-01-01

Post-translational covalent modifications of glutamate receptors remain a hot topic. Early studies have established that this family of receptors, including almost all ionotropic and metabotropic glutamate receptor subtypes, undergoes active phosphorylation at serine, threonine, or tyrosine residues in their intracellular domains. Recent evidence identifies several glutamate receptor subtypes to be direct substrates for palmitoylation at cysteine residues. Other modifications such as ubiquitination and sumoylation at lysine residues also occur to certain glutamate receptors. These modifications are dynamic and reversible in nature and are regulatable by changing synaptic inputs. The regulated modifications significantly impact the receptor in many ways, including interrelated changes in biochemistry (synthesis, subunit assembling, and protein–protein interactions), subcellular redistribution (trafficking, endocytosis, synaptic delivery, and clustering), and physiology, usually associated with changes in synaptic plasticity. Glutamate receptors are enriched in the striatum and cooperate closely with dopamine to regulate striatal signaling. Emerging evidence shows that modification processes of striatal glutamate receptors are sensitive to addictive drugs, such as psychostimulants (cocaine and amphetamine). Altered modifications are believed to be directly linked to enduring receptor/synaptic plasticity and drug-seeking. This review summarizes several major types of modifications of glutamate receptors and analyzes the role of these modifications in striatal signaling and in the pathogenesis of psychostimulant addiction. PMID:21441996

A Novel Post-translational Modification of Nucleolin, SUMOylation at Lys-294, Mediates Arsenite-induced Cell Death by Regulating gadd45α mRNA Stability*

PubMed Central

Zhang, Dongyun; Liang, Yuguang; Xie, Qipeng; Gao, Guangxun; Wei, Jinlong; Huang, Haishan; Li, Jingxia; Gao, Jimin; Huang, Chuanshu

2015-01-01

Nucleolin is a ubiquitously expressed protein and participates in many important biological processes, such as cell cycle regulation and ribosomal biogenesis. The activity of nucleolin is regulated by intracellular localization and post-translational modifications, including phosphorylation, methylation, and ADP-ribosylation. Small ubiquitin-like modifier (SUMO) is a category of recently verified forms of post-translational modifications and exerts various effects on the target proteins. In the studies reported here, we discovered SUMOylational modification of human nucleolin protein at Lys-294, which facilitated the mRNA binding property of nucleolin by maintaining its nuclear localization. In response to arsenic exposure, nucleolin-SUMO was induced and promoted its binding with gadd45α mRNA, which increased gadd45α mRNA stability and protein expression, subsequently causing GADD45α-mediated cell death. On the other hand, ectopic expression of Mn-SOD attenuated the arsenite-generated superoxide radical level, abrogated nucleolin-SUMO, and in turn inhibited arsenite-induced apoptosis by reducing GADD45α expression. Collectively, our results for the first time demonstrate that nucleolin-SUMO at K294R plays a critical role in its nucleus sequestration and gadd45α mRNA binding activity. This novel biological function of nucleolin is distinct from its conventional role as a proto-oncogene. Therefore, our findings here not only reveal a new modification of nucleolin protein and its novel functional paradigm in mRNA metabolism but also expand our understanding of the dichotomous roles of nucleolin in terms of cancer development, which are dependent on multiple intracellular conditions and consequently the appropriate regulations of its modifications, including SUMOylation. PMID:25561743

Modulation of NRF2 signaling pathway by nuclear receptors: implications for cancer.

PubMed

Namani, Akhileshwar; Li, Yulong; Wang, Xiu Jun; Tang, Xiuwen

2014-09-01

Nuclear factor-erythroid 2 p45-related factor 2 (NRF2, also known as Nfe2l2) plays a critical role in regulating cellular defense against electrophilic and oxidative stress by activating the expression of an array of antioxidant response element-dependent genes. On one hand, NRF2 activators have been used in clinical trials for cancer prevention and the treatment of diseases associated with oxidative stress; on the other hand, constitutive activation of NRF2 in many types of tumors contributes to the survival and growth of cancer cells, as well as resistance to anticancer therapy. In this review, we provide an overview of the NRF2 signaling pathway and discuss its role in carcinogenesis. We also introduce the inhibition of NRF2 by nuclear receptors. Further, we address the biological significance of regulation of the NRF2 signaling pathway by nuclear receptors in health and disease. Finally, we discuss the possible impact of NRF2 inhibition by nuclear receptors on cancer therapy. Copyright © 2014 Elsevier B.V. All rights reserved.

Panning for SNuRMs: using cofactor profiling for the rational discovery of selective nuclear receptor modulators.

PubMed

Kremoser, Claus; Albers, Michael; Burris, Thomas P; Deuschle, Ulrich; Koegl, Manfred

2007-10-01

Drugs that target nuclear receptors are clinically, as well as commercially, successful. Their widespread use, however, is limited by an inherent propensity of nuclear receptors to trigger beneficial, as well as adverse, pharmacological effects upon drug activation. Hence, selective drugs that display reduced adverse effects, such as the selective estrogen receptor modulator (SERM) Raloxifene, have been developed by guidance through classical cell culture assays and animal trials. Full agonist and selective modulator nuclear receptor drugs, in general, differ by their ability to recruit certain cofactors to the receptor protein. Hence, systematic cofactor profiling is advancing into an approach for the rationally guided identification of selective NR modulators (SNuRMs) with improved therapeutic ratio.

A SUMO-acetyl switch in PXR biology.

PubMed

Cui, Wenqi; Sun, Mengxi; Zhang, Shupei; Shen, Xunan; Galeva, Nadezhda; Williams, Todd D; Staudinger, Jeff L

2016-09-01

Post-translational modification (PTM) of nuclear receptor superfamily members regulates various aspects of their biology to include sub-cellular localization, the repertoire of protein-binding partners, as well as their stability and mode of degradation. The nuclear receptor pregnane X receptor (PXR, NR1I2) is a master-regulator of the drug-inducible gene expression in liver and intestine. The PXR-mediated gene activation program is primarily recognized to increase drug metabolism, drug transport, and drug efflux pathways in these tissues. The activation of PXR also has important implications in significant human diseases including inflammatory bowel disease and cancer. Our recent investigations reveal that PXR is modified by multiple PTMs to include phosphorylation, SUMOylation, and ubiquitination. Using both primary cultures of hepatocytes and cell-based assays, we show here that PXR is modified through acetylation on lysine residues. Further, we show that increased acetylation of PXR stimulates its increased SUMO-modification to support active transcriptional suppression. Pharmacologic inhibition of lysine de-acetylation using trichostatin A (TSA) alters the sub-cellular localization of PXR in cultured hepatocytes, and also has a profound impact upon PXR transactivation capacity. Both the acetylation and SUMOylation status of the PXR protein is affected by its ability to associate with the lysine de-acetylating enzyme histone de-acetylase (HDAC)3 in a complex with silencing mediator of retinoic acid and thyroid hormone receptor (SMRT). Taken together, our data support a model in which a SUMO-acetyl 'switch' occurs such that acetylation of PXR likely stimulates SUMO-modification of PXR to promote the active repression of PXR-target gene expression. This article is part of a Special Issue entitled: Xenobiotic nuclear receptors: New Tricks for An Old Dog, edited by Dr. Wen Xie. Copyright © 2016 Elsevier B.V. All rights reserved.

The nuclear orphan receptors COUP-TF and ARP-1 positively regulate the trout estrogen receptor gene through enhancing autoregulation.

PubMed Central

Lazennec, G; Kern, L; Valotaire, Y; Salbert, G

1997-01-01

The rainbow trout estrogen receptor (rtER) is a positively autoregulated gene in liver cells. In a previous report, we showed that upregulation is mediated by an estrogen response element (ERE) located in the proximal promoter of the gene and that a half binding site for nuclear receptors (5'-TGACCT-3') located 15 bp upstream of the ERE is involved in the magnitude of the estrogen response. We now report that the human orphan receptor COUP-TF and a COUP-TF-like protein from trout liver are able to bind to the consensus half-site. When cotransfected with the rtER gene proximal promoter, COUP-TF had no regulatory functions on its own. Interestingly, COUP-TF enhanced rtER transactivation properties in the presence of estradiol in a dose-dependent manner when cotransfected with the rtER gene promoter. Unliganded retinoid receptor heterodimers had the same helper function as COUP-TF in the presence of estradiol but were switched to repressors when the ligand all-trans-retinoic acid was added. Mutation of the consensus half-site only slightly reduced COUP-TF helper function, suggesting that it actually results from a complex mechanism that probably involves both DNA binding of COUP-TF to the promoter and protein-protein interaction with another transcription factor bound to the promoter. Nevertheless, a DNA-binding-defective mutant of COUP-TF was also defective in ER helper function. Competition footprinting analysis suggested that COUP-TF actually establishes contacts with the consensus upstream half-site and the downstream ERE half-site that would form a DR-24-like response element. Interaction of COUP-TF with the DR-24 element was confirmed in footprinting assays by using nuclear extracts from Saccharomyces cerevisiae expressing COUP-TF. Finally, interaction of COUP-TF with mutants of the rtER gene promoter showed that COUP-TF recognizes the ERE when the upstream half-site is mutated. These data show that COUP-TF may activate transcription through interaction with

Arsenic degrades PML or PML-RARalpha through a SUMO-triggered RNF4/ubiquitin-mediated pathway.

PubMed

Lallemand-Breitenbach, Valérie; Jeanne, Marion; Benhenda, Shirine; Nasr, Rihab; Lei, Ming; Peres, Laurent; Zhou, Jun; Zhu, Jun; Raught, Brian; de Thé, Hugues

2008-05-01

In acute promyelocytic leukaemia (APL), arsenic trioxide induces degradation of the fusion protein encoded by the PML-RARA oncogene, differentiation of leukaemic cells and produces clinical remissions. SUMOylation of its PML moiety was previously implicated, but the nature of the degradation pathway involved and the role of PML-RARalpha catabolism in the response to therapy have both remained elusive. Here, we demonstrate that arsenic-induced PML SUMOylation triggers its Lys 48-linked polyubiquitination and proteasome-dependent degradation. When exposed to arsenic, SUMOylated PML recruits RNF4, the human orthologue of the yeast SUMO-dependent E3 ubiquitin-ligase, as well as ubiquitin and proteasomes onto PML nuclear bodies. Arsenic-induced differentiation is impaired in cells transformed by a non-degradable PML-RARalpha SUMOylation mutant or in APL cells transduced with a dominant-negative RNF4, directly implicating PML-RARalpha catabolism in the therapeutic response. We thus identify PML as the first protein degraded by SUMO-dependent polyubiquitination. As PML SUMOylation recruits not only RNF4, ubiquitin and proteasomes, but also many SUMOylated proteins onto PML nuclear bodies, these domains could physically integrate the SUMOylation, ubiquitination and degradation pathways.

An improved ChIP-seq peak detection system for simultaneously identifying post-translational modified transcription factors by combinatorial fusion, using SUMOylation as an example.

PubMed

Cheng, Chia-Yang; Chu, Chia-Han; Hsu, Hung-Wei; Hsu, Fang-Rong; Tang, Chung Yi; Wang, Wen-Ching; Kung, Hsing-Jien; Chang, Pei-Ching

2014-01-01

Post-translational modification (PTM) of transcriptional factors and chromatin remodelling proteins is recognized as a major mechanism by which transcriptional regulation occurs. Chromatin immunoprecipitation (ChIP) in combination with high-throughput sequencing (ChIP-seq) is being applied as a gold standard when studying the genome-wide binding sites of transcription factor (TFs). This has greatly improved our understanding of protein-DNA interactions on a genomic-wide scale. However, current ChIP-seq peak calling tools are not sufficiently sensitive and are unable to simultaneously identify post-translational modified TFs based on ChIP-seq analysis; this is largely due to the wide-spread presence of multiple modified TFs. Using SUMO-1 modification as an example; we describe here an improved approach that allows the simultaneous identification of the particular genomic binding regions of all TFs with SUMO-1 modification. Traditional peak calling methods are inadequate when identifying multiple TF binding sites that involve long genomic regions and therefore we designed a ChIP-seq processing pipeline for the detection of peaks via a combinatorial fusion method. Then, we annotate the peaks with known transcription factor binding sites (TFBS) using the Transfac Matrix Database (v7.0), which predicts potential SUMOylated TFs. Next, the peak calling result was further analyzed based on the promoter proximity, TFBS annotation, a literature review, and was validated by ChIP-real-time quantitative PCR (qPCR) and ChIP-reChIP real-time qPCR. The results show clearly that SUMOylated TFs are able to be pinpointed using our pipeline. A methodology is presented that analyzes SUMO-1 ChIP-seq patterns and predicts related TFs. Our analysis uses three peak calling tools. The fusion of these different tools increases the precision of the peak calling results. TFBS annotation method is able to predict potential SUMOylated TFs. Here, we offer a new approach that enhances Ch

How does oxygen rise drive evolution? Clues from oxygen-dependent biosynthesis of nuclear receptor ligands

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jiang, Ying-Ying; Kong, De-Xin; Qin, Tao

2010-01-08

It is well known that oxygen rise greatly facilitated biological evolution. However, the underlying mechanisms remain elusive. Recently, Raymond and Segre revealed that molecular oxygen allows 1000 more metabolic reactions than can occur in anoxic conditions. From the novel metabolites produced in aerobic metabolism, we serendipitously found that some of the metabolites are signaling molecules that target nuclear receptors. Since nuclear signaling systems are indispensable to superior organisms, we speculated that aerobic metabolism may facilitate biological evolution through promoting the establishment of nuclear signaling systems. This hypothesis is validated by the observation that most (97.5%) nuclear receptor ligands are producedmore » by aerobic metabolism, which is further explained in terms of the chemical criteria (appropriate volume and rather high hydrophobicity) of nuclear receptor ligands that aerobic metabolites are more ready than anaerobic counterparts to satisfy these criteria.« less

Identification of Modulators of the Nuclear Receptor Peroxisome Proliferator-Activated Receptor α (PPARα) in a Mouse Liver Gene Expression Compendium

EPA Science Inventory

The nuclear receptor family member peroxisome proliferator-activated receptor α (PPARα) is activated by therapeutic hypolipidemic drugs and environmentally-relevant chemicals to regulate genes involved in lipid transport and catabolism. Chronic activation of PPARα in rodents inc...

A nuclear-receptor-dependent phosphatidylcholine pathway with antidiabetic effects.

PubMed

Lee, Jae Man; Lee, Yoon Kwang; Mamrosh, Jennifer L; Busby, Scott A; Griffin, Patrick R; Pathak, Manish C; Ortlund, Eric A; Moore, David D

2011-05-25

Nuclear hormone receptors regulate diverse metabolic pathways and the orphan nuclear receptor LRH-1 (also known as NR5A2) regulates bile acid biosynthesis. Structural studies have identified phospholipids as potential LRH-1 ligands, but their functional relevance is unclear. Here we show that an unusual phosphatidylcholine species with two saturated 12 carbon fatty acid acyl side chains (dilauroyl phosphatidylcholine (DLPC)) is an LRH-1 agonist ligand in vitro. DLPC treatment induces bile acid biosynthetic enzymes in mouse liver, increases bile acid levels, and lowers hepatic triglycerides and serum glucose. DLPC treatment also decreases hepatic steatosis and improves glucose homeostasis in two mouse models of insulin resistance. Both the antidiabetic and lipotropic effects are lost in liver-specific Lrh-1 knockouts. These findings identify an LRH-1 dependent phosphatidylcholine signalling pathway that regulates bile acid metabolism and glucose homeostasis.

Evolution of the nuclear receptor superfamily: early diversification from an ancestral orphan receptor.

PubMed

Laudet, V

1997-12-01

From a database containing the published nuclear hormone receptor (NR) sequences I constructed an alignment of the C, D and E domains of these molecules. Using this alignment, I have performed tree reconstruction using both distance matrix and parsimony analysis. The robustness of each branch was estimated using bootstrap resampling methods. The trees constructed by these two methods gave congruent topologies. From these analyses I defined six NR subfamilies: (i) a large one clustering thyroid hormone receptors (TRs), retinoic acid receptors (RARs), peroxisome proliferator-activated receptors (PPARs), vitamin D receptors (VDRs) and ecdysone receptors (EcRs) as well as numerous orphan receptors such as RORs or Rev-erbs; (ii) one containing retinoid X receptors (RXRs) together with COUP, HNF4, tailless, TR2 and TR4 orphan receptors; (iii) one containing steroid receptors; (iv) one containing the NGFIB orphan receptors; (v) one containing FTZ-F1 orphan receptors; and finally (vi) one containing to date only one gene, the GCNF1 orphan receptor. The relationships between the six subfamilies are not known except for subfamilies I and IV which appear to be related. Interestingly, most of the liganded receptors appear to be derived when compared with orphan receptors. This suggests that the ligand-binding ability of NRs has been gained by orphan receptors during the course of evolution to give rise to the presently known receptors. The distribution into six subfamilies correlates with the known abilities of the various NRs to bind to DNA as homo- or heterodimers. For example, receptors heterodimerizing efficiently with RXR belong to the first or the fourth subfamilies. I suggest that the ability to heterodimerize evolved once, just before the separation of subfamilies I and IV and that the first NR was able to bind to DNA as a homodimer. From the study of NR sequences existing in vertebrates, arthropods and nematodes, I define two major steps of NR diversification: one

PML Nuclear Bodies

PubMed Central

Lallemand-Breitenbach, Valérie; de Thé, Hugues

2010-01-01

PML nuclear bodies are matrix-associated domains that recruit an astonishing variety of seemingly unrelated proteins. Since their discovery in the early 1960s, PML bodies have fascinated cell biologists because of their beauty and their tight association with cellular disorders. The identification of PML, a gene involved in an oncogenic chromosomal translocation, as the key organizer of these domains drew instant interest onto them. The multiple levels of PML body regulation by a specific posttranslational modification, sumoylation, have raised several unsolved issues. Functionally, PML bodies may sequester, modify or degrade partner proteins, but in many ways, PML bodies still constitute an enigma. PMID:20452955

Med1 subunit of the mediator complex in nuclear receptor-regulated energy metabolism, liver regeneration, and hepatocarcinogenesis.

PubMed

Jia, Yuzhi; Viswakarma, Navin; Reddy, Janardan K

2014-01-01

Several nuclear receptors regulate diverse metabolic functions that impact on critical biological processes, such as development, differentiation, cellular regeneration, and neoplastic conversion. In the liver, some members of the nuclear receptor family, such as peroxisome proliferator-activated receptors (PPARs), constitutive androstane receptor (CAR), farnesoid X receptor (FXR), liver X receptor (LXR), pregnane X receptor (PXR), glucocorticoid receptor (GR), and others, regulate energy homeostasis, the formation and excretion of bile acids, and detoxification of xenobiotics. Excess energy burning resulting from increases in fatty acid oxidation systems in liver generates reactive oxygen species, and the resulting oxidative damage influences liver regeneration and liver tumor development. These nuclear receptors are important sensors of exogenous activators as well as receptor-specific endogenous ligands. In this regard, gene knockout mouse models revealed that some lipid-metabolizing enzymes generate PPARα-activating ligands, while others such as ACOX1 (fatty acyl-CoA oxidase1) inactivate these endogenous PPARα activators. In the absence of ACOX1, the unmetabolized ACOX1 substrates cause sustained activation of PPARα, and the resulting increase in energy burning leads to hepatocarcinogenesis. Ligand-activated nuclear receptors recruit the multisubunit Mediator complex for RNA polymerase II-dependent gene transcription. Evidence indicates that the Med1 subunit of the Mediator is essential for PPARα, PPARγ, CAR, and GR signaling in liver. Med1 null hepatocytes fail to respond to PPARα activators in that these cells do not show induction of peroxisome proliferation and increases in fatty acid oxidation enzymes. Med1-deficient hepatocytes show no increase in cell proliferation and do not give rise to liver tumors. Identification of nuclear receptor-specific coactivators and Mediator subunits should further our understanding of the complexities of metabolic

Skeletal muscle and nuclear hormone receptors: implications for cardiovascular and metabolic disease.

PubMed

Smith, Aaron G; Muscat, George E O

2005-10-01

Skeletal muscle is a major mass peripheral tissue that accounts for approximately 40% of the total body mass and a major player in energy balance. It accounts for >30% of energy expenditure, is the primary tissue of insulin stimulated glucose uptake, disposal, and storage. Furthermore, it influences metabolism via modulation of circulating and stored lipid (and cholesterol) flux. Lipid catabolism supplies up to 70% of the energy requirements for resting muscle. However, initial aerobic exercise utilizes stored muscle glycogen but as exercise continues, glucose and stored muscle triglycerides become important energy substrates. Endurance exercise increasingly depends on fatty acid oxidation (and lipid mobilization from other tissues). This underscores the importance of lipid and glucose utilization as an energy source in muscle. Consequently skeletal muscle has a significant role in insulin sensitivity, the blood lipid profile, and obesity. Moreover, caloric excess, obesity and physical inactivity lead to skeletal muscle insulin resistance, a risk factor for the development of type II diabetes. In this context skeletal muscle is an important therapeutic target in the battle against cardiovascular disease, the worlds most serious public health threat. Major risk factors for cardiovascular disease include dyslipidemia, hypertension, obesity, sedentary lifestyle, and diabetes. These risk factors are directly influenced by diet, metabolism and physical activity. Metabolism is largely regulated by nuclear hormone receptors which function as hormone regulated transcription factors that bind DNA and mediate the patho-physiological regulation of gene expression. Metabolism and activity, which directly influence cardiovascular disease risk factors, are primarily driven by skeletal muscle. Recently, many nuclear receptors expressed in skeletal muscle have been shown to improve glucose tolerance, insulin resistance, and dyslipidemia. Skeletal muscle and nuclear receptors are

Selective regulation of nuclear orphan receptors 4A by adenosine receptor subtypes in human mast cells

PubMed Central

Zhang, Li; Paine, Catherine

2010-01-01

Nuclear orphan receptors 4A (NR4A) are early responsive genes that belong to the superfamily of hormone receptors and comprise NR4A1, NR4A2 and NR4A3. They have been associated to transcriptional activation of multiple genes involved in inflammation, apoptosis and cell cycle control. Here, we establish a link between NR4As and adenosine, a paradoxical inflammatory molecule that can contribute to persistence of inflammation or mediate inflammatory shutdown. Transcriptomics screening of the human mast cell-line HMC-1 revealed a sharp induction of transcriptionally active NR4A2 and NR4A3 by the adenosine analogue NECA. The concomitant treatment of NECA and the adenosine receptor A2A (A2AAR) selective antagonist SCH-58261 exaggerated this effect, suggesting that upregulation of these factors in mast cells is mediated by other AR subtypes (A2B and A3) and that A2AAR activation counteracts NR4A2 and NR4A3 induction. In agreement with this, A2AAR-silencing amplified NR4A induction by NECA. Interestingly, a similar A2AAR modulatory effect was observed on ERK1/2 phosphorylation because A2AAR blockage exacerbated NECA-mediated phosphorylation of ERK1/2. In addition, PKC or MEK1/2 inhibition prevented ERK1/2 phosphorylation and antagonized AR-mediated induction of NR4A2 and NR4A3, suggesting the involvement of these kinases in AR to NR4A signaling. Finally, we observed that selective A2AAR activation with CGS-21680 blocked PMA-induced ERK1/2 phosphorylation and modulated the overexpression of functional nuclear orphan receptors 4A. Taken together, these results establish a novel PKC/ERK/nuclear orphan receptors 4A axis for adenosinergic signaling in mast cells, which can be modulated by A2AAR activation, not only in the context of adenosine but of other mast cell activating stimuli as well. PMID:21234122

Regulation of hepatic energy metabolism by the nuclear receptor PXR.

PubMed

Hakkola, Jukka; Rysä, Jaana; Hukkanen, Janne

2016-09-01

The pregnane X receptor (PXR) is a nuclear receptor that is traditionally thought to be specialized for sensing xenobiotic exposure. In concurrence with this feature PXR was originally identified to regulate drug-metabolizing enzymes and transporters. During the last ten years it has become clear that PXR harbors broader functions. Evidence obtained both in experimental animals and humans indicate that ligand-activated PXR regulates hepatic glucose and lipid metabolism and affects whole body metabolic homeostasis. Currently, the consequences of PXR activation on overall metabolic health are not yet fully understood and varying results on the effect of PXR activation or knockout on metabolic disorders and weight gain have been published in mouse models. Rifampicin and St. John's wort, the prototypical human PXR agonists, impair glucose tolerance in healthy volunteers. Chronic exposure to PXR agonists could potentially represent a risk factor for diabetes and metabolic syndrome. This article is part of a Special Issue entitled: Xenobiotic nuclear receptors: New Tricks for An Old Dog, edited by Dr. Wen Xie. Copyright © 2016 Elsevier B.V. All rights reserved.

Involvement of the orphan nuclear estrogen receptor-related receptor α in osteoclast adhesion and transmigration

PubMed Central

Bonnelye, Edith; Saltel, Frédéric; Chabadel, Anne; Zirngibl, Ralph A; Aubin, Jane E; Jurdic, Pierre

2010-01-01

The orphan nuclear receptor, estrogen receptor-related receptor α (ERRα) is expressed in osteoblasts and osteoclasts (OCs) and has been proposed to be a modulator of estrogen signaling. To determine the role of ERRα in OC biology, we knocked down ERRα activity by transient transfection of an siRNA directed against ERRα in the RAW264.7 monocyte–macrophage cell line that differentiates into OCs in the presence of receptor activator of nuclear factor κB-ligands and macrophage colony-stimulating factor. In parallel, stable RAW cell lines expressing a dominant-negative form of ERRα and green fluorescent protein (RAW-GFP-ERRαΔAF2) were used. Expression of OC markers was assessed by real-time PCR, and adhesion and transmigration tests were performed. Actin cytoskeletal organization was visualized using confocal microscopy. We found that RAW264.7 cells expressing siRNA directed against ERRα and RAW-GFP-ERRαΔAF2 OCs displayed abnormal spreading, and decreased osteopontin and β3 integrin subunit expression compared with the corresponding control cells. Decreased adhesion and the absence of podosome belts concomitant with abnormal localization of c-src were also observed in RAW-GFP-ERRαΔAF2-derived OCs. In addition, RAW-GFP-ERRαΔAF2-derived OCs failed to transmigrate through osteoblast cell layers. Our data show that the impairment of ERRα function does not alter OC precursor proliferation and differentiation but does alter the adhesion/spreading and migration capacities of mature OCs. PMID:20841427

Nuclear receptor TLX inhibits TGF-β signaling in glioblastoma

DOE Office of Scientific and Technical Information (OSTI.GOV)

Johansson, Erik; Zhai, Qiwei; Zeng, Zhao-jun

TLX (also called NR2E1) is an orphan nuclear receptor that maintains stemness of neuronal stem cells. TLX is highly expressed in the most malignant form of glioma, glioblastoma multiforme (GBM), and is important for the proliferation and maintenance of the stem/progenitor cells of the tumor. Transforming Growth Factor-β (TGF-β) is a cytokine regulating many different cellular processes such as differentiation, migration, adhesion, cell death and proliferation. TGF-β has an important function in cancer where it can work as either a tumor suppressor or oncogene, depending on the cancer type and stage of tumor development. Since glioblastoma often have dysfunctional TGF-βmore » signaling we wanted to find out if there is any interaction between TLX and TGF-β in glioblastoma cells. We demonstrate that knockdown of TLX enhances the canonical TGF-β signaling response in glioblastoma cell lines. TLX physically interacts with and stabilizes Smurf1, which can ubiquitinate and target TGF-β receptor II for degradation, whereas knockdown of TLX leads to stabilization of TGF-β receptor II, increased nuclear translocation of Smad2/3 and enhanced expression of TGF-β target genes. The interaction between TLX and TGF-β may play an important role in the regulation of proliferation and tumor-initiating properties of glioblastoma cells. - Highlights: • TLX knockdown enhances TGF-β dependent Smad signaling in glioblastoma cells • TLX knockdown increases the protein level of TGF-β receptor II. • TLX stabilizes and retains Smurf1 in the cytoplasm. • TLX enhances Smurf1-dependent ubiquitination and degradation of TGF-β receptor II.« less

Nuclear hormone receptors in parasitic helminths

PubMed Central

Wu, Wenjie; LoVerde, Philip T

2010-01-01

Nuclear receptors (NRs) belong to a large protein superfamily that are important transcriptional modulators in metazoans. Parasitic helminths include parasitic worms from the Lophotrochozoa (Platyhelminths) and Ecdysozoa (Nematoda). NRs in parasitic helminths diverged into two different evolutionary lineages. NRs in parasitic Platyhelminths have orthologues in Deuterostomes, in arthropods or both with a feature of extensive gene loss and gene duplication within different gene groups. NRs in parasitic Nematoda follow the nematode evolutionary lineage with a feature of multiple duplication of SupNRs and gene loss. PMID:20600585

A protein that interacts with members of the nuclear hormone receptor family: identification and cDNA cloning.

PubMed Central

Zeiner, M; Gehring, U

1995-01-01

In search of proteins which interact with activated steroid hormone receptors, we screened a human liver lambda gt11 expression library with the glucocorticoid receptor. We identified and cloned a cDNA sequence of 1322 bp that encodes a protein of 274 aa. This protein consists predominantly of hydrophilic amino acids and contains a putative bipartite nuclear localization signal. The in vitro translated receptor-associating protein runs in SDS/polyacrylamide gels with an apparent molecular mass of 46 kDa. By use of the bacterially expressed fusion protein with glutathione S-transferase we have found that interaction is not limited to the glucocorticoid receptor but included other nuclear receptors--most notably, the estrogen and thyroid receptors. Binding also occurs with the glucocorticoid receptor complexed with the antiglucocorticoid RU 38486, with the estrogen receptor complexed with the antiestrogen 4-hydroxytamoxifen or ICI 164,384, and even with receptors not complexed with ligand. Association with steroid hormone receptors depends on prior receptor activation--i.e., release from heat shock proteins. The sequence identified here appears to be a general partner protein for nuclear hormone receptors, with the gene being expressed in a variety of mammalian tissues. Images Fig. 2 Fig. 3 Fig. 4 PMID:8524784

Bmal1 is a direct transcriptional target of the orphan nuclear receptor, NR2F1

USDA-ARS?s Scientific Manuscript database

Orphan nuclear receptor NR2F1 (also known as COUP-TFI, Chicken Ovalbumin Upstream Promoter Transcription Factor I) is a highly conserved member of the nuclear receptor superfamily. NR2F1 plays a critical role during embryonic development, particularly in the central and peripheral nervous systems a...

Nutrient-sensing nuclear receptors PPARα and FXR control liver energy balance.

PubMed

Preidis, Geoffrey A; Kim, Kang Ho; Moore, David D

2017-04-03

The nuclear receptors PPARα (encoded by NR1C1) and farnesoid X receptor (FXR, encoded by NR1H4) are activated in the liver in the fasted and fed state, respectively. PPARα activation induces fatty acid oxidation, while FXR controls bile acid homeostasis, but both nuclear receptors also regulate numerous other metabolic pathways relevant to liver energy balance. Here we review evidence that they function coordinately to control key nutrient pathways, including fatty acid oxidation and gluconeogenesis in the fasted state and lipogenesis and glycolysis in the fed state. We have also recently reported that these receptors have mutually antagonistic impacts on autophagy, which is induced by PPARα but suppressed by FXR. Secretion of multiple blood proteins is a major drain on liver energy and nutrient resources, and we present preliminary evidence that the liver secretome may be directly suppressed by PPARα, but induced by FXR. Finally, previous studies demonstrated a striking deficiency in bile acid levels in malnourished mice that is consistent with results in malnourished children. We present evidence that hepatic targets of PPARα and FXR are dysregulated in chronic undernutrition. We conclude that PPARα and FXR function coordinately to integrate liver energy balance.

Combined therapeutic potential of nuclear receptors with receptor tyrosine kinase inhibitors in lung cancer

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wairagu, Peninah M.; Institute of Lifestyle Medicine, Wonju College of Medicine, Yonsei University, Wonju, Gangwon-do 220-701; Nuclear Receptor Research Consortium, Wonju College of Medicine, Yonsei University, Wonju, Gangwon-do 220-701

2014-05-09

Highlights: • The 48 NR genes and 48 biological anti-cancer targets are profiled in paired-cells. • Growth inhibition by NR ligands or TKIs is target receptor level-dependent. • T0901317 with gefitinib/PHA665752 shows additive growth inhibition in lung cells. - Abstract: Cancer heterogeneity is a big hurdle in achieving complete cancer treatment, which has led to the emergence of combinational therapy. In this study, we investigated the potential use of nuclear receptor (NR) ligands for combinational therapy with other anti-cancer drugs. We first profiled all 48 NRs and 48 biological anti-cancer targets in four pairs of lung cell lines, where eachmore » pair was obtained from the same patient. Two sets of cell lines were normal and the corresponding tumor cell lines while the other two sets consisted of primary versus metastatic tumor cell lines. Analysis of the expression profile revealed 11 NRs and 15 cancer targets from the two pairs of normal versus tumor cell lines, and 9 NRs and 9 cancer targets from the primary versus metastatic tumor cell lines had distinct expression patterns in each category. Finally, the evaluation of nuclear receptor ligand T0901317 for liver X receptor (LXR) demonstrated its combined therapeutic potential with tyrosine kinase inhibitors. The combined treatment of cMET inhibitor PHA665752 or EGFR inhibitor gefitinib with T0901317 showed additive growth inhibition in both H2073 and H1993 cells. Mechanistically, the combined treatment suppressed cell cycle progression by inhibiting cyclinD1 and cyclinB expression. Taken together, this study provides insight into the potential use of NR ligands in combined therapeutics with other biological anti-cancer drugs.« less

Non-canonical modulators of nuclear receptors.

PubMed

Tice, Colin M; Zheng, Ya-Jun

2016-09-01

Like G protein-coupled receptors (GPCRs) and protein kinases, nuclear receptors (NRs) are a rich source of pharmaceutical targets. Over 80 NR-targeting drugs have been approved for 18 NRs. The focus of drug discovery in NRs has hitherto been on identifying ligands that bind to the canonical ligand binding pockets of the C-terminal ligand binding domains (LBDs). Due to the development of drug resistance and selectivity concerns, there has been considerable interest in exploring other, non-canonical ligand binding sites. Unfortunately, the potencies of compounds binding at other sites have generally not been sufficient for clinical development. However, the situation has changed dramatically over the last 3years, as compounds with sufficient potency have been reported for several NR targets. Here we review recent developments in this area from a medicinal chemistry point of view in the hope of stimulating further interest in this area of research. Copyright © 2016 Elsevier Ltd. All rights reserved.

Transcriptional regulation of human Paraoxonase 1 by nuclear receptors.

PubMed

Ponce-Ruiz, N; Murillo-González, F E; Rojas-García, A E; Mackness, Mike; Bernal-Hernández, Y Y; Barrón-Vivanco, B S; González-Arias, C A; Medina-Díaz, I M

2017-04-25

Paraoxonase 1 (PON1) is a calcium-dependent lactonase synthesized primarily in the liver and secreted into the plasma, where it is associates with high density lipoproteins (HDL). PON1 acts as antioxidant preventing low-density lipoprotein (LDL) oxidation, a process considered critical in the initiation and progression of atherosclerosis. Additionally, PON1 hydrolyzes and detoxifies some toxic metabolites of organophosphorus compounds (OPs). Thus, PON1 activity and expression levels are important for determining susceptibility to OPs intoxication and risk of developing diseases related to inflammation and oxidative stress. Increasing evidence has demonstrated the modulation of PON1 expression by many factors is due to interaction with nuclear receptors (NRs). Here, we briefly review the studies in this area and discuss the role of nuclear receptors in the regulation of PON1 expression, as well as how understanding these mechanisms may allow us to manipulate PON1 levels to improve drug efficacy and treat disease. Copyright © 2017 Elsevier B.V. All rights reserved.

Huntingtin interacting protein 1 modulates the transcriptional activity of nuclear hormone receptors.

PubMed

Mills, Ian G; Gaughan, Luke; Robson, Craig; Ross, Theodora; McCracken, Stuart; Kelly, John; Neal, David E

2005-07-18

Internalization of activated receptors regulates signaling, and endocytic adaptor proteins are well-characterized in clathrin-mediated uptake. One of these adaptor proteins, huntingtin interacting protein 1 (HIP1), induces cellular transformation and is overexpressed in some prostate cancers. We have discovered that HIP1 associates with the androgen receptor through a central coiled coil domain and is recruited to DNA response elements upon androgen stimulation. HIP1 is a novel androgen receptor regulator, significantly repressing transcription when knocked down using a silencing RNA approach and activating transcription when overexpressed. We have also identified a functional nuclear localization signal at the COOH terminus of HIP1, which contributes to the nuclear translocation of the protein. In conclusion, we have discovered that HIP1 is a nucleocytoplasmic protein capable of associating with membranes and DNA response elements and regulating transcription.

Huntingtin interacting protein 1 modulates the transcriptional activity of nuclear hormone receptors

PubMed Central

Mills, Ian G.; Gaughan, Luke; Robson, Craig; Ross, Theodora; McCracken, Stuart; Kelly, John; Neal, David E.

2005-01-01

Internalization of activated receptors regulates signaling, and endocytic adaptor proteins are well-characterized in clathrin-mediated uptake. One of these adaptor proteins, huntingtin interacting protein 1 (HIP1), induces cellular transformation and is overexpressed in some prostate cancers. We have discovered that HIP1 associates with the androgen receptor through a central coiled coil domain and is recruited to DNA response elements upon androgen stimulation. HIP1 is a novel androgen receptor regulator, significantly repressing transcription when knocked down using a silencing RNA approach and activating transcription when overexpressed. We have also identified a functional nuclear localization signal at the COOH terminus of HIP1, which contributes to the nuclear translocation of the protein. In conclusion, we have discovered that HIP1 is a nucleocytoplasmic protein capable of associating with membranes and DNA response elements and regulating transcription. PMID:16027218

0610009K11Rik, a testis-specific and germ cell nuclear receptor-interacting protein

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zhang Heng; Denhard, Leslie A.; Zhou Huaxin

Using an in silico approach, a putative nuclear receptor-interacting protein 0610009K11Rik was identified in mouse testis. We named this gene testis-specific nuclear receptor-interacting protein-1 (Tnrip-1). Tnrip-1 was predominantly expressed in the testis of adult mouse tissues. Expression of Tnrip-1 in the testis was regulated during postnatal development, with robust expression in 14-day-old or older testes. In situ hybridization analyses showed that Tnrip-1 is highly expressed in pachytene spermatocytes and spermatids. Consistent with its mRNA expression, Tnrip-1 protein was detected in adult mouse testes. Immunohistochemical studies showed that Tnrip-1 is a nuclear protein and mainly expressed in pachytene spermatocytes and roundmore » spermatids. Moreover, co-immunoprecipitation analyses showed that endogenous Tnrip-1 protein can interact with germ cell nuclear receptor (GCNF) in adult mouse testes. Our results suggest that Tnrip-1 is a testis-specific and GCNF-interacting protein which may be involved in the modulation of GCNF-mediated gene transcription in spermatogenic cells within the testis.« less

Phosphorylation of a conserved serine in the deoxyribonucleic acid binding domain of nuclear receptors alters intracellular localization.

PubMed

Sun, Kai; Montana, Vedrana; Chellappa, Karthikeyani; Brelivet, Yann; Moras, Dino; Maeda, Yutaka; Parpura, Vladimir; Paschal, Bryce M; Sladek, Frances M

2007-06-01

Nuclear receptors (NRs) are a superfamily of transcription factors whose genomic functions are known to be activated by lipophilic ligands, but little is known about how to deactivate them or how to turn on their nongenomic functions. One obvious mechanism is to alter the nuclear localization of the receptors. Here, we show that protein kinase C (PKC) phosphorylates a highly conserved serine (Ser) between the two zinc fingers of the DNA binding domain of orphan receptor hepatocyte nuclear factor 4alpha (HNF4alpha). This Ser (S78) is adjacent to several positively charged residues (Arg or Lys), which we show here are involved in nuclear localization of HNF4alpha and are conserved in nearly all other NRs, along with the Ser/threonine (Thr). A phosphomimetic mutant of HNF4alpha (S78D) reduced DNA binding, transactivation ability, and protein stability. It also impaired nuclear localization, an effect that was greatly enhanced in the MODY1 mutant Q268X. Treatment of the hepatocellular carcinoma cell line HepG2 with PKC activator phorbol 12-myristate 13-acetate also resulted in increased cytoplasmic localization of HNF4alpha as well as decreased endogenous HNF4alpha protein levels in a proteasome-dependent fashion. We also show that PKC phosphorylates the DNA binding domain of other NRs (retinoic acid receptor alpha, retinoid X receptor alpha, and thyroid hormone receptor beta) and that phosphomimetic mutants of the same Ser/Thr result in cytoplasmic localization of retinoid X receptor alpha and peroxisome proliferator-activated receptor alpha. Thus, phosphorylation of this conserved Ser between the two zinc fingers may be a common mechanism for regulating the function of NRs.

Rapid, portable detection of endocrine disrupting chemicals through ligand-nuclear hormone receptor interactions.

PubMed

Hunt, J Porter; Schinn, Song-Min; Jones, Matthew D; Bundy, Bradley C

2017-12-04

Endocrine disrupting chemicals (EDC) are structurally diverse compounds that can interact with nuclear hormone receptors, posing significant risk to human and ecological health. Unfortunately, many conventional biosensors have been too structure-specific, labor-intensive or laboratory-oriented to detect broad ranges of EDC effectively. Recently, several technological advances are providing more rapid, portable, and affordable detection of endocrine-disrupting activity through ligand-nuclear hormone receptor interactions. Here, we overview these recent advances applied to EDC biosensors - including cell lyophilization, cell immobilization, cell-free systems, smartphone-based signal detection, and improved competitive binding assays.

SUMO-modified insulin-like growth factor 1 receptor (IGF-1R) increases cell cycle progression and cell proliferation.

PubMed

Lin, Yingbo; Liu, Hongyu; Waraky, Ahmed; Haglund, Felix; Agarwal, Prasoon; Jernberg-Wiklund, Helena; Warsito, Dudi; Larsson, Olle

2017-10-01

Increasing number of studies have shown nuclear localization of the insulin-like growth factor 1 receptor (nIGF-1R) in tumor cells and its links to adverse clinical outcome in various cancers. Any obvious cell physiological roles of nIGF-1R have, however, still not been disclosed. Previously, we reported that IGF-1R translocates to cell nucleus and modulates gene expression by binding to enhancers, provided that the receptor is SUMOylated. In this study, we constructed stable transfectants of wild type IGF1R (WT) and triple-SUMO-site-mutated IGF1R (TSM) using igf1r knockout mouse fibroblasts (R-). Cell clones (R-WT and R-TSM) expressing equal amounts of IGF-1R were selected for experiments. Phosphorylation of IGF-1R, Akt, and Erk upon IGF-1 stimulation was equal in R-WT and R-TSM. WT was confirmed to enter nuclei. TSM did also undergo nuclear translocation, although to a lesser extent. This may be explained by that TSM heterodimerizes with insulin receptor, which is known to translocate to cell nuclei. R-WT proliferated substantially faster than R-TSM, which did not differ significantly from the empty vector control. Upon IGF-1 stimulation G1-S-phase progression of R-WT increased from 12 to 38%, compared to 13 to 20% of R-TSM. The G1-S progression of R-WT correlated with increased expression of cyclin D1, A, and CDK2, as well as downregulation of p27. This suggests that SUMO-IGF-1R affects upstream mechanisms that control and coordinate expression of cell cycle regulators. Further studies to identify such SUMO-IGF-1R dependent mechanisms seem important. © 2017 The Authors. Journal of Cellular Physiology Published by Wiley Periodicals Inc.

Discrimination between NL1- and NL2-Mediated Nuclear Localization of the Glucocorticoid Receptor

PubMed Central

Savory, Joanne G. A.; Hsu, Brian; Laquian, Ian R.; Giffin, Ward; Reich, Terry; Haché, Robert J. G.; Lefebvre, Yvonne A.

1999-01-01

Glucocorticoid receptor (GR) cycles between a free liganded form that is localized to the nucleus and a heat shock protein (hsp)-immunophilin-complexed, unliganded form that is usually localized to the cytoplasm but that can also be nuclear. In addition, rapid nucleocytoplasmic exchange or shuttling of the receptor underlies its localization. Nuclear import of liganded GR is mediated through a well-characterized sequence, NL1, adjacent to the receptor DNA binding domain and a second, uncharacterized motif, NL2, that overlaps with the ligand binding domain. In this study we report that rapid nuclear import (half-life [t1/2] of 4 to 6 min) of agonist- and antagonist-treated GR and the localization of unliganded, hsp-associated GRs to the nucleus in G0 are mediated through NL1 and correlate with the binding of GR to pendulin/importin α. By contrast, NL2-mediated nuclear transfer of GR occurred more slowly (t1/2 = 45 min to 1 h), was agonist specific, and appeared to be independent of binding to importin α. Together, these results suggest that NL2 mediates the nuclear import of GR through an alternative nuclear import pathway. Nuclear export of GR was inhibited by leptomycin B, suggesting that the transfer of GR to the cytoplasm is mediated through the CRM1-dependent pathway. Inhibition of GR nuclear export by leptomycin B enhanced the nuclear localization of both unliganded, wild-type GR and hormone-treated NL1− GR. These results highlight that the subcellular localization of both liganded and unliganded GRs is determined, at least in part, by a flexible equilibrium between the rates of nuclear import and export. PMID:9891038

Nuclear receptor coactivators: regulators of steroid action in brain and behaviour.

PubMed

Tetel, M J; Acharya, K D

2013-11-01

Steroid hormones act in specific regions of the brain to alter behaviour and physiology. Although it has been well established that the bioavailability of the steroid and the expression of its receptor is critical for understanding steroid action in the brain, the importance of nuclear receptor coactivators in the brain is becoming more apparent. The present review focuses on the function of the p160 family of coactivators, which includes steroid receptor coactivator-1 (SRC-1), SRC-2 and SRC-3, in steroid receptor action in the brain. The expression, regulation and function of these coactivators in steroid-dependent gene expression in both brain and behaviour are discussed. © 2013 British Society for Neuroendocrinology.

A muscle-specific knockout implicates nuclear receptor coactivator MED1 in the regulation of glucose and energy metabolism.

PubMed

Chen, Wei; Zhang, Xiaoting; Birsoy, Kivanc; Roeder, Robert G

2010-06-01

As conventional transcriptional factors that are activated in diverse signaling pathways, nuclear receptors play important roles in many physiological processes that include energy homeostasis. The MED1 subunit of the Mediator coactivator complex plays a broad role in nuclear receptor-mediated transcription by anchoring the Mediator complex to diverse promoter-bound nuclear receptors. Given the significant role of skeletal muscle, in part through the action of nuclear receptors, in glucose and fatty acid metabolism, we generated skeletal muscle-specific Med1 knockout mice. Importantly, these mice show enhanced insulin sensitivity and improved glucose tolerance as well as resistance to high-fat diet-induced obesity. Furthermore, the white muscle of these mice exhibits increased mitochondrial density and expression of genes specific to type I and type IIA fibers, indicating a fast-to-slow fiber switch, as well as markedly increased expression of the brown adipose tissue-specific UCP-1 and Cidea genes that are involved in respiratory uncoupling. These dramatic results implicate MED1 as a powerful suppressor in skeletal muscle of genetic programs implicated in energy expenditure and raise the significant possibility of therapeutical approaches for metabolic syndromes and muscle diseases through modulation of MED1-nuclear receptor interactions.

Nuclear receptor TLX inhibits TGF-β signaling in glioblastoma.

PubMed

Johansson, Erik; Zhai, Qiwei; Zeng, Zhao-Jun; Yoshida, Takeshi; Funa, Keiko

2016-05-01

TLX (also called NR2E1) is an orphan nuclear receptor that maintains stemness of neuronal stem cells. TLX is highly expressed in the most malignant form of glioma, glioblastoma multiforme (GBM), and is important for the proliferation and maintenance of the stem/progenitor cells of the tumor. Transforming Growth Factor-β (TGF-β) is a cytokine regulating many different cellular processes such as differentiation, migration, adhesion, cell death and proliferation. TGF-β has an important function in cancer where it can work as either a tumor suppressor or oncogene, depending on the cancer type and stage of tumor development. Since glioblastoma often have dysfunctional TGF-β signaling we wanted to find out if there is any interaction between TLX and TGF-β in glioblastoma cells. We demonstrate that knockdown of TLX enhances the canonical TGF-β signaling response in glioblastoma cell lines. TLX physically interacts with and stabilizes Smurf1, which can ubiquitinate and target TGF-β receptor II for degradation, whereas knockdown of TLX leads to stabilization of TGF-β receptor II, increased nuclear translocation of Smad2/3 and enhanced expression of TGF-β target genes. The interaction between TLX and TGF-β may play an important role in the regulation of proliferation and tumor-initiating properties of glioblastoma cells. Copyright © 2016. Published by Elsevier Inc.

PML nuclear bodies: regulation, function and therapeutic perspectives.

PubMed

Sahin, Umut; Lallemand-Breitenbach, Valérie; de Thé, Hugues

2014-11-01

PML nuclear bodies (NBs) were first described by electron microscopy and rediscovered through their treatment-reversible disruption in a rare leukaemia. They recruit multiple partner proteins and now emerge as interferon- and oxidative stress-responsive sumoylation factories. NBs mediate interferon-induced viral restriction, enhance proteolysis, finely tune metabolism and enforce stress-induced senescence. Apart from being markers of cellular stress, PML NBs could be harnessed pharmacologically in a number of conditions, including cancer, viral infection or neurodegenerative diseases. Copyright © 2014 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sekhri, Palak; Tao, Tao; Kaplan, Feige

As the sole E2 enzyme for SUMOylation, Ubc9 is predominantly nuclear. However, the underlying mechanisms of Ubc9 nuclear localization are still not well understood. Here we show that RNAi-depletion of Imp13, an importin known to mediate Ubc9 nuclear import, reduces both Ubc9 nuclear accumulation and global SUMOylation. Furthermore, Ubc9-R13A or Ubc9-H20D mutation previously shown to interrupt the interaction of Ubc9 with nucleus-enriched SUMOs reduces the nuclear enrichment of Ubc9, suggesting that the interaction of Ubc9 with the nuclear SUMOs may enhance Ubc9 nuclear retention. Moreover, Ubc9-R17E mutation, which is known to disrupt the interaction of Ubc9 with both SUMOs andmore » Imp13, causes a greater decrease in Ubc9 nuclear accumulation than Ubc9-R13A or Ubc9-H20D mutation. Lastly, Ubc9-K74A/S89D mutations that perturb the interaction of Ubc9 with nucleus-enriched SUMOylation-consensus motifs has no effect on Ubc9 nuclear localization. Altogether, our results have elucidated that the amino acid residues within the N-terminal region of Ubc9 play a pivotal role in regulation of Ubc9 nuclear localization. - Highlights: • Imp13-mediated nuclear import of Ubc9 is critical for global SUMOylation. • Ubc9 mutations disrupting Ubc9-SUMO interaction decrease Ubc9 nuclear accumulation. • N-terminal amino acid residues of Ubc9 are critical for Ubc9 nuclear enrichment.« less

Dose-dependent difference of nuclear receptors involved in murine liver hypertrophy by piperonyl butoxide.

PubMed

Sakamoto, Yohei; Yoshida, Midori; Tamura, Kei; Takahashi, Miwa; Kodama, Yukio; Inoue, Kaoru

2015-12-01

Nuclear receptors play important roles in chemically induced liver hypertrophy in rodents. To clarify the involvement of constitutive androstane receptor (CAR) and other nuclear receptors in mouse liver hypertrophy induced by different doses of piperonyl butoxide (PBO), wild-type and CAR-knockout mice were administered PBO (200, 1,000, or 5,000 ppm) in the basal diet for 1 week. Increased liver weight and diffuse hepatocellular hypertrophy were observed at 5,000 ppm for both genotypes, accompanied by increased Cyp3a11 mRNA and CYP3A protein expression, suggesting that CAR-independent pathway, possibly pregnane X receptor (PXR), plays a major role in the induction of hypertrophy. Moreover, wild-type mice at 5,000 ppm showed enhanced hepatocellular hypertrophy and strong positive staining for CYP2B in the centrilobular area, suggesting the localized contribution of CAR. At 1,000 ppm, only wild-type mice showed liver weight increase and centrilobular hepatocellular hypertrophy concurrent with elevated Cyp2b10 mRNA expression and strong CYP2B staining, indicating that CAR was essential at 1,000 ppm. We concluded that high-dose PBO induced hypertrophy via CAR and another pathway, while lower dose of PBO induced a pathway mediated predominantly by CAR. The dose-responsiveness on liver hypertrophy is important for understanding the involvement of nuclear receptors.

Featured Article: Nuclear export of opioid growth factor receptor is CRM1 dependent.

PubMed

Kren, Nancy P; Zagon, Ian S; McLaughlin, Patricia J

2016-02-01

Opioid growth factor receptor (OGFr) facilitates growth inhibition in the presence of its specific ligand opioid growth factor (OGF), chemically termed [Met(5)]-enkephalin. The function of the OGF-OGFr axis requires the receptor to translocate to the nucleus. However, the mechanism of nuclear export of OGFr is unknown. In this study, endogenous OGFr, as well as exogenously expressed OGFr-EGFP, demonstrated significant nuclear accumulation in response to leptomycin B (LMB), an inhibitor of CRM1-dependent nuclear export, suggesting that OGFr is exported in a CRM1-dependent manner. One consensus sequence for a nuclear export signal (NES) was identified. Mutation of the associated leucines, L217 L220 L223 and L225, to alanine resulted in decreased nuclear accumulation. NES-EGFP responded to LMB, indicating that this sequence is capable of functioning as an export signal in isolation. To determine why the sequence functions differently in isolation than as a full length protein, the localization of subNES was evaluated in the presence and absence of MG132, a potent inhibitor of proteosomal degradation. MG132 had no effect of subNES localization. The role of tandem repeats located at the C-terminus of OGFr was examined for their role in nuclear trafficking. Six of seven tandem repeats were removed to form deltaTR. DeltaTR localized exclusively to the nucleus indicating that the tandem repeats may contribute to the localization of the receptor. Similar to the loss of cellular proliferation activity (i.e. inhibition) recorded with subNES, deltaTR also demonstrated a significant loss of inhibitory activity indicating that the repeats may be integral to receptor function. These experiments reveal that OGFr contains one functional NES, L217 L220 L223 and L225 and can be exported from the nucleus in a CRM1-dependent manner. © 2015 by the Society for Experimental Biology and Medicine.

Analysis of the Heat Shock Response in Mouse Liver Reveals Transcriptional Dependence on the Nuclear Receptor Peroxisome Proliferator-Activated Receptor alpha (PPARα)

EPA Science Inventory

BACKGROUND: The nuclear receptor peroxisome proliferator-activated receptor alpha (PPARalpha) regulates responses to chemical or physical stress in part by altering expression of genes involved in proteome maintenance. Many of these genes are also transcriptionally regulated by h...

Mode of Action and Human Relevance Analysis for Nuclear Receptor-Mediated Liver Toxicity: A Case Study with Phenobarbital as a Model Constitutive Androstane Receptor (CAR) Activator

EPA Science Inventory

The constitutive androstane receptor (CAR) and pregnane X receptor (PXR) are key nuclear receptors involved in the regulation of cellular responses. to exposure to many xenobiotics and various physiological processes. Phenobarbital (PB) is a non­ genotoxic i...

Role of molecular chaperones and TPR-domain proteins in the cytoplasmic transport of steroid receptors and their passage through the nuclear pore.

PubMed

Galigniana, Mario D; Echeverría, Pablo C; Erlejman, Alejandra G; Piwien-Pilipuk, Graciela

2010-01-01

In the absence of hormone, corticosteroid receptors such as GR (glucocorticoid receptor) and (mineralocorticoid receptor) are primarily located in the cytoplasm. Upon steroid-binding, they rapidly accumulate in the nucleus. Regardless of their primary location, these receptors and many other nuclear factors undergo a constant and dynamic nucleocytoplasmic shuttling. All members of the steroid receptor family are known to form large oligomeric structures with the heat-shock proteins of 90-kDa (hsp90) and 70-kDa (hsp70), the small acidic protein p23, and a tetratricopeptide repeat (TPR) -domain protein such as FK506-binding proteins (FKBPs), cyclophilins (CyPs) or the serine/threonine protein phosphatase 5 (PP5). It has always been stated that the dissociation of the chaperone heterocomplex (a process normally referred to as receptor "transformation") is the first step that permits the nuclear import of steroid receptors. However the experimental evidence is consistent with a model where the chaperone machinery is required for the retrotransport of the receptor through the cytoplasm and also facilitates the passage through the nuclear pore. Recent evidence indicates that the hsp90-based chaperone system also interacts with structures of the nuclear pore such as importin β and the integral nuclear pore glycoprotein Nup62 facilitating the passage of the untransformed receptor through the nuclear pore.

Components of the CCR4-NOT complex function as nuclear hormone receptor coactivators via association with the NRC-interacting Factor NIF-1.

PubMed

Garapaty, Shivani; Mahajan, Muktar A; Samuels, Herbert H

2008-03-14

CCR4-NOT is an evolutionarily conserved, multicomponent complex known to be involved in transcription as well as mRNA degradation. Various subunits (e.g. CNOT1 and CNOT7/CAF1) have been reported to be involved in influencing nuclear hormone receptor activities. Here, we show that CCR4/CNOT6 and RCD1/CNOT9, members of the CCR4-NOT complex, potentiate nuclear receptor activity. RCD1 interacts in vivo and in vitro with NIF-1 (NRC-interacting factor), a previously characterized nuclear receptor cotransducer that activates nuclear receptors via its interaction with NRC. As with NIF-1, RCD1 and CCR4 do not directly associate with nuclear receptors; however, they enhance ligand-dependent transcriptional activation by nuclear hormone receptors. CCR4 mediates its effect through the ligand binding domain of nuclear receptors and small interference RNA-mediated silencing of endogenous CCR4 results in a marked decrease in nuclear receptor activation. Furthermore, knockdown of CCR4 results in an attenuated stimulation of RARalpha target genes (e.g. Sox9 and HoxA1) as shown by quantitative PCR assays. The silencing of endogenous NIF-1 also resulted in a comparable decrease in the RAR-mediated induction of both Sox9 and HoxA1. Furthermore, CCR4 associates in vivo with NIF-1. In addition, the CCR4-enhanced transcriptional activation by nuclear receptors is dependent on NIF-1. The small interference RNA-mediated knockdown of NIF-1 blocks the ligand-dependent potentiating effect of CCR4. Our results suggest that CCR4 plays a role in the regulation of certain endogenous RARalpha target genes and that RCD1 and CCR4 might mediate their function through their interaction with NIF-1.

Regulation of Liver Energy Balance by the Nuclear Receptors Farnesoid X Receptor and Peroxisome Proliferator Activated Receptor α.

PubMed

Kim, Kang Ho; Moore, David D

2017-01-01

The liver undergoes major changes in substrate utilization and metabolic output over the daily feeding and fasting cycle. These changes occur acutely in response to hormones such as insulin and glucagon, with rapid changes in signaling pathways mediated by protein phosphorylation and other post-translational modifications. They are also reflected in chronic alterations in gene expression in response to nutrient-sensitive transcription factors. Among these, the nuclear receptors farnesoid X receptor (FXR) and peroxisome proliferator activated receptor α (PPARα) provide an intriguing, coordinated response to maintain energy balance in the liver. FXR is activated in the fed state by bile acids returning to the liver, while PPARα is activated in the fasted state in response to the free fatty acids produced by adipocyte lipolysis or possibly other signals. Key Messages: Previous studies indicate that FXR and PPARα have opposing effects on each other's primary targets in key metabolic pathways including gluconeogenesis. Our more recent work shows that these 2 nuclear receptors coordinately regulate autophagy: FXR suppresses this pathway of nutrient and energy recovery, while PPARα activates it. Another recent study indicates that FXR activates the complement and coagulation pathway, while earlier studies identify this as a negative target of PPARα. Since secretion is a very energy- and nutrient-intensive process for hepatocytes, it is possible that FXR licenses it in the nutrient-rich fed state, while PPARα represses it to spare resources in the fasted state. Energy balance is a potential connection linking FXR and PPARα regulation of autophagy and secretion, 2 seemingly unrelated aspects of hepatocyte function. FXR and PPARα act coordinately to promote energy balance and homeostasis in the liver by regulating autophagy and potentially protein secretion. It is quite likely that their impact extends to additional pathways relevant to hepatic energy balance, and

The emerging roles of orphan nuclear receptors in prostate cancer.

PubMed

Wu, Dinglan; Cheung, Alyson; Wang, Yuliang; Yu, Shan; Chan, Franky L

2016-08-01

Orphan nuclear receptors are members of the nuclear receptor (NR) superfamily and are so named because their endogenous physiological ligands are either unknown or may not exist. Because of their important regulatory roles in many key physiological processes, dysregulation of signalings controlled by these receptors is associated with many diseases including cancer. Over years, studies of orphan NRs have become an area of great interest because their specific physiological and pathological roles have not been well-defined, and some of them are promising drug targets for diseases. The recently identified synthetic small molecule ligands, acting as agonists or antagonists, to these orphan NRs not only help to understand better their functional roles but also highlight that the signalings mediated by these ligand-independent NRs in diseases could be therapeutically intervened. This review is a summary of the recent advances in elucidating the emerging functional roles of orphan NRs in cancers, especially prostate cancer. In particular, some orphan NRs, RORγ, TR2, TR4, COUP-IFII, ERRα, DAX1 and SHP, exhibit crosstalk or interference with androgen receptor (AR) signaling in either normal or malignant prostatic cells, highlighting their involvement in prostate cancer progression as androgen and AR signaling pathway play critical roles in this process. We also propose that a better understanding of the mechanism of actions of these orphan NRs in prostate gland or prostate cancer could help to evaluate their potential value as therapeutic targets for prostate cancer. Copyright © 2016 Elsevier B.V. All rights reserved.

A Boolean Network Model of Nuclear Receptor Mediated Cell Cycle Progression

EPA Science Inventory

Nuclear receptors (NRs) are ligand-activated transcription factors that regulate a broad range of cellular processes. Hormones, lipids and xenobiotics have been shown to activate NRs with a range of consequences on development, metabolism, oxidative stress, apoptosis, and prolif...

Novel protective role of the circadian nuclear receptor retinoic acid-related orphan receptor-α in diabetic cardiomyopathy.

PubMed

Zhao, Yichao; Xu, Longwei; Ding, Song; Lin, Nan; Ji, Qingqi; Gao, Lingchen; Su, Yuanyuan; He, Ben; Pu, Jun

2017-04-01

Diabetic cardiomyopathy is a major complication that significantly contributes to morbidity and mortality in diabetics with few therapies. Moreover, antidiabetic drugs reported inconsistent or even adverse cardiovascular effects, suggesting that it is important to exploit novel therapeutic targets against diabetic cardiomyopathy. Here, we observed that the nuclear melatonin receptor, the retinoic acid-related orphan receptor-α (RORα), was downregulated in diabetic hearts. By utilizing a mouse line with RORα disruption, we demonstrated that RORα deficiency led to significantly augmented diastolic dysfunction and cardiac remodeling induced by diabetes. Microscopic and molecular analyses further indicated that the detrimental effects of RORα deficiency were associated with aggravated myocardial apoptosis, autophagy dysfunction, and oxidative stress by disrupting antioxidant gene expression. By contrast, restoration of cardiac RORα levels in transgenic mice significantly improved cardiac functional and structural parameters at 8 weeks after diabetes induction. Consistent with genetic manipulation, pharmacological activation of RORα by melatonin and SR1078 (a synthetic agonist) showed beneficial effects against diabetic cardiomyopathy, while the RORα inhibitor SR3335 significantly exacerbated cardiac impairments in diabetic mice. Collectively, these findings suggest that cardiac-targeted manipulation of nuclear melatonin receptor RORα may hold promise for delaying diabetic cardiomyopathy development. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

The antidepressant fluoxetine normalizes the nuclear glucocorticoid receptor evoked by psychosocial stress

NASA Astrophysics Data System (ADS)

Mitić, M.; Simić, I.; Djordjević, J.; Radojčić, M. B.; Adžić, M.

2011-12-01

Dysregulation of the hypothalamic-pituitary-adrenal (HPA) axis has been implicated in the pathophysiology of depression and stress disorders. Glucocorticoids, key regulators of the stress response, exert diverse effects on cellular processes in the hippocampus. Beside non-genomic pathways, glucocorticoid effects are mediated through activation of the glucocorticoid receptor (GR), a ligand activated transcriptional factor that belongs to the nuclear hormone receptor superfamily. We analysed the GR protein levels both in the cytoplasmic and nuclear compartments of the hippocampus of Wistar rats exposed to chronic psychosocial isolation stress upon chronic fluoxetine (FLU) treatment. Under chronic stress, corticosterone levels (CORT) were decreased compared to the control, and treatment with FLU did not change its level in the stressed rats. At the molecular level, FLU normalized the level of nuclear GR protein in the hippocampus of the stressed rats. Discrepancy between normalization of nuclear GR in the hippocampus and lack of normalization of HPA axis activity judged by CORT, suggests that other brain structures such as the amygdale and prefrontal cortex that also regulate HPA axis activity, seem not to be normalized by the FLU treatment used in our study.

Nuclear localisation of calreticulin in vivo is enhanced by its interaction with glucocorticoid receptors.

PubMed

Roderick, H L; Campbell, A K; Llewellyn, D H

1997-03-24

The multi-functional protein calreticulin (CRT) is normally found within the lumen of the endoplasmic reticulum (ER). However, some of its proposed functions require it to be located within the nucleus, where its presence is contentious. We have investigated this in live COS7, HeLa and LM(TK-) cells using green fluorescent protein (GFP)-fusion proteins. GFP-CRT, and GFP, with an ER signal peptide and a KDEL sequence (ER-GFP), were localised to the ER. In addition, GFP-CRT was located in the nucleus of all the cell types at low levels. The higher levels of nuclear fluorescence in LM(TK-) and HeLa cells suggested that glucocorticoid receptors might enhance nuclear localisation of calreticulin. Dexamethasone treatment of LM(TK-) cells doubled the amount of nuclear GFP-CRT, but did not affect the localisation of a GFP-CRT fusion in which the glucocorticoid receptor-binding N-domain of calreticulin had been deleted. Thus, despite ER targeting and retention signals, calreticulin is also located within the nucleus where its presence increases due to its interaction with glucocorticoid receptors.

Structural Overview of the Nuclear Receptor Superfamily: Insights into Physiology and Therapeutics

PubMed Central

Huang, Pengxiang; Chandra, Vikas; Rastinejad, Fraydoon

2013-01-01

As ligand-regulated transcription factors, the nuclear hormone receptors are nearly ideal drug targets, with internal pockets that bind to hydrophobic, drug-like molecules and well-characterized ligand-induced conformational changes that recruit transcriptional coregulators to promoter elements. Yet, due to the multitude of genes under the control of a single receptor, the major challenge has been the identification of ligands with gene-selective actions, impacting disease outcomes through a narrow subset of target genes and not across their entire gene-regulatory repertoire. Here, we summarize the concepts and work to date underlying the development of steroidal and nonsteroidal receptor ligands, including the use of crystal structures, high-throughput screens, and rational design approaches for finding useful therapeutic molecules. Difficulties in finding selective receptor modulators require a more complete understanding of receptor interdomain communications, posttranslational modifications, and receptor-protein interactions that could be exploited for target gene selectivity. PMID:20148675

Antidiabetic actions of a phosphatidylcholine ligand for nuclear receptor LRH-1

PubMed Central

Lee, Jae Man; Lee, Yoon Kwang; Mamrosh, Jennifer L.; Busby, Scott A.; Griffin, Patrick R.; Pathak, Manish C.; Ortlund, Eric A.; Moore, David D.

2011-01-01

Nuclear hormone receptors regulate diverse metabolic pathways and the orphan nuclear receptor LRH-1 (NR5A2) regulates bile acid biosynthesis1,2. Structural studies have identified phospholipids as potential LRH-1 ligands3–5, but their functional relevance is unclear. Here we show that an unusual phosphatidylcholine species with two saturated 12 carbon fatty acid acyl side chains (dilauroyl phosphatidylcholine, DLPC) is an LRH-1 agonist ligand in vitro. DLPC treatment induces bile acid biosynthetic enzymes in mouse liver, increases bile acid levels, and lowers hepatic triglycerides and serum glucose. DLPC treatment also decreases hepatic steatosis and improves glucose homeostasis in two mouse models of insulin resistance. Both the antidiabetic and lipotropic effects are lost in liver specific Lrh-1 knockouts. These findings identify an LRH-1 dependent phosphatidylcholine signaling pathway that regulates bile acid metabolism and glucose homeostasis. PMID:21614002

Cmr1/WDR76 defines a nuclear genotoxic stress body linking genome integrity and protein quality control.

PubMed

Gallina, Irene; Colding, Camilla; Henriksen, Peter; Beli, Petra; Nakamura, Kyosuke; Offman, Judith; Mathiasen, David P; Silva, Sonia; Hoffmann, Eva; Groth, Anja; Choudhary, Chunaram; Lisby, Michael

2015-03-30

DNA replication stress is a source of genomic instability. Here we identify changed mutation rate 1 (Cmr1) as a factor involved in the response to DNA replication stress in Saccharomyces cerevisiae and show that Cmr1--together with Mrc1/Claspin, Pph3, the chaperonin containing TCP1 (CCT) and 25 other proteins--define a novel intranuclear quality control compartment (INQ) that sequesters misfolded, ubiquitylated and sumoylated proteins in response to genotoxic stress. The diversity of proteins that localize to INQ indicates that other biological processes such as cell cycle progression, chromatin and mitotic spindle organization may also be regulated through INQ. Similar to Cmr1, its human orthologue WDR76 responds to proteasome inhibition and DNA damage by relocalizing to nuclear foci and physically associating with CCT, suggesting an evolutionarily conserved biological function. We propose that Cmr1/WDR76 plays a role in the recovery from genotoxic stress through regulation of the turnover of sumoylated and phosphorylated proteins.

Cmr1/WDR76 defines a nuclear genotoxic stress body linking genome integrity and protein quality control

PubMed Central

Gallina, Irene; Colding, Camilla; Henriksen, Peter; Beli, Petra; Nakamura, Kyosuke; Offman, Judith; Mathiasen, David P.; Silva, Sonia; Hoffmann, Eva; Groth, Anja; Choudhary, Chunaram; Lisby, Michael

2015-01-01

DNA replication stress is a source of genomic instability. Here we identify changed mutation rate 1 (Cmr1) as a factor involved in the response to DNA replication stress in Saccharomyces cerevisiae and show that Cmr1—together with Mrc1/Claspin, Pph3, the chaperonin containing TCP1 (CCT) and 25 other proteins—define a novel intranuclear quality control compartment (INQ) that sequesters misfolded, ubiquitylated and sumoylated proteins in response to genotoxic stress. The diversity of proteins that localize to INQ indicates that other biological processes such as cell cycle progression, chromatin and mitotic spindle organization may also be regulated through INQ. Similar to Cmr1, its human orthologue WDR76 responds to proteasome inhibition and DNA damage by relocalizing to nuclear foci and physically associating with CCT, suggesting an evolutionarily conserved biological function. We propose that Cmr1/WDR76 plays a role in the recovery from genotoxic stress through regulation of the turnover of sumoylated and phosphorylated proteins. PMID:25817432

Nuclear Receptor Variants in Liver Disease

PubMed Central

Müllenbach, Roman; Weber, Susanne N.; Lammert, Frank

2012-01-01

This review aims to provide a snapshot of the actual state of knowledge on genetic variants of nuclear receptors (NR) involved in regulating important aspects of liver metabolism. It recapitulates recent evidence for the application of NR in genetic diagnosis of monogenic (“Mendelian”) liver disease and their use in clinical diagnosis. Genetic analysis of multifactorial liver diseases such as viral hepatitis or fatty liver disease identifies key players in disease predisposition and progression. Evidence from these analyses points towards a role of NR polymorphisms in common diseases, linking regulatory networks to complex and variable phenotypes. The new insights into NR variants also offer perspectives and cautionary advice for their use as handles towards diagnosis and treatment. PMID:22523693

An arginine-rich motif of ring finger protein 4 (RNF4) oversees the recruitment and degradation of the phosphorylated and SUMOylated Krüppel-associated box domain-associated protein 1 (KAP1)/TRIM28 protein during genotoxic stress.

PubMed

Kuo, Ching-Ying; Li, Xu; Kong, Xiang-Qian; Luo, Cheng; Chang, Che-Chang; Chung, Yiyin; Shih, Hsiu-Ming; Li, Keqin Kathy; Ann, David K

2014-07-25

Krüppel-associated box domain-associated protein 1 (KAP1) is a universal transcriptional corepressor that undergoes multiple posttranslational modifications (PTMs), including SUMOylation and Ser-824 phosphorylation. However, the functional interplay of KAP1 PTMs in regulating KAP1 turnover during DNA damage response remains unclear. To decipher the role and cross-talk of multiple KAP1 PTMs, we show here that DNA double strand break-induced KAP1 Ser-824 phosphorylation promoted the recruitment of small ubiquitin-like modifier (SUMO)-targeted ubiquitin E3 ligase, ring finger protein 4 (RNF4), and subsequent RNF4-mediated, SUMO-dependent degradation. Besides the SUMO interacting motif (SIM), a previously unrecognized, but evolutionarily conserved, arginine-rich motif (ARM) in RNF4 acts as a novel recognition motif for selective target recruitment. Results from combined mutagenesis and computational modeling studies suggest that RNF4 utilizes concerted bimodular recognition, namely SIM for Lys-676 SUMOylation and ARM for Ser(P)-824 of simultaneously phosphorylated and SUMOylated KAP1 (Ser(P)-824-SUMO-KAP1). Furthermore, we proved that arginines 73 and 74 within the ARM of RNF4 are required for efficient recruitment to KAP1 or accelerated degradation of promyelocytic leukemia protein (PML) under stress. In parallel, results of bimolecular fluorescence complementation assays validated the role of the ARM in recognizing Ser(P)-824 in living cells. Taken together, we establish that the ARM is required for RNF4 to efficiently target Ser(P)-824-SUMO-KAP1, conferring ubiquitin Lys-48-mediated proteasomal degradation in the context of double strand breaks. The conservation of such a motif may possibly explain the requirement for timely substrate selectivity determination among a myriad of SUMOylated proteins under stress conditions. Thus, the ARM dynamically regulates the SIM-dependent recruitment of targets to RNF4, which could be critical to dynamically fine-tune the

An Arginine-rich Motif of Ring Finger Protein 4 (RNF4) Oversees the Recruitment and Degradation of the Phosphorylated and SUMOylated Krüppel-associated Box Domain-associated Protein 1 (KAP1)/TRIM28 Protein during Genotoxic Stress*

PubMed Central

Kuo, Ching-Ying; Li, Xu; Kong, Xiang-Qian; Luo, Cheng; Chang, Che-Chang; Chung, Yiyin; Shih, Hsiu-Ming; Li, Keqin Kathy; Ann, David K.

2014-01-01

Krüppel-associated box domain-associated protein 1 (KAP1) is a universal transcriptional corepressor that undergoes multiple posttranslational modifications (PTMs), including SUMOylation and Ser-824 phosphorylation. However, the functional interplay of KAP1 PTMs in regulating KAP1 turnover during DNA damage response remains unclear. To decipher the role and cross-talk of multiple KAP1 PTMs, we show here that DNA double strand break-induced KAP1 Ser-824 phosphorylation promoted the recruitment of small ubiquitin-like modifier (SUMO)-targeted ubiquitin E3 ligase, ring finger protein 4 (RNF4), and subsequent RNF4-mediated, SUMO-dependent degradation. Besides the SUMO interacting motif (SIM), a previously unrecognized, but evolutionarily conserved, arginine-rich motif (ARM) in RNF4 acts as a novel recognition motif for selective target recruitment. Results from combined mutagenesis and computational modeling studies suggest that RNF4 utilizes concerted bimodular recognition, namely SIM for Lys-676 SUMOylation and ARM for Ser(P)-824 of simultaneously phosphorylated and SUMOylated KAP1 (Ser(P)-824-SUMO-KAP1). Furthermore, we proved that arginines 73 and 74 within the ARM of RNF4 are required for efficient recruitment to KAP1 or accelerated degradation of promyelocytic leukemia protein (PML) under stress. In parallel, results of bimolecular fluorescence complementation assays validated the role of the ARM in recognizing Ser(P)-824 in living cells. Taken together, we establish that the ARM is required for RNF4 to efficiently target Ser(P)-824-SUMO-KAP1, conferring ubiquitin Lys-48-mediated proteasomal degradation in the context of double strand breaks. The conservation of such a motif may possibly explain the requirement for timely substrate selectivity determination among a myriad of SUMOylated proteins under stress conditions. Thus, the ARM dynamically regulates the SIM-dependent recruitment of targets to RNF4, which could be critical to dynamically fine-tune the

In vitro nuclear receptor inhibition and cytotoxicity of hydraulic fracturing chemicals and their binary mixtures.

PubMed

Bain, Peter A; Kumar, Anu

2018-05-01

The widespread use of hydraulic fracturing (HF) in oil and gas extraction operations has led to concern over environmental risks posed by chemicals used in HF fluids. Here we employed a suite of stable luciferase reporter gene assays to investigate the potential for selected HF chemicals or geogenics to activate or antagonise nuclear receptor signalling. We screened three biocides (bronopol [BP], glutaraldehyde [GA], and tetrakis(hydroxymethyl)phosphonium sulfate [THPS]), a surfactant (2-butoxyethanol), a friction reducer (polyacrylamide), and a coal seam geogenic (o-cresol) for their potential to act as agonists or antagonists of the estrogen receptor, androgen receptor, progesterone receptor (PR), glucocorticoid receptor or peroxisome proliferator-activated receptor gamma (PPARγ). None of the chemicals induced luciferase activity in any of assays used in the study. In antagonistic mode, BP, GA and THPS caused reductions in luciferase activity in the reporter assays at higher concentrations (50-100 μM), while at low concentrations (2-10 μM) GA and THPS enhanced luciferase activity in some assays relative to controls. None of the other tested chemicals exhibited antagonism in the selected assays. In most cases, altered receptor signalling only occurred at concentrations exhibiting cytotoxicity. However, PPARγ activity, and to a lesser extent PR activity, were inhibited by THPS at sub-cytotoxic concentrations. The majority of binary combinations tested exhibited significantly less-than-additive cytotoxicity, and none of the combinations exhibited synergistic cytotoxicity. In summary, the results of the present study indicate that the selected chemicals are not likely to function as direct agonists of the nuclear receptors tested, and only one chemical, THPS was an apparent partial antagonist of two nuclear receptors. Copyright © 2017. Published by Elsevier Ltd.

SUMO and Nucleocytoplasmic Transport.

PubMed

Ptak, Christopher; Wozniak, Richard W

2017-01-01

The transport of proteins between the nucleus and cytoplasm occurs through nuclear pore complexes and is facilitated by numerous transport factors. These transport processes are often regulated by post-translational modification or, reciprocally, transport can function to control post-translational modifications through regulated transport of key modifying enzymes. This interplay extends to relationships between nucleocytoplasmic transport and SUMO-dependent pathways. Examples of protein sumoylation inhibiting or stimulating nucleocytoplasmic transport have been documented, both through its effects on the physical properties of cargo molecules and by directly regulating the functions of components of the nuclear transport machinery. Conversely, the nuclear transport machinery regulates the localization of target proteins and enzymes controlling dynamics of sumoylation and desumoylation thereby affecting the sumoylation state of target proteins. These inter-relationships between SUMO and the nucleocytoplasmic transport machinery, and the varied ways in which they occur, are discussed.

The Peptide Near the C Terminus Regulates Receptor CAR Nuclear Translocation Induced by Xenochemicals in Mouse Liver

PubMed Central

Zelko, Igor; Sueyoshi, Tatsuya; Kawamoto, Takeshi; Moore, Rick; Negishi, Masahiko

2001-01-01

In response to phenobarbital (PB) and other PB-type inducers, the nuclear receptor CAR translocates to the mouse liver nucleus (T. Kawamoto et al., Mol. Cell. Biol. 19:6318–6322, 1999). To define the translocation mechanism, fluorescent protein-tagged human CAR (hCAR) was expressed in the mouse livers using the in situ DNA injection and gene delivery systems. As in the wild-type hCAR, the truncated receptor lacking the C-terminal 10 residues (i.e., AF2 domain) translocated to the nucleus, indicating that the PB-inducible translocation is AF2 independent. Deletion of the 30 C-terminal residues abolished the receptor translocation, and subsequent site-directed mutagenesis delineated the PB-inducible translocation activity of the receptor to the peptide L313GLL316AEL319. Ala mutations of Leu313, Leu316, or Leu319 abrogated the translocation of CAR in the livers, while those of Leu312 or Leu315 did not affect the nuclear translocation. The leucine-rich peptide dictates the nuclear translocation of hCAR in response to various PB-type inducers and appears to be conserved in the mouse and rat receptors. PMID:11283262

Genomic characterization and regulation of CYP3a13: role of xenobiotics and nuclear receptors.

PubMed

Anakk, Sayeepriyadarshini; Kalsotra, Auinash; Shen, Qi; Vu, Mary T; Staudinger, Jeffrey L; Davies, Peter J A; Strobel, Henry W

2003-09-01

We report that CYP3a13 gene, located on mouse chromosome 5, spans 27.5 Kb and contains 13 exons. The transcription start site is 35 bp upstream of the coding region and results in a 109 bp 5' untranslated region. CYP3a13 promoter shows putative binding sites for retinoid X receptor, pregnane X receptor, and estrogen receptor. CYP3a13 shows a broad tissue distribution with predominant expression in liver. Although CYP3a13 shares 92% nucleotide identity with the female-specific rat CYP3A9, its expression does not exhibit sexual dimorphism. Ligand activation of peroxisomal proliferator-activated receptor-gamma and retinoid X receptor inhibit expression of CYP3a13 at the transcription level in a tissue-specific manner. Another novel finding is hepatic induction of CYP3a13 by dexamethasone occurring only in pregnane X receptor null mice. We also report that pregnane X receptor is essential to maintain robust in vivo basal levels of CYP3a13 in contrast to CYP3a11. CYP3a13 protein expressed in vitro can metabolize clinically active drugs ethylmorphine and erythromycin, as well as benzphetamine. We conclude that CYP3a13 is regulated differentially by various nuclear receptors. In humans this may lead to altered drug metabolism, as many of the newly synthesized ligands/drugs targeted toward these nuclear receptors could influence CYP3A gene expression.

Regulation of Stress-Inducible Phosphoprotein 1 Nuclear Retention by Protein Inhibitor of Activated STAT PIAS1

PubMed Central

Soares, Iaci N.; Caetano, Fabiana A.; Pinder, Jordan; Rodrigues, Bruna Roz; Beraldo, Flavio H.; Ostapchenko, Valeriy G.; Durette, Chantal; Pereira, Grace Schenatto; Lopes, Marilene H.; Queiroz-Hazarbassanov, Nicolle; Cunha, Isabela W.; Sanematsu, Paulo I.; Suzuki, Sergio; Bleggi-Torres, Luiz F.; Schild-Poulter, Caroline; Thibault, Pierre; Dellaire, Graham; Martins, Vilma R.; Prado, Vania F.; Prado, Marco A. M.

2013-01-01

Stress-inducible phosphoprotein 1 (STI1), a cochaperone for Hsp90, has been shown to regulate multiple pathways in astrocytes, but its contributions to cellular stress responses are not fully understood. We show that in response to irradiation-mediated DNA damage stress STI1 accumulates in the nucleus of astrocytes. Also, STI1 haploinsufficiency decreases astrocyte survival after irradiation. Using yeast two-hybrid screenings we identified several nuclear proteins as STI1 interactors. Overexpression of one of these interactors, PIAS1, seems to be specifically involved in STI1 nuclear retention and in directing STI1 and Hsp90 to specific sub-nuclear regions. PIAS1 and STI1 co-immunoprecipitate and PIAS1 can function as an E3 SUMO ligase for STI. Using mass spectrometry we identified five SUMOylation sites in STI1. A STI1 mutant lacking these five sites is not SUMOylated, but still accumulates in the nucleus in response to increased expression of PIAS1, suggesting the possibility that a direct interaction with PIAS1 could be responsible for STI1 nuclear retention. To test this possibility, we mapped the interaction sites between PIAS1 and STI1 using yeast-two hybrid assays and surface plasmon resonance and found that a large domain in the N-terminal region of STI1 interacts with high affinity with amino acids 450–480 of PIAS1. Knockdown of PIAS1 in astrocytes impairs the accumulation of nuclear STI1 in response to irradiation. Moreover, a PIAS1 mutant lacking the STI1 binding site is unable to increase STI1 nuclear retention. Interestingly, in human glioblastoma multiforme PIAS1 expression is increased and we found a significant correlation between increased PIAS1 expression and STI1 nuclear localization. These experiments provide evidence that direct interaction between STI1 and PIAS1 is involved in the accumulation of nuclear STI1. This retention mechanism could facilitate nuclear chaperone activity. PMID:23938469

System-wide identification of wild-type SUMO-2 conjugation sites

PubMed Central

Hendriks, Ivo A.; D'Souza, Rochelle C.; Chang, Jer-Gung; Mann, Matthias; Vertegaal, Alfred C. O.

2015-01-01

SUMOylation is a reversible post-translational modification (PTM) regulating all nuclear processes. Identification of SUMOylation sites by mass spectrometry (MS) has been hampered by bulky tryptic fragments, which thus far necessitated the use of mutated SUMO. Here we present a SUMO-specific protease-based methodology which circumvents this problem, dubbed Protease-Reliant Identification of SUMO Modification (PRISM). PRISM allows for detection of SUMOylated proteins as well as identification of specific sites of SUMOylation while using wild-type SUMO. The method is generic and could be widely applied to study lysine PTMs. We employ PRISM in combination with high-resolution MS to identify SUMOylation sites from HeLa cells under standard growth conditions and in response to heat shock. We identified 751 wild-type SUMOylation sites on endogenous proteins, including 200 dynamic SUMO sites in response to heat shock. Thus, we have developed a method capable of quantitatively studying wild-type mammalian SUMO at the site-specific and system-wide level. PMID:26073453

Structural basis for corepressor assembly by the orphan nuclear receptor TLX

PubMed Central

Zhou, X. Edward; He, Yuanzheng; Searose-Xu, Kelvin; Zhang, Chun-Li; Tsai, Chih-Cheng; Melcher, Karsten

2015-01-01

The orphan nuclear receptor TLX regulates neural stem cell self-renewal in the adult brain and functions primarily as a transcription repressor through recruitment of Atrophin corepressors, which bind to TLX via a conserved peptide motif termed the Atro box. Here we report crystal structures of the human and insect TLX ligand-binding domain in complex with Atro box peptides. In these structures, TLX adopts an autorepressed conformation in which its helix H12 occupies the coactivator-binding groove. Unexpectedly, H12 in this autorepressed conformation forms a novel binding pocket with residues from helix H3 that accommodates a short helix formed by the conserved ALXXLXXY motif of the Atro box. Mutations that weaken the TLX–Atrophin interaction compromise the repressive activity of TLX, demonstrating that this interaction is required for Atrophin to confer repressor activity to TLX. Moreover, the autorepressed conformation is conserved in the repressor class of orphan nuclear receptors, and mutations of corresponding residues in other members of this class of receptors diminish their repressor activities. Together, our results establish the functional conservation of the autorepressed conformation and define a key sequence motif in the Atro box that is essential for TLX-mediated repression. PMID:25691470

Structural basis for corepressor assembly by the orphan nuclear receptor TLX.

PubMed

Zhi, Xiaoyong; Zhou, X Edward; He, Yuanzheng; Searose-Xu, Kelvin; Zhang, Chun-Li; Tsai, Chih-Cheng; Melcher, Karsten; Xu, H Eric

2015-02-15

The orphan nuclear receptor TLX regulates neural stem cell self-renewal in the adult brain and functions primarily as a transcription repressor through recruitment of Atrophin corepressors, which bind to TLX via a conserved peptide motif termed the Atro box. Here we report crystal structures of the human and insect TLX ligand-binding domain in complex with Atro box peptides. In these structures, TLX adopts an autorepressed conformation in which its helix H12 occupies the coactivator-binding groove. Unexpectedly, H12 in this autorepressed conformation forms a novel binding pocket with residues from helix H3 that accommodates a short helix formed by the conserved ALXXLXXY motif of the Atro box. Mutations that weaken the TLX-Atrophin interaction compromise the repressive activity of TLX, demonstrating that this interaction is required for Atrophin to confer repressor activity to TLX. Moreover, the autorepressed conformation is conserved in the repressor class of orphan nuclear receptors, and mutations of corresponding residues in other members of this class of receptors diminish their repressor activities. Together, our results establish the functional conservation of the autorepressed conformation and define a key sequence motif in the Atro box that is essential for TLX-mediated repression. © 2015 Zhi et al.; Published by Cold Spring Harbor Laboratory Press.

Structural basis for corepressor assembly by the orphan nuclear receptor TLX

DOE PAGES

Zhi, Xiaoyong; Zhou, X. Edward; He, Yuanzheng; ...

2015-02-15

The orphan nuclear receptor TLX regulates neural stem cell self-renewal in the adult brain and functions primarily as a transcription repressor through recruitment of Atrophin corepressors, which bind to TLX via a conserved peptide motif termed the Atro box. Here we report crystal structures of the human and insect TLX ligand-binding domain in complex with Atro box peptides. In these structures, TLX adopts an autorepressed conformation in which its helix H12 occupies the coactivator-binding groove. Unexpectedly, H12 in this autorepressed conformation forms a novel binding pocket with residues from helix H3 that accommodates a short helix formed by the conservedmore » ALXXLXXY motif of the Atro box. Mutations that weaken the TLX–Atrophin interaction compromise the repressive activity of TLX, demonstrating that this interaction is required for Atrophin to confer repressor activity to TLX. Moreover, the autorepressed conformation is conserved in the repressor class of orphan nuclear receptors, and mutations of corresponding residues in other members of this class of receptors diminish their repressor activities. Together, our results establish the functional conservation of the autorepressed conformation and define a key sequence motif in the Atro box that is essential for TLX-mediated repression.« less

Evidence for triclosan-induced activation of human and rodent xenobiotic nuclear receptors

EPA Science Inventory

The bacteriostat triclosan (2,4,40-trichloro-20-hydroxydiphenylether) (TCS) decreases rat serum thyroxine via putative nuclear receptor (NR) interaction(s) and subsequent transcriptional up-regulation of hepatic catabolism and clearance. However, due to the evolutionary divergenc...

Nuclear receptor coactivators function in estrogen receptor- and progestin receptor-dependent aspects of sexual behavior in female rats

PubMed Central

Molenda-Figueira, Heather A.; Williams, Casey A.; Griffin, Andreana L.; Rutledge, Eric M.; Blaustein, Jeffrey D.; Tetel, Marc J.

2008-01-01

The ovarian hormones, estradiol (E) and progesterone (P) facilitate the expression of sexual behavior in female rats. E and P mediate many of these behavioral effects by binding to their respective intracellular receptors in specific brain regions. Nuclear receptor coactivators, including Steroid Receptor Coactivator-1 (SRC-1) and CREB Binding Protein (CBP), dramatically enhance ligand-dependent steroid receptor transcriptional activity in vitro. Previously, our lab has shown that SRC-1 and CBP modulate estrogen receptor (ER)-mediated induction of progestin receptor (PR) gene expression in the ventromedial nucleus of the hypothalamus (VMN) and hormone-dependent sexual receptivity in female rats. Female sexual behaviors can be activated by high doses of E alone in ovariectomized rats, and thus are believed to be ER-dependent. However, the full repertoire of female sexual behavior, in particular, proceptive behaviors such as hopping, darting and ear wiggling, are considered to be PR-dependent. In the present experiments, the function of SRC-1 and CBP in distinct ER- (Exp. 1) and PR- (Exp. 2) dependent aspects of female sexual behavior was investigated. In Exp. 1, infusion of antisense oligodeoxynucleotides to SRC-1 and CBP mRNA into the VMN decreased lordosis intensity in rats treated with E alone, suggesting that these coactivators modulate ER-mediated female sexual behavior. In Exp. 2, antisense to SRC-1 and CBP mRNA around the time of P administration reduced PR-dependent ear wiggling and hopping and darting. Taken together, these data suggest that SRC-1 and CBP modulate ER and PR action in brain and influence distinct aspects of hormone-dependent sexual behaviors. These findings support our previous studies and provide further evidence that SRC-1 and CBP function together to regulate ovarian hormone action in behaviorally-relevant brain regions. PMID:16769066

SUMO Ligase Protein Inhibitor of Activated STAT1 (PIAS1) Is a Constituent Promyelocytic Leukemia Nuclear Body Protein That Contributes to the Intrinsic Antiviral Immune Response to Herpes Simplex Virus 1

PubMed Central

Brown, James R.; Conn, Kristen L.; Wasson, Peter; Charman, Matthew; Tong, Lily; Grant, Kyle; McFarlane, Steven

2016-01-01

ABSTRACT Aspects of intrinsic antiviral immunity are mediated by promyelocytic leukemia nuclear body (PML-NB) constituent proteins. During herpesvirus infection, these antiviral proteins are independently recruited to nuclear domains that contain infecting viral genomes to cooperatively promote viral genome silencing. Central to the execution of this particular antiviral response is the small ubiquitin-like modifier (SUMO) signaling pathway. However, the participating SUMOylation enzymes are not fully characterized. We identify the SUMO ligase protein inhibitor of activated STAT1 (PIAS1) as a constituent PML-NB protein. We show that PIAS1 localizes at PML-NBs in a SUMO interaction motif (SIM)-dependent manner that requires SUMOylated or SUMOylation-competent PML. Following infection with herpes simplex virus 1 (HSV-1), PIAS1 is recruited to nuclear sites associated with viral genome entry in a SIM-dependent manner, consistent with the SIM-dependent recruitment mechanisms of other well-characterized PML-NB proteins. In contrast to that of Daxx and Sp100, however, the recruitment of PIAS1 is enhanced by PML. PIAS1 promotes the stable accumulation of SUMO1 at nuclear sites associated with HSV-1 genome entry, whereas the accumulation of other evaluated PML-NB proteins occurs independently of PIAS1. We show that PIAS1 cooperatively contributes to HSV-1 restriction through mechanisms that are additive to those of PML and cooperative with those of PIAS4. The antiviral mechanisms of PIAS1 are counteracted by ICP0, the HSV-1 SUMO-targeted ubiquitin ligase, which disrupts the recruitment of PIAS1 to nuclear domains that contain infecting HSV-1 genomes through mechanisms that do not directly result in PIAS1 degradation. IMPORTANCE Adaptive, innate, and intrinsic immunity cooperatively and efficiently restrict the propagation of viral pathogens. Intrinsic immunity mediated by constitutively expressed cellular proteins represents the first line of intracellular defense against

SUMO Ligase Protein Inhibitor of Activated STAT1 (PIAS1) Is a Constituent Promyelocytic Leukemia Nuclear Body Protein That Contributes to the Intrinsic Antiviral Immune Response to Herpes Simplex Virus 1.

PubMed

Brown, James R; Conn, Kristen L; Wasson, Peter; Charman, Matthew; Tong, Lily; Grant, Kyle; McFarlane, Steven; Boutell, Chris

2016-07-01

Aspects of intrinsic antiviral immunity are mediated by promyelocytic leukemia nuclear body (PML-NB) constituent proteins. During herpesvirus infection, these antiviral proteins are independently recruited to nuclear domains that contain infecting viral genomes to cooperatively promote viral genome silencing. Central to the execution of this particular antiviral response is the small ubiquitin-like modifier (SUMO) signaling pathway. However, the participating SUMOylation enzymes are not fully characterized. We identify the SUMO ligase protein inhibitor of activated STAT1 (PIAS1) as a constituent PML-NB protein. We show that PIAS1 localizes at PML-NBs in a SUMO interaction motif (SIM)-dependent manner that requires SUMOylated or SUMOylation-competent PML. Following infection with herpes simplex virus 1 (HSV-1), PIAS1 is recruited to nuclear sites associated with viral genome entry in a SIM-dependent manner, consistent with the SIM-dependent recruitment mechanisms of other well-characterized PML-NB proteins. In contrast to that of Daxx and Sp100, however, the recruitment of PIAS1 is enhanced by PML. PIAS1 promotes the stable accumulation of SUMO1 at nuclear sites associated with HSV-1 genome entry, whereas the accumulation of other evaluated PML-NB proteins occurs independently of PIAS1. We show that PIAS1 cooperatively contributes to HSV-1 restriction through mechanisms that are additive to those of PML and cooperative with those of PIAS4. The antiviral mechanisms of PIAS1 are counteracted by ICP0, the HSV-1 SUMO-targeted ubiquitin ligase, which disrupts the recruitment of PIAS1 to nuclear domains that contain infecting HSV-1 genomes through mechanisms that do not directly result in PIAS1 degradation. Adaptive, innate, and intrinsic immunity cooperatively and efficiently restrict the propagation of viral pathogens. Intrinsic immunity mediated by constitutively expressed cellular proteins represents the first line of intracellular defense against infection. PML

Effects of different ligands on epidermal growth factor receptor (EGFR) nuclear translocation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Faria, Jerusa A.Q.A.; Andrade, Carolina de; Goes, Alfredo M.

The epidermal growth factor receptor (EGFR) is activated through binding to specific ligands and generates signals for proliferation, differentiation, migration, and cell survival. Recent data show the role of nuclear EGFR in tumors. Although many EGFR ligands are upregulated in cancers, little is known about their effects on EGFR nuclear translocation. We have compared the effects of six EGFR ligands (EGF, HB-EGF, TGF-α, β-Cellulin, amphiregulin, and epiregulin) on nuclear translocation of EGFR, receptor phosphorylation, migration, and proliferation. Cell fractionation and confocal immunofluorescence detected EGFR in the nucleus after EGF, HB-EGF, TGF-α and β-Cellulin stimulation in a dose-dependent manner. In contrast,more » amphiregulin and epiregulin did not generate nuclear translocation of EGFR. EGF, HB-EGF, TGF-α and β-Cellulin showed correlations between a higher rate of wound closure and increased phosphorylation of residues in the carboxy-terminus of EGFR, compared to amphiregulin and epiregulin. The data indicate that EGFR is translocated to the nucleus after stimulation with EGF, HB-EGF, TGF-α and β-Cellulin, and that these ligands are related to increased phosphorylation of EGFR tyrosine residues, inducing migration of SkHep-1 cells. - Highlights: • EGF, HB-EGF, TGF-α, β-Cellulin are involved in the EGFR nuclear translocation. • Amphiregulin and epiregulin did not promote nuclear translocation of EGFR. • EGF, HB-EGF, TGF-α and β-Cellulin have a role in SkHep-1 cells migration. • EGFR ligands associated with better prognosis don't stimulate EGFR translocation.« less

Transcriptional corepressor SMILE recruits SIRT1 to inhibit nuclear receptor estrogen receptor-related receptor gamma transactivation.

PubMed

Xie, Yuan-Bin; Park, Jeong-Hoh; Kim, Don-Kyu; Hwang, Jung Hwan; Oh, Sangmi; Park, Seung Bum; Shong, Minho; Lee, In-Kyu; Choi, Hueng-Sik

2009-10-16

SMILE (small heterodimer partner interacting leucine zipper protein) has been identified as a corepressor of the glucocorticoid receptor, constitutive androstane receptor, and hepatocyte nuclear factor 4alpha. Here we show that SMILE also represses estrogen receptor-related receptor gamma (ERRgamma) transactivation. Knockdown of SMILE gene expression increases ERRgamma activity. SMILE directly interacts with ERRgamma in vitro and in vivo. Domain mapping analysis showed that SMILE binds to the AF2 domain of ERRgamma. SMILE represses ERRgamma transactivation partially through competition with coactivators PGC-1alpha, PGC-1beta, and GRIP1. Interestingly, the repression of SMILE on ERRgamma is released by SIRT1 inhibitors, a catalytically inactive SIRT1 mutant, and SIRT1 small interfering RNA but not by histone protein deacetylase inhibitor. In vivo glutathione S-transferase pulldown and coimmunoprecipitation assays validated that SMILE physically interacts with SIRT1. Furthermore, the ERRgamma inverse agonist GSK5182 enhances the interaction of SMILE with ERRgamma and SMILE-mediated repression. Knockdown of SMILE or SIRT1 blocks the repressive effect of GSK5182. Moreover, chromatin immunoprecipitation assays revealed that GSK5182 augments the association of SMILE and SIRT1 on the promoter of the ERRgamma target PDK4. GSK5182 and adenoviral overexpression of SMILE cooperate to repress ERRgamma-induced PDK4 gene expression, and this repression is released by overexpression of a catalytically defective SIRT1 mutant. Finally, we demonstrated that ERRgamma regulates SMILE gene expression, which in turn inhibits ERRgamma. Overall, these findings implicate SMILE as a novel corepressor of ERRgamma and recruitment of SIRT1 as a novel repressive mechanism for SMILE and ERRgamma inverse agonist.

Nuclear receptor-mediated regulation of carboxylesterase expression and activity.

PubMed

Staudinger, Jeff L; Xu, Chenshu; Cui, Yue J; Klaassen, Curtis D

2010-03-01

Emerging evidence demonstrates that several nuclear receptor (NR) family members regulate drug-inducible expression and activity of several important carboxylesterase (CES) enzymes in mammalian liver and intestine. Numerous clinically prescribed anticancer prodrugs, carbamate and pyrethroid insecticides, environmental toxicants and procarcinogens are substrates for CES enzymes. Moreover, a key strategy used in rational drug design frequently utilizes an ester linkage methodology to selectively target a prodrug, or to improve the water solubility of a novel compound. This review summarizes the current state of knowledge regarding NR-mediated regulation of CES enzymes in mammals and highlights their importance in drug metabolism, drug-drug interactions and toxicology. New knowledge regarding the transcriptional regulation of CES enzymes by NR proteins pregnane x receptor (NR1I2) and constitutive androstane receptor (NR1I3) has recently come to light through the use of knockout and transgenic mouse models. Novel insights regarding the species-specific cross-regulation of glucocorticoid receptor (NR3C1) and PPAR-alpha (NR1C1) signaling and CES gene expression are discussed. Elucidation of the role of NR-mediated regulation of CES enzymes in liver and intestine will have a significant impact on rational drug design and the development of novel prodrugs, especially for patients on combination therapy.

A Boolean Network Model of Nuclear Receptor Mediated Cell Cycle Progression (S)

EPA Science Inventory

Nuclear receptors (NRs) are ligand-activated transcription factors that regulate a broad range of cellular processes. Hormones, lipids and xenobiotics have been shown to activate NRs with a range of consequences on development, metabolism, oxidative stress, apoptosis, and prolif...

Body Mass Index Influences the Prognostic Impact of Combined Nuclear Insulin Receptor and Estrogen Receptor Expression in Primary Breast Cancer.

PubMed

Björner, Sofie; Rosendahl, Ann H; Simonsson, Maria; Markkula, Andrea; Jirström, Karin; Borgquist, Signe; Rose, Carsten; Ingvar, Christian; Jernström, Helena

2017-01-01

The prognostic importance of tumor-specific nuclear insulin receptor (InsR) expression in breast cancer is unclear, while membrane and cytoplasmic localization of InsR is better characterized. The insulin signaling network is influenced by obesity and may interact with the estrogen receptor α (ERα) signaling. The purpose was to investigate the interplay between nuclear InsR, ER, body mass index (BMI), and prognosis. Tumor-specific expression of nuclear InsR was evaluated by immunohistochemistry in tissue microarrays from 900 patients with primary invasive breast cancer without preoperative treatment, included in a population-based cohort in Sweden (2002-2012) in relation to prognosis. Patients were followed for up to 11 years during which 107 recurrences were observed. Nuclear InsR + expression was present in 214 patients (23.8%) and increased with longer time between surgery and staining ( P  < 0.001). There were significant effect modifications by ER status and BMI in relation to clinical outcomes. Nuclear InsR + conferred higher recurrence-risk in patients with ER + tumors, but lower risk in patients with ER - tumors ( P interaction  = 0.003). Normal-weight patients with nuclear InsR + tumors had higher recurrence-risk, while overweight or obese patients had half the recurrence-risk compared to patients with nuclear InsR - tumors ( P interaction  = 0.007). Normal-weight patients with a nuclear InsR - /ER + tumor had the lowest risk for recurrence compared to all other nuclear InsR/ER combinations [HR adj 0.50, 95% confidence interval (CI): 0.25-0.97], while overweight or obese patients with nuclear InsR - /ER - tumors had the worst prognosis (HR adj 7.75, 95% CI: 2.04-29.48). Nuclear InsR was more prognostic than ER among chemotherapy-treated patients. In summary, nuclear InsR may have prognostic impact among normal-weight patients with ER + tumors and in overweight or obese patients with ER - tumors. Normal-weight patients with nuclear Ins

Characterization of germ cell-specific expression of the orphan nuclear receptor, germ cell nuclear factor.

PubMed

Katz, D; Niederberger, C; Slaughter, G R; Cooney, A J

1997-10-01

Nuclear receptors, such as those for androgens, estrogens, and progesterones, control many reproductive processes. Proteins with structures similar to these receptors, but for which ligands have not yet been identified, have been termed orphan nuclear receptors. One of these orphans, germ cell nuclear factor (GCNF), has been shown to be germ cell specific in the adult and, therefore, may also participate in the regulation of reproductive functions. In this paper, we examine more closely the expression patterns of GCNF in germ cells to begin to define spatio-temporal domains of its activity. In situ hybridization showed that GCNF messenger RNA (mRNA) is lacking in the testis of hypogonadal mutant mice, which lack developed spermatids, but is present in the wild-type testis. Thus, GCNF is, indeed, germ cell specific in the adult male. Quantitation of the specific in situ hybridization signal in wild-type testis reveals that GCNF mRNA is most abundant in stage VII round spermatids. Similarly, Northern analysis and specific in situ hybridization show that GCNF expression first occurs in testis of 20-day-old mice, when round spermatids first emerge. Therefore, in the male, GCNF expression occurs postmeiotically and may participate in the morphological changes of the maturing spermatids. In contrast, female expression of GCNF is shown in growing oocytes that have not completed the first meiotic division. Thus, GCNF in the female is expressed before the completion of meiosis. Finally, the nature of the two different mRNAs that hybridize to the GCNF complementary DNA was studied. Although both messages contain the DNA binding domain, only the larger message is recognized by a probe from the extreme 3' untranslated region. In situ hybridization with these differential probes demonstrates that both messages are present in growing oocytes. In addition, the coding region and portions of the 3' untranslated region of the GCNF complementary DNA are conserved in the rat.

Cloning retinoid and peroxisome proliferator-activated nuclear receptors of the Pacific oyster and in silico binding to environmental chemicals

PubMed Central

Vogeler, Susanne; Galloway, Tamara S.; Isupov, Michail

2017-01-01

Disruption of nuclear receptors, a transcription factor superfamily regulating gene expression in animals, is one proposed mechanism through which pollution causes effects in aquatic invertebrates. Environmental pollutants have the ability to interfere with the receptor’s functions through direct binding and inducing incorrect signals. Limited knowledge of invertebrate endocrinology and molecular regulatory mechanisms, however, impede the understanding of endocrine disruptive effects in many aquatic invertebrate species. Here, we isolated three nuclear receptors of the Pacific oyster, Crassostrea gigas: two isoforms of the retinoid X receptor, CgRXR-1 and CgRXR-2, a retinoic acid receptor ortholog CgRAR, and a peroxisome proliferator-activated receptor ortholog CgPPAR. Computer modelling of the receptors based on 3D crystal structures of human proteins was used to predict each receptor’s ability to bind to different ligands in silico. CgRXR showed high potential to bind and be activated by 9-cis retinoic acid and the organotin tributyltin (TBT). Computer modelling of CgRAR revealed six residues in the ligand binding domain, which prevent the successful interaction with natural and synthetic retinoid ligands. This supports an existing theory of loss of retinoid binding in molluscan RARs. Modelling of CgPPAR was less reliable due to high discrepancies in sequence to its human ortholog. Yet, there are suggestions of binding to TBT, but not to rosiglitazone. The effect of potential receptor ligands on early oyster development was assessed after 24h of chemical exposure. TBT oxide (0.2μg/l), all-trans retinoic acid (ATRA) (0.06 mg/L) and perfluorooctanoic acid (20 mg/L) showed high effects on development (>74% abnormal developed D-shelled larvae), while rosiglitazone (40 mg/L) showed no effect. The results are discussed in relation to a putative direct (TBT) disruption effect on nuclear receptors. The inability of direct binding of ATRA to CgRAR suggests either a

Nuclear receptors HR96 and ultraspiracle from the fall armyworm (Spodoptera frugiperda), developmental expression and induction by xenobiotics.

PubMed

Giraudo, Maeva; Audant, Pascaline; Feyereisen, René; Le Goff, Gaëlle

2013-05-01

The fall armyworm Spodoptera frugiperda is a major polyphagous pest in agriculture and little is known on how this insect can adapt to the diverse and potentially toxic plant allelochemicals that they ingest or to insecticides. To investigate the involvement of nuclear receptors in the response of S. frugiperda to its chemical environment, we cloned SfHR96, a nuclear receptor orthologous to the mammalian xenobiotic receptors, pregnane X receptor (PXR) and constitutive androstane receptor (CAR). We also cloned ultraspiracle (USP), the ortholog of retinoid X receptor (RXR) that serves as partner of dimerization of PXR and CAR. Cloning of SfUSP revealed the presence of two isoforms, SfUSP-1 and SfUSP-2 in this species, that differ in their N-terminal region. The expression of these receptors as well as the ecdysone receptor was studied during specific steps of development in different tissues. SfHR96 was constitutively expressed in larval midgut, fat body and Malpighian tubules throughout the last two instars and pupal stage, as well as in Sf9 cells. EcR and SfUSP-2 showed peaks of expression before larval moults and during metamorphosis, whereas SfUSP-1 was mainly expressed in the pre-pupal stage. Receptor induction was followed after exposure of larvae or cells to 11 chemical compounds. SfHR96 was not inducible by the tested compounds. EcR was significantly induced by the 20-hydroxyecdysone agonist, methoxyfenozide, and SfUSP showed an increase expression when exposed to the juvenile hormone analog, methoprene. The cloning of these nuclear receptors is a first step in understanding the important capacities of adaptation of this insect pest. Copyright © 2013 Elsevier Ltd. All rights reserved.

Role of osteoprotegerin/receptor activator of nuclear factor kappa B/receptor activator of nuclear factor kappa B ligand axis in nonalcoholic fatty liver disease.

PubMed

Pacifico, Lucia; Andreoli, Gian Marco; D'Avanzo, Miriam; De Mitri, Delia; Pierimarchi, Pasquale

2018-05-21

Concomitantly with the increase in the prevalences of overweight/obesity, nonalcoholic fatty liver disease (NAFLD) has worldwide become the main cause of chronic liver disease in both adults and children. Patients with fatty liver display features of metabolic syndrome (MetS), like insulin resistance (IR), glucose intolerance, hypertension and dyslipidemia. Recently, epidemiological studies have linked obesity, MetS, and NAFLD to decreased bone mineral density and osteoporosis, highlighting an intricate interplay among bone, adipose tissue, and liver. Osteoprotegerin (OPG), an important symbol of the receptor activator of nuclear factor-B ligand/receptor activator of nuclear factor kappa B/OPG system activation, typically considered for its role in bone metabolism, may also play critical roles in the initiation and perpetuation of obesity-related comorbidities. Clinical data have indicated that OPG concentrations are associated with hypertension, left ventricular hypertrophy, vascular calcification, endothelial dysfunction, and severity of liver damage in chronic hepatitis C. Nonetheless, the relationship between circulating OPG and IR as a key feature of MetS as well as between OPG and NAFLD remains uncertain. Thus, the aims of the present review are to provide the existent knowledge on these associations and to discuss briefly the underlying mechanisms linking OPG and NAFLD.

Fatty Acid Amide Hydrolase (FAAH) Inhibition Enhances Memory Acquisition through Activation of PPAR-alpha Nuclear Receptors

ERIC Educational Resources Information Center

Mazzola, Carmen; Medalie, Julie; Scherma, Maria; Panlilio, Leigh V.; Solinas, Marcello; Tanda, Gianluigi; Drago, Filippo; Cadet, Jean Lud; Goldberg, Steven R.; Yasar, Sevil

2009-01-01

Inhibitors of fatty acid amide hydrolase (FAAH) increase endogenous levels of anandamide (a cannabinoid CB[subscript 1]-receptor ligand) and oleoylethanolamide and palmitoylethanolamide (OEA and PEA, ligands for alpha-type peroxisome proliferator-activated nuclear receptors, PPAR-alpha) when and where they are naturally released in the brain.…

Cell-type-specific regulation of the retinoic acid receptor mediated by the orphan nuclear receptor TLX.

PubMed

Kobayashi, M; Yu, R T; Yasuda, K; Umesono, K

2000-12-01

Malformations in the eye can be caused by either an excess or deficiency of retinoids. An early target gene of the retinoid metabolite, retinoic acid (RA), is that encoding one of its own receptors, the retinoic acid receptor beta (RARbeta). To better understand the mechanisms underlying this autologous regulation, we characterized the chick RARbeta2 promoter. The region surrounding the transcription start site of the avian RARbeta2 promoter is over 90% conserved with the corresponding region in mammals and confers strong RA-dependent transactivation in primary cultured embryonic retina cells. This response is selective for RAR but not retinoid X receptor-specific agonists, demonstrating a principal role for RAR(s) in retina cells. Retina cells exhibit a far higher sensitivity to RA than do fibroblasts or osteoblasts, a property we found likely due to expression of the orphan nuclear receptor TLX. Ectopic expression of TLX in fibroblasts resulted in increased sensitivity to RA induction, an effect that is conserved between chick and mammals. We have identified a cis element, the silencing element relieved by TLX (SET), within the RARbeta2 promoter region which confers TLX- and RA-dependent transactivation. These results indicate an important role for TLX in autologous regulation of the RARbeta gene in the eye.

Cell-Type-Specific Regulation of the Retinoic Acid Receptor Mediated by the Orphan Nuclear Receptor TLX†

PubMed Central

Kobayashi, Mime; Yu, Ruth T.; Yasuda, Kunio; Umesono, Kazuhiko

2000-01-01

Malformations in the eye can be caused by either an excess or deficiency of retinoids. An early target gene of the retinoid metabolite, retinoic acid (RA), is that encoding one of its own receptors, the retinoic acid receptor β (RARβ). To better understand the mechanisms underlying this autologous regulation, we characterized the chick RARβ2 promoter. The region surrounding the transcription start site of the avian RARβ2 promoter is over 90% conserved with the corresponding region in mammals and confers strong RA-dependent transactivation in primary cultured embryonic retina cells. This response is selective for RAR but not retinoid X receptor-specific agonists, demonstrating a principal role for RAR(s) in retina cells. Retina cells exhibit a far higher sensitivity to RA than do fibroblasts or osteoblasts, a property we found likely due to expression of the orphan nuclear receptor TLX. Ectopic expression of TLX in fibroblasts resulted in increased sensitivity to RA induction, an effect that is conserved between chick and mammals. We have identified a cis element, the silencing element relieved by TLX (SET), within the RARβ2 promoter region which confers TLX- and RA-dependent transactivation. These results indicate an important role for TLX in autologous regulation of the RARβ gene in the eye. PMID:11073974

Genetic incorporation of Nε-acetyllysine reveals a novel acetylation-sumoylation switch in yeast.

PubMed

Kim, Sang-Woo; Lee, Kyung Jin; Kim, Sinil; Kim, Jihyo; Cho, Kyukwang; Ro, Hyeon-Su; Park, Hee-Sung

2017-11-01

The lysine acetylation of proteins plays a key role in regulating protein functions, thereby controlling a wide range of cellular processes. Despite the prevalence and significance of lysine acetylation in eukaryotes, however, its systematic study has been challenged by the technical limitations of conventional approaches for selective lysine acetylation in vivo. Here, we report the in vivo study of lysine acetylation via the genetic incorporation of N ε -acetyllysine in yeast. We demonstrate that a newly discovered acetylation-sumoylation switch precisely controls the localization and cellular function of the yeast septin protein, Cdc11, during the cell cycle. This approach should facilitate the comprehensive in vivo study of lysine acetylation across a wide range of proteins in eukaryotic organisms. This article is part of a Special Issue entitled "Biochemistry of Synthetic Biology - Recent Developments" Guest Editor: Dr. Ilka Heinemann and Dr. Patrick O'Donoghue. Copyright © 2017 Elsevier B.V. All rights reserved.

Control of energy balance by hypothalamic gene circuitry involving two nuclear receptors, neuron-derived orphan receptor 1 and glucocorticoid receptor.

PubMed

Kim, Sun-Gyun; Lee, Bora; Kim, Dae-Hwan; Kim, Juhee; Lee, Seunghee; Lee, Soo-Kyung; Lee, Jae W

2013-10-01

Nuclear receptors (NRs) regulate diverse physiological processes, including the central nervous system control of energy balance. However, the molecular mechanisms for the central actions of NRs in energy balance remain relatively poorly defined. Here we report a hypothalamic gene network involving two NRs, neuron-derived orphan receptor 1 (NOR1) and glucocorticoid receptor (GR), which directs the regulated expression of orexigenic neuropeptides agouti-related peptide (AgRP) and neuropeptide Y (NPY) in response to peripheral signals. Our results suggest that the anorexigenic signal leptin induces NOR1 expression likely via the transcription factor cyclic AMP response element-binding protein (CREB), while the orexigenic signal glucocorticoid mobilizes GR to inhibit NOR1 expression by antagonizing the action of CREB. Also, NOR1 suppresses glucocorticoid-dependent expression of AgRP and NPY. Consistently, relative to wild-type mice, NOR1-null mice showed significantly higher levels of AgRP and NPY and were less responsive to leptin in decreasing the expression of AgRP and NPY. These results identify mutual antagonism between NOR1 and GR to be a key rheostat for peripheral metabolic signals to centrally control energy balance.

Nuclear export receptor CRM1 recognizes diverse conformations in nuclear export signals.

PubMed

Fung, Ho Yee Joyce; Fu, Szu-Chin; Chook, Yuh Min

2017-03-10

Nuclear export receptor CRM1 binds highly variable nuclear export signals (NESs) in hundreds of different cargoes. Previously we have shown that CRM1 binds NESs in both polypeptide orientations (Fung et al., 2015). Here, we show crystal structures of CRM1 bound to eight additional NESs which reveal diverse conformations that range from loop-like to all-helix, which occupy different extents of the invariant NES-binding groove. Analysis of all NES structures show 5-6 distinct backbone conformations where the only conserved secondary structural element is one turn of helix that binds the central portion of the CRM1 groove. All NESs also participate in main chain hydrogen bonding with human CRM1 Lys568 side chain, which acts as a specificity filter that prevents binding of non-NES peptides. The large conformational range of NES backbones explains the lack of a fixed pattern for its 3-5 hydrophobic anchor residues, which in turn explains the large array of peptide sequences that can function as NESs.

Deciphering the Regulatory Logic of an Ancient, Ultraconserved Nuclear Receptor Enhancer Module

PubMed Central

Bagamasbad, Pia D.; Bonett, Ronald M.; Sachs, Laurent; Buisine, Nicolas; Raj, Samhitha; Knoedler, Joseph R.; Kyono, Yasuhiro; Ruan, Yijun; Ruan, Xiaoan

2015-01-01

Cooperative, synergistic gene regulation by nuclear hormone receptors can increase sensitivity and amplify cellular responses to hormones. We investigated thyroid hormone (TH) and glucocorticoid (GC) synergy on the Krüppel-like factor 9 (Klf9) gene, which codes for a zinc finger transcription factor involved in development and homeostasis of diverse tissues. We identified regions of the Xenopus and mouse Klf9 genes 5–6 kb upstream of the transcription start sites that supported synergistic transactivation by TH plus GC. Within these regions, we found an orthologous sequence of approximately 180 bp that is highly conserved among tetrapods, but absent in other chordates, and possesses chromatin marks characteristic of an enhancer element. The Xenopus and mouse approximately 180-bp DNA element conferred synergistic transactivation by hormones in transient transfection assays, so we designate this the Klf9 synergy module (KSM). We identified binding sites within the mouse KSM for TH receptor, GC receptor, and nuclear factor κB. TH strongly increased recruitment of liganded GC receptor and serine 5 phosphorylated (initiating) RNA polymerase II to chromatin at the KSM, suggesting a mechanism for transcriptional synergy. The KSM is transcribed to generate long noncoding RNAs, which are also synergistically induced by combined hormone treatment, and the KSM interacts with the Klf9 promoter and a far upstream region through chromosomal looping. Our findings support that the KSM plays a central role in hormone regulation of vertebrate Klf9 genes, it evolved in the tetrapod lineage, and has been maintained by strong stabilizing selection. PMID:25866873

Allosteric Pathways in the PPARγ-RXRα nuclear receptor complex

NASA Astrophysics Data System (ADS)

Ricci, Clarisse G.; Silveira, Rodrigo L.; Rivalta, Ivan; Batista, Victor S.; Skaf, Munir S.

2016-01-01

Understanding the nature of allostery in DNA-nuclear receptor (NR) complexes is of fundamental importance for drug development since NRs regulate the transcription of a myriad of genes in humans and other metazoans. Here, we investigate allostery in the peroxisome proliferator-activated/retinoid X receptor heterodimer. This important NR complex is a target for antidiabetic drugs since it binds to DNA and functions as a transcription factor essential for insulin sensitization and lipid metabolism. We find evidence of interdependent motions of Ω-loops and PPARγ-DNA binding domain with contacts susceptible to conformational changes and mutations, critical for regulating transcriptional functions in response to sequence-dependent DNA dynamics. Statistical network analysis of the correlated motions, observed in molecular dynamics simulations, shows preferential allosteric pathways with convergence centers comprised of polar amino acid residues. These findings are particularly relevant for the design of allosteric modulators of ligand-dependent transcription factors.

Nuclear hormone receptor NHR-49 controls fat consumption and fatty acid composition in C. elegans.

PubMed

Van Gilst, Marc R; Hadjivassiliou, Haralambos; Jolly, Amber; Yamamoto, Keith R

2005-02-01

Mammalian nuclear hormone receptors (NHRs), such as liver X receptor, farnesoid X receptor, and peroxisome proliferator-activated receptors (PPARs), precisely control energy metabolism. Consequently, these receptors are important targets for the treatment of metabolic diseases, including diabetes and obesity. A thorough understanding of NHR fat regulatory networks has been limited, however, by a lack of genetically tractable experimental systems. Here we show that deletion of the Caenorhabditis elegans NHR gene nhr-49 yielded worms with elevated fat content and shortened life span. Employing a quantitative RT-PCR screen, we found that nhr-49 influenced the expression of 13 genes involved in energy metabolism. Indeed, nhr-49 served as a key regulator of fat usage, modulating pathways that control the consumption of fat and maintain a normal balance of fatty acid saturation. We found that the two phenotypes of the nhr-49 knockout were linked to distinct pathways and were separable: The high-fat phenotype was due to reduced expression of enzymes in fatty acid beta-oxidation, and the shortened adult life span resulted from impaired expression of a stearoyl-CoA desaturase. Despite its sequence relationship with the mammalian hepatocyte nuclear factor 4 receptor, the biological activities of nhr-49 were most similar to those of the mammalian PPARs, implying an evolutionarily conserved role for NHRs in modulating fat consumption and composition. Our findings in C. elegans provide novel insights into how NHR regulatory networks are coordinated to govern fat metabolism.

Estrogen-related receptor β (ERRβ) – renaissance receptor or receptor renaissance?

PubMed Central

Divekar, Shailaja D.; Tiek, Deanna M.; Fernandez, Aileen; Riggins, Rebecca B.

2016-01-01

Estrogen-related receptors (ERRs) are founding members of the orphan nuclear receptor (ONR) subgroup of the nuclear receptor superfamily. Twenty-seven years of study have yet to identify cognate ligands for the ERRs, though they have firmly placed ERRα and ERRγ at the intersection of cellular metabolism and oncogenesis. The pace of discovery for novel functions of ERRβ, however, has until recently been somewhat slower than that of its family members. ERRβ has also been largely ignored in summaries and perspectives of the ONR literature. Here, we provide an overview of established and emerging knowledge of ERRβ in mouse, man, and other species, highlighting unique aspects of ERRβ biology that set it apart from the other two estrogen-related receptors, with a focus on the impact of alternative splicing on the structure and function of this receptor. PMID:27507929

Nuclear pool of phosphatidylinositol 4 phosphate 5 kinase 1α is modified by polySUMO-2 during apoptosis.

PubMed

Chakrabarti, Rajarshi; Bhowmick, Debajit; Bhargava, Varsha; Bhar, Kaushik; Siddhanta, Anirban

2013-09-20

Phosphatidylinositol 4 phosphate 5 kinase 1α (PIP5K) is mainly localized in the cytosol and plasma membrane. Studies have also indicated its prominent association with nuclear speckles. The exact nature of this nuclear pool of PIP5K is not clear. Using biochemical and microscopic techniques, we have demonstrated that the nuclear pool of PIP5K is modified by SUMO-1 in HEK-293 cells stably expressing PIP5K. Moreover, this SUMOylated pool of PIP5K increased during apoptosis. PolySUMO-2 chain conjugated PIP5K was detected by pull-down experiment using affinity-tagged RNF4, a polySUMO-2 binding protein, during late apoptosis. Copyright © 2013 Elsevier Inc. All rights reserved.

rigor mortis encodes a novel nuclear receptor interacting protein required for ecdysone signaling during Drosophila larval development.

PubMed

Gates, Julie; Lam, Geanette; Ortiz, José A; Losson, Régine; Thummel, Carl S

2004-01-01

Pulses of the steroid hormone ecdysone trigger the major developmental transitions in Drosophila, including molting and puparium formation. The ecdysone signal is transduced by the EcR/USP nuclear receptor heterodimer that binds to specific response elements in the genome and directly regulates target gene transcription. We describe a novel nuclear receptor interacting protein encoded by rigor mortis (rig) that is required for ecdysone responses during larval development. rig mutants display defects in molting, delayed larval development, larval lethality, duplicated mouth parts, and defects in puparium formation--phenotypes that resemble those seen in EcR, usp, E75A and betaFTZ-F1 mutants. Although the expression of these nuclear receptor genes is essentially normal in rig mutant larvae, the ecdysone-triggered switch in E74 isoform expression is defective. rig encodes a protein with multiple WD-40 repeats and an LXXLL motif, sequences that act as specific protein-protein interaction domains. Consistent with the presence of these elements and the lethal phenotypes of rig mutants, Rig protein interacts with several Drosophila nuclear receptors in GST pull-down experiments, including EcR, USP, DHR3, SVP and betaFTZ-F1. The ligand binding domain of betaFTZ-F1 is sufficient for this interaction, which can occur in an AF-2-independent manner. Antibody stains reveal that Rig protein is present in the brain and imaginal discs of second and third instar larvae, where it is restricted to the cytoplasm. In larval salivary gland and midgut cells, however, Rig shuttles between the cytoplasm and nucleus in a spatially and temporally regulated manner, at times that correlate with the major lethal phase of rig mutants and major switches in ecdysone-regulated gene expression. Taken together, these data indicate that rig exerts essential functions during larval development through gene-specific effects on ecdysone-regulated transcription, most likely as a cofactor for one or more

pSumo-CD: predicting sumoylation sites in proteins with covariance discriminant algorithm by incorporating sequence-coupled effects into general PseAAC.

PubMed

Jia, Jianhua; Zhang, Liuxia; Liu, Zi; Xiao, Xuan; Chou, Kuo-Chen

2016-10-15

Sumoylation is a post-translational modification (PTM) process, in which small ubiquitin-related modifier (SUMO) is attaching by covalent bonds to substrate protein. It is critical to many different biological processes such as replicating genome, expressing gene, localizing and stabilizing proteins; unfortunately, it is also involved with many major disorders including Alzheimer's and Parkinson's diseases. Therefore, for both basic research and drug development, it is important to identify the sumoylation sites in proteins. To address such a problem, we developed a predictor called pSumo-CD by incorporating the sequence-coupled information into the general pseudo-amino acid composition (PseAAC) and introducing the covariance discriminant (CD) algorithm, in which a bias-adjustment term, which has the function to automatically adjust the errors caused by the bias due to the imbalance of training data, had been incorporated. Rigorous cross-validations indicated that the new predictor remarkably outperformed the existing state-of-the-art prediction method for the same purpose. For the convenience of most experimental scientists, a user-friendly web-server for pSumo-CD has been established at http://www.jci-bioinfo.cn/pSumo-CD, by which users can easily obtain their desired results without the need to go through the complicated mathematical equations involved. jjia@gordonlifescience.org, xxiao@gordonlifescience.org or kcchou@gordonlifescience.orgSupplementary information: Supplementary data are available at Bioinformatics online. © The Author 2016. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

Annotation of the Nuclear Receptors in an Estuarine Fish species, Fundulus heteroclitus

PubMed Central

Baldwin, William S.; Boswell, W. Tyler; Ginjupalli, Gautam; Litoff, Elizabeth J.

2017-01-01

The nuclear receptors (NRs) are ligand-dependent transcription factors that respond to various internal as well as external cues such as nutrients, pheromones, and steroid hormones that play crucial roles in regulation and maintenance of homeostasis and orchestrating the physiological and stress responses of an organism. We annotated the Fundulus heteroclitus (mummichog; Atlantic killifish) nuclear receptors. Mummichog are a non-migratory, estuarine fish with a limited home range often used in environmental research as a field model for studying ecological and evolutionary responses to variable environmental conditions such as salinity, oxygen, temperature, pH, and toxic compounds because of their hardiness. F. heteroclitus have at least 74 NRs spanning all seven gene subfamilies. F. heteroclitus is unique in that no RXRα member was found within the genome. Interestingly, some of the NRs are highly conserved between species, while others show a higher degree of divergence such as PXR, SF1, and ARα. Fundulus like other fish species show expansion of the RAR (NR1B), Rev-erb (NR1D), ROR (NR1F), COUPTF (NR2F), ERR (NR3B), RXR (NR2B), and to a lesser extent the NGF (NR4A), and NR3C steroid receptors (GR/AR). Of particular interest is the co-expansion of opposing NRs, Reverb-ROR, and RAR/RXR-COUPTF. PMID:28804711

Retinoic Acid Inducible Gene 1 Protein (RIG1)-like Receptor Pathway is Required for Efficient Nuclear Reprogramming

PubMed Central

Sayed, Nazish; Ospino, Frank; Himmati, Farhan; Lee, Jieun; Chanda, Palas; Mocarski, Edward S.; Cooke, John P.

2017-01-01

We have revealed a critical role for innate immune signaling in nuclear reprogramming to pluripotency, and in the nuclear reprogramming required for somatic cell transdifferentiation. Activation of innate immune signaling causes global changes in the expression and activity of epigenetic modifiers to promote epigenetic plasticity. In our previous papers, we focused on the role of toll-like receptor 3 (TLR3) in this signaling pathway. Here we define the role of another innate immunity pathway known to participate in the response to viral RNA, the retinoic acid-inducible gene 1 receptor (RIG-1)-like receptor (RLR) pathway. This pathway is represented by the sensors of viral RNA, RIG-1, LGP2 and MDA5. We first found that TLR3 deficiency only causes a partial inhibition of nuclear reprogramming to pluripotency in mouse tail-tip fibroblasts, which motivated us to determine the contribution of RLR. We found that knockdown of iPS-1, the common adaptor protein for the RLR family, substantially reduced nuclear reprogramming induced by retroviral or by mmRNA expression of Oct 4, Sox2, KLF4 and cMYC (OSKM). Importantly a double knockdown of both RLR and TLR3 pathway led to a further decrease in iPSC colonies suggesting an additive effect of both these pathways on nuclear reprogramming. Furthermore, in murine embryonic fibroblasts expressing a dox-inducible cassette of the genes encoding OSKM, an RLR agonist increased the yield of iPSCs. Similarly, the RLR agonist enhanced nuclear reprogramming by cell permeant peptides of the Yamanaka factors. Finally, in the dox-inducible system, RLR activation promotes activating histone marks in the promoter region of pluripotency genes. To conclude, innate immune signaling mediated by RLR plays a critical role in nuclear reprogramming. Manipulation of innate immune signaling may facilitate nuclear reprogramming to achieve pluripotency. PMID:28276156

DOE Office of Scientific and Technical Information (OSTI.GOV)

Nishida, Tamotsu, E-mail: nishida@gene.mie-u.ac.jp; Yamada, Yoshiji

Parkin-interacting substrate (PARIS), a member of the family of Krüppel-associated box (KRAB)-containing zinc-finger transcription factors, is a substrate of the ubiquitin E3 ligase parkin. PARIS represses the expression of peroxisome proliferator-activated receptor γ coactivator-1α (PGC-1α), although the underlying mechanisms remain largely unknown. In the present study, we demonstrate that PARIS can be SUMOylated, and its SUMOylation plays a role in the repression of PGC-1a promoter activity. Protein inhibitor of activated STAT y (PIASy) was identified as an interacting protein of PARIS and shown to enhance its SUMOylation. PIASy repressed PGC-1a promoter activity, and this effect was attenuated by PARIS inmore » a manner dependent on its SUMOylation status. Co-expression of SUMO-1 with PIASy completely repressed PGC-1a promoter activity independently of PARIS expression. PARIS-mediated PGC-1a promoter repression depended on the activity of histone deacetylases (HDAC), whereas PIASy repressed the PGC-1a promoter in an HDAC-independent manner. Taken together, these results suggest that PARIS and PIASy modulate PGC-1a gene transcription through distinct molecular mechanisms. -- Highlights: •PARIS can be SUMOylated in vivo and in vitro. •SUMOylation of PARIS functions in the repression of PGC-1a promoter activity. •PIASy interacts with PARIS and enhances its SUMOylation. •PIASy influences PARIS-mediated repression of PGC-1a promoter activity.« less

SUMOylation of HSP27 by small ubiquitin-like modifier 2/3 promotes proliferation and invasion of hepatocellular carcinoma cells.

PubMed

Ge, Haize; Du, Juan; Xu, Jingman; Meng, Xiangliang; Tian, Jinchuan; Yang, Jie; Liang, Huimin

2017-08-03

Primary hepatocellular carcinoma (PHC) is a major health problem worldwide and is one of the 10 most commonly diagnosed cancers in China. Heat shock protein 27 (HSP27) were found to be overexpressed in a wide range of malignancies including PHC, however, post-translational modification of HSP27 still needs exploration in PHC. Recently, SUMOylation, an important post-translational modification associating with the development of many kinds of cancers has been intensively studied. In the current study, mRNA and protein level of HSP27 in archived tumor samples representing various pathological characteristics of PHC were examined, and modification of HSP27 by SUMO2/3 was investigated. HSP27 were expressed abundantly in patients' tumor tissues, and found to be associated with pathological progression. Besides, HSP27 was also elevated significantly in liver cancer cell lines Huh7 and HepG2 compared with human hepatocyte cells L02. Furthermore, knockdown of HSP27 was found to be associated with the decreased proliferation and invasion ability in Huh7 and HepG2 cells. Immunofluorescence assay showed that HSP27 and SUMO2/3 were co-localized in the subcellular, and co-immunoprecipitation verified the interaction between HSP27 and SUMO2/3. Overexpression of SUMO2/3 upregulated the HSP27 protein level and promotes Huh7 and HepG2 cell proliferation and invasion, and vice versa when the SUMO2/3 was knockdown. Taken together, increased protein level of HSP27 through SUMO2/3-mediated SUMOylation plays crucial roles in the progression of PHC, and this finding may shed light on developing potential therapeutic targets for PHC.

Atropisomers of 2,2',3,3',6,6'-hexachlorobiphenyl (PCB 136) exhibit stereoselective effects on activation of nuclear receptors in vitro.

PubMed

Pěnčíková, Kateřina; Brenerová, Petra; Svržková, Lucie; Hrubá, Eva; Pálková, Lenka; Vondráček, Jan; Lehmler, Hans-Joachim; Machala, Miroslav

2017-11-09

PCB 136 is an environmentally relevant chiral PCB congener, which has been found in vivo to be present in form of rotational isomers (atropisomers). Its atropselective biotransformation or neurotoxic effects linked with sensitization of ryanodine receptor suggest that it might interact also with other intracellular receptors in a stereospecific manner. However, possible atropselective effects of PCB 136 on nuclear receptor transactivation remain unknown. Therefore, in this study, atropselective effects of PCB 136 on nuclear receptors controlling endocrine signaling and/or expression of xenobiotic and steroid hormone catabolism were investigated. PCB136 atropisomers were found to exert differential effects on estrogen receptor (ER) activation; (+)-PCB 136 was estrogenic, while (-)-PCB 136 was antiestrogenic. In contrast, inhibition of androgen receptor (AR) activity was not stereospecific. Both PCB136 stereoisomers induced the constitutive androgen receptor (CAR)-dependent gene expression; however, no significant stereospecificity of PCB 136 atropisomers was observed. PCB136 was a partial inducer of the pregnane X receptor (PXR)-dependent gene expression. Here, (-)-PCB 136 was a significantly more potent inducer of PXR activity than (+)-PCB 136. Taken together, the present results indicate that at least two nuclear receptors participating in endocrine regulation or metabolism, ER and PXR, could be regulated in an atropselective manner by chiral PCB 136. The enantioselective enrichment of PCB atropisomers in animal and human tissues may thus have significant consequences for endocrine-disrupting effects of chiral ortho-substituted PCB congeners.

A comprehensive data mining study shows that most nuclear receptors act as newly proposed homeostasis-associated molecular pattern receptors.

PubMed

Wang, Luqiao; Nanayakkara, Gayani; Yang, Qian; Tan, Hongmei; Drummer, Charles; Sun, Yu; Shao, Ying; Fu, Hangfei; Cueto, Ramon; Shan, Huimin; Bottiglieri, Teodoro; Li, Ya-Feng; Johnson, Candice; Yang, William Y; Yang, Fan; Xu, Yanjie; Xi, Hang; Liu, Weiqing; Yu, Jun; Choi, Eric T; Cheng, Xiaoshu; Wang, Hong; Yang, Xiaofeng

2017-10-24

Nuclear receptors (NRs) can regulate gene expression; therefore, they are classified as transcription factors. Despite the extensive research carried out on NRs, still several issues including (1) the expression profile of NRs in human tissues, (2) how the NR expression is modulated during atherosclerosis and metabolic diseases, and (3) the overview of the role of NRs in inflammatory conditions are not fully understood. To determine whether and how the expression of NRs are regulated in physiological/pathological conditions, we took an experimental database analysis to determine expression of all 48 known NRs in 21 human and 17 murine tissues as well as in pathological conditions. We made the following significant findings: (1) NRs are differentially expressed in tissues, which may be under regulation by oxygen sensors, angiogenesis pathway, stem cell master regulators, inflammasomes, and tissue hypo-/hypermethylation indexes; (2) NR sequence mutations are associated with increased risks for development of cancers and metabolic, cardiovascular, and autoimmune diseases; (3) NRs have less tendency to be upregulated than downregulated in cancers, and autoimmune and metabolic diseases, which may be regulated by inflammation pathways and mitochondrial energy enzymes; and (4) the innate immune sensor inflammasome/caspase-1 pathway regulates the expression of most NRs. Based on our findings, we propose a new paradigm that most nuclear receptors are anti-inflammatory homeostasis-associated molecular pattern receptors (HAMPRs). Our results have provided a novel insight on NRs as therapeutic targets in metabolic diseases, inflammations, and malignancies.

Heterodimers of Retinoic Acid Receptors and Thyroid Hormone Receptors Display Unique Combinatorial Regulatory Properties

PubMed Central

Lee, Sangho; Privalsky, Martin L.

2009-01-01

Nuclear receptors are ligand-regulated transcription factors that regulate key aspects of metazoan development, differentiation, and homeostasis. Nuclear receptors recognize target genes by binding to specific DNA recognition sequences, denoted hormone response elements (HREs). Many nuclear receptors can recognize HREs as either homodimers or heterodimers. Retinoid X receptors (RXRs), in particular, serve as important heterodimer partners for many other nuclear receptors, including thyroid hormone receptors (TRs), and RXR/TR heterodimers have been proposed to be the primary mediators of target gene regulation by T3 hormone. Here, we report that the retinoic acid receptors (RARs), a distinct class of nuclear receptors, are also efficient heterodimer partners for TRs. These RAR/TR heterodimers form with similar affinities as RXR/TR heterodimers on an assortment of consensus and natural HREs, and preferentially assemble with the RAR partner 5′ of the TR moiety. The corepressor and coactivator recruitment properties of these RAR/TR heterodimers and their transcriptional activities in vivo are distinct from those observed with the corresponding RXR heterodimers. Our studies indicate that RXRs are not unique in their ability to partner with TRs, and that RARs can also serve as robust heterodimer partners and combinatorial regulators of T3-modulated gene expression. PMID:15650024

RNA-induced silencing complex (RISC) Proteins PACT, TRBP, and Dicer are SRA binding nuclear receptor coregulators

PubMed Central

Redfern, Andrew D.; Colley, Shane M.; Beveridge, Dianne J.; Ikeda, Naoya; Epis, Michael R.; Li, Xia; Foulds, Charles E.; Stuart, Lisa M.; Barker, Andrew; Russell, Victoria J.; Ramsay, Kerry; Kobelke, Simon J.; Li, Xiaotao; Hatchell, Esme C.; Payne, Christine; Giles, Keith M.; Messineo, Adriana; Gatignol, Anne; Lanz, Rainer B.; O’Malley, Bert W.; Leedman, Peter J.

2013-01-01

The cytoplasmic RNA-induced silencing complex (RISC) contains dsRNA binding proteins, including protein kinase RNA activator (PACT), transactivation response RNA binding protein (TRBP), and Dicer, that process pre-microRNAs into mature microRNAs (miRNAs) that target specific mRNA species for regulation. There is increasing evidence for important functional interactions between the miRNA and nuclear receptor (NR) signaling networks, with recent data showing that estrogen, acting through the estrogen receptor, can modulate initial aspects of nuclear miRNA processing. Here, we show that the cytoplasmic RISC proteins PACT, TRBP, and Dicer are steroid receptor RNA activator (SRA) binding NR coregulators that target steroid-responsive promoters and regulate NR activity and downstream gene expression. Furthermore, each of the RISC proteins, together with Argonaute 2, associates with SRA and specific pre-microRNAs in both the nucleus and cytoplasm, providing evidence for links between NR-mediated transcription and some of the factors involved in miRNA processing. PMID:23550157

RNA-induced silencing complex (RISC) Proteins PACT, TRBP, and Dicer are SRA binding nuclear receptor coregulators.

PubMed

Redfern, Andrew D; Colley, Shane M; Beveridge, Dianne J; Ikeda, Naoya; Epis, Michael R; Li, Xia; Foulds, Charles E; Stuart, Lisa M; Barker, Andrew; Russell, Victoria J; Ramsay, Kerry; Kobelke, Simon J; Li, Xiaotao; Hatchell, Esme C; Payne, Christine; Giles, Keith M; Messineo, Adriana; Gatignol, Anne; Lanz, Rainer B; O'Malley, Bert W; Leedman, Peter J

2013-04-16

The cytoplasmic RNA-induced silencing complex (RISC) contains dsRNA binding proteins, including protein kinase RNA activator (PACT), transactivation response RNA binding protein (TRBP), and Dicer, that process pre-microRNAs into mature microRNAs (miRNAs) that target specific mRNA species for regulation. There is increasing evidence for important functional interactions between the miRNA and nuclear receptor (NR) signaling networks, with recent data showing that estrogen, acting through the estrogen receptor, can modulate initial aspects of nuclear miRNA processing. Here, we show that the cytoplasmic RISC proteins PACT, TRBP, and Dicer are steroid receptor RNA activator (SRA) binding NR coregulators that target steroid-responsive promoters and regulate NR activity and downstream gene expression. Furthermore, each of the RISC proteins, together with Argonaute 2, associates with SRA and specific pre-microRNAs in both the nucleus and cytoplasm, providing evidence for links between NR-mediated transcription and some of the factors involved in miRNA processing.

Nuclear Receptor TLX in Development and Diseases.

PubMed

Sun, Guoqiang; Cui, Qi; Shi, Yanhong

2017-01-01

The nuclear receptor TLX (NR2E1) is a transcription factor that is critical for neural development and adult neurogenesis through its actions in regulating neural stem cell proliferation, self-renewal, and fate determination. These roles are primarily executed by regulating TLX downstream target genes involved in myriad pathways such as cell cycle progression, RNA processing, angiogenesis, and senescence. Recent studies suggest that dysregulation of TLX pathways plays an important role in the pathogenesis of human neurological disorders and brain tumors. Here, we will highlight recent progress in the roles of TLX in brain development and adult neurogenesis, and the relevance of TLX to neurological diseases and brain tumors. We will also discuss the potential of TLX as a therapeutic target for these disorders. © 2017 Elsevier Inc. All rights reserved.

Brain nuclear receptors and body weight regulation

PubMed Central

O’Malley, Bert W.; Elmquist, Joel K.

2017-01-01

Neural pathways, especially those in the hypothalamus, integrate multiple nutritional, hormonal, and neural signals, resulting in the coordinated control of body weight balance and glucose homeostasis. Nuclear receptors (NRs) sense changing levels of nutrients and hormones, and therefore play essential roles in the regulation of energy homeostasis. Understanding the role and the underlying mechanisms of NRs in the context of energy balance control may facilitate the identification of novel targets to treat obesity. Notably, NRs are abundantly expressed in the brain, and emerging evidence indicates that a number of these brain NRs regulate multiple aspects of energy balance, including feeding, energy expenditure and physical activity. In this Review we summarize some of the recent literature regarding effects of brain NRs on body weight regulation and discuss mechanisms underlying these effects. PMID:28218618

Integrated in silico and in vivo approaches to investigate effects of BDE-99 mediated by the nuclear receptors on developing zebrafish.

PubMed

Zhang, Li; Jin, Yaru; Han, Zhihua; Liu, Hongling; Shi, Laihao; Hua, Xiaoxue; Doering, Jon A; Tang, Song; Giesy, John P; Yu, Hongxia

2018-03-01

One of the most abundant polybrominated diphenyl ethers (PBDEs) is 2,2',4,4',5-pentabromodiphenyl ether (BDE-99), which persists and potentially bioaccumulates in aquatic wildlife. Previous studies in mammals have shown that BDE-99 affects development and disrupts certain endocrine functions through signaling pathways mediated by nuclear receptors. However, fewer studies have investigated the potential of BDE-99 to interact with nuclear receptors in aquatic vertebrates such as fish. In the present study, interactions between BDE-99 and nuclear receptors were investigated by in silico and in vivo approaches. This PBDE was able to dock into the ligand-binding domain of zebrafish aryl hydrocarbon receptor 2 (AhR2) and pregnane X receptor (PXR). It had a significant effect on the transcriptional profiles of genes associated with AhR or PXR. Based on the developed cytoscape of all zebrafish genes, it was also inferred that AhR and PXR could interact via cross-talk. In addition, both the in silico and in vivo approaches found that BDE-99 affected peroxisome proliferator-activated receptor alpha (PPARα), glucocorticoid receptor, and thyroid receptor. Collectively, our results demonstrate for the first time detailed in silico evidence that BDE-99 can bind to and interact with zebrafish AhR and PXR. These findings can be used to elaborate the molecular mechanism of BDE-99 and guide more objective environmental risk assessments. Environ Toxicol Chem 2018;37:780-787. © 2017 SETAC. © 2017 SETAC.

Surface localization of the nuclear receptor CAR in influenza A virus-infected cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Takahashi, Tadanobu; Department of Biochemistry, School of Pharmaceutical Sciences, University of Shizuoka, CREST, JST, and COE Program in the 21st Century, Shizuoka 422-8526; Moriyama, Yusuke

Constitutive active/androstane receptor CAR is a member of the nuclear receptors which regulate transcription of xenobiotic metabolism enzymes. CAR is usually localized in the cytosol and nucleus. Here, we found that CAR was localized at the cell surface of influenza A virus (IAV)-infected cells. Additionally, we demonstrated that expression of a viral envelope glycoprotein, either hemagglutinin (HA) or neuraminidase (NA), but not viral nucleoprotein (NP), was responsible for this localization. This report is the first demonstration of CAR at the surface of tissue culture cells, and suggests that CAR may exert the IAV infection mechanism.

Identification of Modulators of the Nuclear Receptor ...

EPA Pesticide Factsheets

The nuclear receptor family member peroxisome proliferator-activated receptor α (PPARα) is activated by therapeutic hypolipidemic drugs and environmentally-relevant chemicals to regulate genes involved in lipid transport and catabolism. Chronic activation of PPARα in rodents increases in liver cancer incidence, whereas suppression of PPARα activity can lead to hepatocellular steatosis. Analytical approaches were developed to identify biosets (i.e., gene expression differences between two conditions) in a genomic database in which PPARα activity was altered. A gene expression signature of 131 PPARα-dependent genes was built using profiles from the livers of wild-type and PPARα-null mice after exposure to three structurally diverse PPARα activators (WY-14,643, fenofibrate and perfluorohexane sulfonate). A rank-based test (Running Fisher’s test (p-value ≤ 10-4)) was used to evaluate the similarity between the PPARα signature and a test set of 48 and 31 biosets positive or negative, respectively for PPARα activation; the test resulted in a balanced accuracy of 98%. The signature was used to identify factors that activate or suppress PPARα in an annotated mouse liver/primary hepatocyte gene expression database of ~1850 biosets. In addition to the expected activation of PPARα by fibrate drugs, di(2-ethylhexyl) phthalate, and perfluorinated compounds, PPARα was activated by benzofuran, galactosamine and TCDD and suppressed by hepatotoxins acetami

MTA family of coregulators in nuclear receptor biology and pathology

PubMed Central

Manavathi, Bramanandam; Singh, Kamini; Kumar, Rakesh

2007-01-01

Nuclear receptors (NRs) rely on coregulators (coactivators and corepressors) to modulate the transcription of target genes. By interacting with nucleosome remodeling complexes, NR coactivators potentiate transcription, whereas corepressors inhibit transcription of the target genes. Metastasis-associated proteins (MTA) represent an emerging family of novel NR coregulators. In general, MTA family members form independent nucleosome remodeling and deacetylation (NuRD) complexes and repress the transcription of different genes by recruiting histone deacetylases onto their target genes. However, MTA1 also acts as a coactivator in a promoter-context dependent manner. Recent findings that repression of estrogen receptor transactivation functions by MTA1, MTA1s, and MTA2 and regulation of MTA3 by estrogen signaling have indicated the significance of these proteins in NR signaling. Here, we highlight the action of MTA proteins on NR signaling and their roles in pathophysiological conditions. PMID:18174918

Plasticity of an Ultrafast Interaction between Nucleoporins and Nuclear Transport Receptors

PubMed Central

Milles, Sigrid; Mercadante, Davide; Aramburu, Iker Valle; Jensen, Malene Ringkjøbing; Banterle, Niccolò; Koehler, Christine; Tyagi, Swati; Clarke, Jane; Shammas, Sarah L.; Blackledge, Martin; Gräter, Frauke; Lemke, Edward A.

2015-01-01

Summary The mechanisms by which intrinsically disordered proteins engage in rapid and highly selective binding is a subject of considerable interest and represents a central paradigm to nuclear pore complex (NPC) function, where nuclear transport receptors (NTRs) move through the NPC by binding disordered phenylalanine-glycine-rich nucleoporins (FG-Nups). Combining single-molecule fluorescence, molecular simulations, and nuclear magnetic resonance, we show that a rapidly fluctuating FG-Nup populates an ensemble of conformations that are prone to bind NTRs with near diffusion-limited on rates, as shown by stopped-flow kinetic measurements. This is achieved using multiple, minimalistic, low-affinity binding motifs that are in rapid exchange when engaging with the NTR, allowing the FG-Nup to maintain an unexpectedly high plasticity in its bound state. We propose that these exceptional physical characteristics enable a rapid and specific transport mechanism in the physiological context, a notion supported by single molecule in-cell assays on intact NPCs. PMID:26456112

Nuclear Membranes ETB Receptors Mediate ET-1-induced Increase of Nuclear Calcium in Human Left Ventricular Endocardial Endothelial Cells.

PubMed

Jules, Farah; Avedanian, Levon; Al-Khoury, Johny; Keita, Ramatoulaye; Normand, Alexandre; Bkaily, Ghassan; Jacques, Danielle

2015-07-01

In fetal human left ventricular endocardial endothelial cells (EECLs), both plasma membrane (PM) ET(A)R and ET(B)R were reported to mediate ET-1-induced increase of intracellular calcium [Ca](i); however, this effect was mediated by ET(A)R in right EECs (EECRs). In this study, we verified whether, as for the PM, nuclear membranes (NMs) ET-1 receptors activation in EECLs and EECRs induce an increase of nuclear calcium ([Ca](n)) and if this effect is mediated through the same receptor type as in PM. Using a plasmalemma-perforated technique and 3D confocal microscopy, our results showed that, as in PM intact cells, superfusion of nuclei of both cell types with cytosolic ET-1 induced a concentration-dependent sustained increase of [Ca](n). In EECRs, the ET(A)R antagonist prevented the effect of ET-1 on [Ca](n) without affecting EECLs. However, in both cell types, the effect of cytosolic ET-1 on [Ca](n) was prevented by the ETBR antagonist. In conclusion, both NMs' ET(A)R and ET(B)R mediated the effect of cytosolic ET-1 on [Ca](n) in EECRs. In contrast, only NMs' ET(B)R activation mediated the effect of cytosolic ET-1 in EECLs. Hence, the type of NMs' receptors mediating the effect of ET-1 on [Ca](n) are different from those of PM mediating the increase in [Ca](i).

LASSO-ing Potential Nuclear Receptor Agonists and Antagonists: A New Computational Method for Database Screening

EPA Science Inventory

Nuclear receptors (NRs) are important biological macromolecular transcription factors that are implicated in multiple biological pathways and may interact with other xenobiotics that are endocrine disruptors present in the environment. Examples of important NRs include the androg...

Nuclear receptor TLX prevents retinal dystrophy and recruits the corepressor atrophin1

PubMed Central

Zhang, Chun-Li; Zou, Yuhua; Yu, Ruth T.; Gage, Fred H.; Evans, Ronald M.

2006-01-01

During mammalian embryogenesis, precise coordination of progenitor cell proliferation and differentiation is essential for proper organ size and function. The involvement of TLX (NR2E1), an orphan nuclear receptor, has been implicated in ocular development, as Tlx−/− mice exhibit visual impairment. Using genetic and biochemical approaches, we show that TLX modulates retinal progenitor cell proliferation and cell cycle re-entry by directly regulating the expression of Pten and its target cyclin D1. Additionally, TLX finely tunes the progenitor differentiation program by modulating the phospholipase C and mitogen-activated protein kinase (MAPK) pathways and the expression of an array of cell type-specific transcriptional regulators. Consequently, Tlx−/− mice have a dramatic reduction in retina thickness and enhanced generation of S-cones, and develop severe early onset retinal dystrophy. Furthermore, TLX interacts with atrophin1 (Atn1), a corepressor that is involved in human neurodegenerative dentatorubral-pallidoluysian atrophy (DRPLA) and that is essential for development of multiple tissues. Together, these results reveal a molecular strategy by which an orphan nuclear receptor can precisely orchestrate tissue-specific proliferation and differentiation programs to prevent retinal malformation and degeneration. PMID:16702404

Nuclear receptor TLX prevents retinal dystrophy and recruits the corepressor atrophin1.

PubMed

Zhang, Chun-Li; Zou, Yuhua; Yu, Ruth T; Gage, Fred H; Evans, Ronald M

2006-05-15

During mammalian embryogenesis, precise coordination of progenitor cell proliferation and differentiation is essential for proper organ size and function. The involvement of TLX (NR2E1), an orphan nuclear receptor, has been implicated in ocular development, as Tlx-/- mice exhibit visual impairment. Using genetic and biochemical approaches, we show that TLX modulates retinal progenitor cell proliferation and cell cycle re-entry by directly regulating the expression of Pten and its target cyclin D1. Additionally, TLX finely tunes the progenitor differentiation program by modulating the phospholipase C and mitogen-activated protein kinase (MAPK) pathways and the expression of an array of cell type-specific transcriptional regulators. Consequently, Tlx-/- mice have a dramatic reduction in retina thickness and enhanced generation of S-cones, and develop severe early onset retinal dystrophy. Furthermore, TLX interacts with atrophin1 (Atn1), a corepressor that is involved in human neurodegenerative dentatorubral-pallidoluysian atrophy (DRPLA) and that is essential for development of multiple tissues. Together, these results reveal a molecular strategy by which an orphan nuclear receptor can precisely orchestrate tissue-specific proliferation and differentiation programs to prevent retinal malformation and degeneration.

Investigating the Mechanisms Underlying Neuronal Death in Ischemia Using In Vitro Oxygen-Glucose Deprivation: Potential Involvement of Protein SUMOylation

PubMed Central

CIMAROSTI, HELENA; HENLEY, JEREMY M.

2012-01-01

It is well established that brain ischemia can cause neuronal death via different signaling cascades. The relative importance and interrelationships between these pathways, however, remain poorly understood. Here is presented an overview of studies using oxygen-glucose deprivation of organotypic hippocampal slice cultures to investigate the molecular mechanisms involved in ischemia. The culturing techniques, setup of the oxygen-glucose deprivation model, and analytical tools are reviewed. The authors focus on SUMOylation, a posttranslational protein modification that has recently been implicated in ischemia from whole animal studies as an example of how these powerful tools can be applied and could be of interest to investigate the molecular pathways underlying ischemic cell death. PMID:19029060

The oxidative stress mediator 4-hydroxynonenal is an intracellular agonist of the nuclear receptor peroxisome proliferator-activated receptor-β/δ (PPARβ/δ)

PubMed Central

Coleman, Jeffrey D.; Prabhu, K. Sandeep; Thompson, Jerry T.; Reddy, P. Sreenivasula; Peters, Jeffrey M.; Peterson, Blake R.; Reddy, C. Channa; Vanden Heuvel, John P.

2007-01-01

Liver insufficiency and damage is a major cause of death and disease worldwide and may result from exposure to environmental toxicants, specific combinations or dosages of pharmaceuticals and microbial metabolites. The generation of reactive intermediates, in particular 4-hydroxynonenal (4-HNE), is a common event in liver damage caused by a variety of hepatotoxic drugs and solvents. The peroxisome proliferator-activated receptors (PPARs) are nuclear receptors that are involved in the transcriptional regulation of lipid metabolism as well as other biological functions. Importantly, we have observed that the PPARβ/δ−/− mouse is more susceptible to chemically-induced hepatotoxicity than its wildtype counterpart, and our objective in this study was to elucidate the mechanism(s) by which PPARβ/δ confers protection to hepatocytes. We hypothesized that PPARβ/δ plays a protective role by responding to toxic lipids and altering gene expression accordingly. In support, oxidized-VLDL and constituents including 13-S-hydroxyoctadeca-dienoic acid (13(S)-HODE) and 4-HNE are PPARβ/δ ligands. A structure-activity relationship was established where 4-HNE and 4-hydroperoxynonenal (4-HpNE) enhanced the activity of the PPARβ/δ subtype while 4-hyroxy-hexenal (4-HHE), 4-oxo-2-Nonenal (4-ONE), and trans-4,5-epoxy-2(E)-decenal did not activate this receptor. Increasing PPARβ/δ activity with a synthetic agonist decreased sensitivity of hepatocytes to 4-HNE and other toxic agents, whereas inhibition of this receptor had the opposite result. Gene expression microarray analysis identified several important PPARβ/δ-regulated detoxification enzymes involved in 4-HNE metabolism that are regulated at the transcript level. This research established 4-HNE as an endogenous modulator of PPARβ/δ activity and raises the possibility that agonists of this nuclear receptor may be utilized to prevent or treat liver disease associated with oxidative damage. PMID:17382197

Abnormal XPD-induced nuclear receptor transactivation in DNA repair disorders: trichothiodystrophy and xeroderma pigmentosum.

PubMed

Zhou, Xiaolong; Khan, Sikandar G; Tamura, Deborah; Ueda, Takahiro; Boyle, Jennifer; Compe, Emmanuel; Egly, Jean-Marc; DiGiovanna, John J; Kraemer, Kenneth H

2013-08-01

XPD (ERCC2) is a DNA helicase involved in nucleotide excision repair and in transcription as a structural bridge tying the transcription factor IIH (TFIIH) core with the cdk-activating kinase complex, which phosphorylates nuclear receptors. Mutations in XPD are associated with several different phenotypes, including trichothiodystrophy (TTD), with sulfur-deficient brittle hair, bone defects, and developmental abnormalities without skin cancer, xeroderma pigmentosum (XP), with pigmentary abnormalities and increased skin cancer, or XP/TTD with combined features, including skin cancer. We describe the varied clinical features and mutations in nine patients examined at the National Institutes of Health who were compound heterozygotes for XPD mutations but had different clinical phenotypes: four TTD, three XP, and two combined XP/TTD. We studied TFIIH-dependent transactivation by nuclear receptor for vitamin D (VDR) and thyroid in cells from these patients. The vitamin D stimulation ratio of CYP24 and osteopontin was associated with specific pairs of mutations (reduced in 5, elevated in 1) but not correlated with distinct clinical phenotypes. Thyroid receptor stimulation ratio for KLF9 was not significantly different from normal. XPD mutations frequently were associated with abnormal VDR stimulation in compound heterozygote patients with TTD, XP, or XP/TTD.

Abnormal XPD-induced nuclear receptor transactivation in DNA repair disorders: trichothiodystrophy and xeroderma pigmentosum

PubMed Central

Zhou, Xiaolong; Khan, Sikandar G; Tamura, Deborah; Ueda, Takahiro; Boyle, Jennifer; Compe, Emmanuel; Egly, Jean-Marc; DiGiovanna, John J; Kraemer, Kenneth H

2013-01-01

XPD (ERCC2) is a DNA helicase involved in nucleotide excision repair and in transcription as a structural bridge tying the transcription factor IIH (TFIIH) core with the cdk-activating kinase complex, which phosphorylates nuclear receptors. Mutations in XPD are associated with several different phenotypes, including trichothiodystrophy (TTD), with sulfur-deficient brittle hair, bone defects, and developmental abnormalities without skin cancer, xeroderma pigmentosum (XP), with pigmentary abnormalities and increased skin cancer, or XP/TTD with combined features, including skin cancer. We describe the varied clinical features and mutations in nine patients examined at the National Institutes of Health who were compound heterozygotes for XPD mutations but had different clinical phenotypes: four TTD, three XP, and two combined XP/TTD. We studied TFIIH-dependent transactivation by nuclear receptor for vitamin D (VDR) and thyroid in cells from these patients. The vitamin D stimulation ratio of CYP24 and osteopontin was associated with specific pairs of mutations (reduced in 5, elevated in 1) but not correlated with distinct clinical phenotypes. Thyroid receptor stimulation ratio for KLF9 was not significantly different from normal. XPD mutations frequently were associated with abnormal VDR stimulation in compound heterozygote patients with TTD, XP, or XP/TTD. PMID:23232694

Development of an image analysis screen for estrogen receptor alpha (ERα) ligands through measurement of nuclear translocation dynamics.

PubMed

Dull, Angie; Goncharova, Ekaterina; Hager, Gordon; McMahon, James B

2010-11-01

We have developed a robust high-content assay to screen for novel estrogen receptor alpha (ERα) agonists and antagonists by quantitation of cytoplasmic to nuclear translocation of an estrogen receptor chimera in 384-well plates. The screen utilizes a green fluorescent protein tagged-glucocorticoid/estrogen receptor (GFP-GRER) chimera which consisted of the N-terminus of the glucocorticoid receptor fused to the human ER ligand binding domain. The GFP-GRER exhibited cytoplasmic localization in the absence of ERα ligands, and translocated to the nucleus in response to stimulation with ERα agonists or antagonists. The BD Pathway 435 imaging system was used for image acquisition, analysis of translocation dynamics, and cytotoxicity measurements. The assay was validated with known ERα agonists and antagonists, and the Library of Pharmacologically Active Compounds (LOPAC 1280). Additionally, screening of crude natural product extracts demonstrated the robustness of the assay, and the ability to quantitate the effects of toxicity on nuclear translocation dynamics. The GFP-GRER nuclear translocation assay was very robust, with z' values >0.7, CVs <5%, and has been validated with known ER ligands, and inclusion of cytotoxicity filters will facilitate screening of natural product extracts. This assay has been developed for future primary screening of synthetic, pure natural products, and natural product extracts libraries available at the National Cancer Institute at Frederick. Copyright © 2010 Elsevier Ltd. All rights reserved.

Transcription control and neuronal differentiation by agents that activate the LXR nuclear receptor family.

PubMed

Schmidt, A; Vogel, R; Holloway, M K; Rutledge, S J; Friedman, O; Yang, Z; Rodan, G A; Friedman, E

1999-09-10

LXR and PPAR receptors belong to the nuclear receptor superfamily of transcriptional activating factors. Using ligand-dependent transcription assays, we found that 5-tetradecyloxy-2-furancarboxylic acid (TOFA) transactivates chimeric receptors composed of the glucocorticoid receptor DNA binding domain and the ligand binding regions of PPARalpha, PPARbeta (NUC-1) and LXRbeta (NER) receptors. In the same assays, ligands for PPARs (oleic acid, WY-14643 and L-631,033) and LXRs (hydroxycholesterols) maintain their respective receptor selectivity. TOFA and hydroxycholesterols also stimulate transcription from a minimal fibrinogen promoter that is under the control of AP-1 or NF-kappaB transcription factor binding sites. In addition to their effects on transcription, these LXRbeta activators induce neuronal differentiation in rat pheochromocytoma cells. TOFA and the natural LXR agonist, 22 (R)-hydroxycholesterol, stimulate neurite outgrowth in 55 and 28% of cells, respectively. No neurite outgrowth was induced by the related 22(S)-hydroxycholesterol, which does not activate the LXR family. These results suggest that the hydroxycholesterol signaling pathway has a complex effect on transcription that mediates the activity of TOFA and hydroxycholesterol on neuronal differentiation in pheochromocytoma cells.

Toll-like receptor 4 promotes proliferation and apoptosis resistance in human papillomavirus-related cervical cancer cells through the Toll-like receptor 4/nuclear factor-κB pathway.

PubMed

Jiang, Ninghong; Xie, Feng; Guo, Qisang; Li, Ming-Qing; Xiao, Jingjing; Sui, Long

2017-06-01

Toll-like receptor 4 is overexpressed in various tumors, including cervical carcinoma. However, the role of Toll-like receptor 4 in cervical cancer remains controversial, and the underlying mechanisms are largely elusive. Therefore, Toll-like receptor 4 in cervical cancer and related mechanisms were investigated in this study. Quantitative reverse transcription polymerase chain reaction and western blot analyses were used to detect messenger RNA and protein levels in HeLa, Caski, and C33A cells with different treatments. Proliferation was quantified using Cell Counting Kit-8. Cell cycle distribution and apoptosis were assessed by flow cytometry. Higher levels of Toll-like receptor 4 expression were found in human papillomavirus-positive cells compared to human papillomavirus-negative cells. Proliferation of HeLa and Caski cells was promoted in lipopolysaccharide-stimulated groups but suppressed in short hairpin RNA-transfected groups. Apoptosis rates were lower in lipopolysaccharide-stimulated groups relative to short hairpin RNA-transfected groups. In addition, G2-phase distribution was enhanced when Toll-like receptor 4 was downregulated. Moreover, the pNF-κBp65 level was positively correlated with the Toll-like receptor 4 level in HeLa and Caski cells, though when an nuclear factor-κB inhibitor was applied to lipopolysaccharide-stimulated groups, the patterns of proliferation and apoptosis were opposite to those of the lipopolysaccharide-stimulated groups without inhibitor treatment. In conclusion, these data suggest that Toll-like receptor 4 promotes proliferation and apoptosis resistance in human papillomavirus-related cervical cancer cells at least in part through the Toll-like receptor 4/nuclear factor-κB pathway, which may be correlated with the occurrence and development of cervical carcinoma.

Orphan nuclear receptor TR3 acts in autophagic cell death via mitochondrial signaling pathway.

PubMed

Wang, Wei-jia; Wang, Yuan; Chen, Hang-zi; Xing, Yong-zhen; Li, Feng-wei; Zhang, Qian; Zhou, Bo; Zhang, Hong-kui; Zhang, Jie; Bian, Xue-li; Li, Li; Liu, Yuan; Zhao, Bi-xing; Chen, Yan; Wu, Rong; Li, An-zhong; Yao, Lu-ming; Chen, Ping; Zhang, Yi; Tian, Xu-yang; Beermann, Friedrich; Wu, Mian; Han, Jiahuai; Huang, Pei-qiang; Lin, Tianwei; Wu, Qiao

2014-02-01

Autophagy is linked to cell death, yet the associated mechanisms are largely undercharacterized. We discovered that melanoma, which is generally resistant to drug-induced apoptosis, can undergo autophagic cell death with the participation of orphan nuclear receptor TR3. A sequence of molecular events leading to cellular demise is launched by a specific chemical compound, 1-(3,4,5-trihydroxyphenyl)nonan-1-one, newly acquired from screening a library of TR3-targeting compounds. The autophagic cascade comprises TR3 translocation to mitochondria through interaction with the mitochondrial outer membrane protein Nix, crossing into the mitochondrial inner membrane through Tom40 and Tom70 channel proteins, dissipation of mitochondrial membrane potential by the permeability transition pore complex ANT1-VDAC1 and induction of autophagy. This process leads to excessive mitochondria clearance and irreversible cell death. It implicates a new approach to melanoma therapy through activation of a mitochondrial signaling pathway that integrates a nuclear receptor with autophagy for cell death.

Nuclear receptors and metabolism: from feast to famine.

PubMed

Hong, Suk-Hyun; Ahmadian, Maryam; Yu, Ruth T; Atkins, Annette R; Downes, Michael; Evans, Ronald M

2014-05-01

The ability to adapt to cycles of feast and famine is critical for survival. Communication between multiple metabolic organs must be integrated to properly metabolise nutrients. By controlling networks of genes in major metabolic organs, nuclear hormone receptors (NHRs) play central roles in regulating metabolism in a tissue-specific manner. NHRs also establish daily rhythmicity by controlling the expression of core clock genes both centrally and peripherally. Recent findings show that many of the metabolic effects of NHRs are mediated through certain members of the fibroblast growth factor (FGF) family. This review focuses on the roles of NHRs in critical metabolic organs, including adipose tissue, liver and muscle, during the fed and fasted states, as well as their roles in circadian metabolism and downstream regulation of FGFs.

Identification of SR1078, a synthetic agonist for the orphan nuclear receptors RORα and RORγ.

PubMed

Wang, Yongjun; Kumar, Naresh; Nuhant, Philippe; Cameron, Michael D; Istrate, Monica A; Roush, William R; Griffin, Patrick R; Burris, Thomas P

2010-11-19

The retinoic acid receptor-related receptors (RORs) are members of the nuclear receptor (NR) superfamily of transcription factors. Several NRs are still characterized as orphan receptors because ligands have not yet been identified for these proteins. Here, we describe the identification of a synthetic RORα/RORγ ligand, SR1078. SR1078 modulates the conformation of RORγ in a biochemical assay and activates RORα and RORγ driven transcription. Furthermore, SR1078 stimulates expression of endogenous ROR target genes in HepG2 cells that express both RORα and RORγ. Pharmacokinetic studies indicate that SR1078 displays reasonable exposure following injection into mice, and consistent with SR1078 functioning as a RORα/RORγ agonist, expression of two ROR target genes, glucose-6-phosphatase and fibroblast growth factor 21, were stimulated in the liver. Thus, we have identified the first synthetic RORα/γ agonist, and this compound can be utilized as a chemical tool to probe the function of these receptors both in vitro and in vivo.

Identification of a Synthetic Agonist for the Orphan Nuclear Receptors RORα and RORγ, SR1078

PubMed Central

Wang, Yongjun; Kumar, Naresh; Nuhant, Philippe; Cameron, Michael D.; Istrate, Monica A.; Roush, William R.; Griffin, Patrick R.; Burris, Thomas P.

2010-01-01

The retinoic acid receptor-related receptors (RORs) are members of the nuclear receptor (NR) superfamily of transcription factors. Several NRs are still characterized as orphan receptors since ligands have not yet been identified for these proteins. Here, we describe the identification of a synthetic RORα/RORγ ligand, SR1078. SR1078 modulates the conformation of RORγ in a biochemical assay and activates RORα and RORγ driven transcription. Furthermore, SR1078 stimulates expression of endogenous ROR target genes in HepG2 cells that express both RORα and RORγ. Pharmacokinetic studies indicate that SR1078 displays reasonable exposure following injection into mice and consistent with SR1078 functioning as a RORα/RORγ agonist, expression of two ROR target genes, glucose-6-phosphatase and fibroblast growth factor 21, were stimulated in the liver. Thus, we have identified the first synthetic RORα/γ agonist and this compound can be utilized as a chemical tool to probe the function of these receptors both in vitro and in vivo. PMID:20735016

Controlling Androgen receptor nuclear localization by dendrimer conjugates

NASA Astrophysics Data System (ADS)

Wang, Haoyu

Androgen Receptor (AR) antagonists, such as bicalutamide and flutamide have been used widely in the treatment of prostate cancer. Although initial treatment is effective, prostate cancer cells often acquire antiandrogen resistance with prolonged treatment. AR over-expression and AR mutations contribute to the development of antiandrogen resistant cancer. Second generation antiandrogens such as enzalutamide are more effective and show reduced AR nuclear localization. In this study, derivatives of PAN52, a small molecule antiandrogen previously developed in our lab, were conjugated to the surface of generation 4 and generation 6 PAMAM dendrimers to obtain antiandrogen PAMAM dendrimer conjugates (APDC). APDCs readily enter cells and associate with AR in the cytoplasm. Due to their large size and positive charge, they can not enter the nucleus, thus retaining AR in the cytoplasm. In addition, APDCs are effective in decreasing AR mediated transcription and cell proliferation. APDC is the first AR antagonists that inhibit DHT-induced nuclear localization of AR. By inhibiting AR nuclear localization, APDC represents a new class of antiandrogens that offer an alternative approach to addressing antiandrogen-resistant prostate cancer. Lysine post-translational modification of AR Nuclear Localization Sequence (NLS) has great impact on AR cellular localization. It is of interest to understand which modifications modulate AR translocation into the nucleus. In this study, we prepared dendrimer-based acetyltransferase mimetic (DATM), DATM is able to catalytically acetylate AR in CWR22Rv1 cells, which will be a useful tool for studying AR modification effect on AR cellular localization. Derivatives of DATM, which transfer other chemical groups to AR, can be prepared similarly, and with more dendrimer based AR modification tools prepared in future, we will be able to understand and control AR cellular localization through AR modification.

PML/RARA oxidation and arsenic binding initiate the antileukemia response of As2O3.

PubMed

Jeanne, Marion; Lallemand-Breitenbach, Valérie; Ferhi, Omar; Koken, Marcel; Le Bras, Morgane; Duffort, Stéphanie; Peres, Laurent; Berthier, Caroline; Soilihi, Hassane; Raught, Brian; de Thé, Hugues

2010-07-13

As(2)O(3) cures acute promyelocytic leukemia (APL) by initiating PML/RARA oncoprotein degradation, through sumoylation of its PML moiety. However, how As(2)O(3) initiates PML sumoylation has remained largely unexplained. As(2)O(3) binds vicinal cysteines and increases reactive oxygen species (ROS) production. We demonstrate that upon As(2)O(3) exposure, PML undergoes ROS-initiated intermolecular disulfide formation and binds arsenic directly. Disulfide-linked PML or PML/RARA multimers form nuclear matrix-associated nuclear bodies (NBs), become sumoylated and are degraded. Hematopoietic progenitors transformed by an As(2)O(3)-binding PML/RARA mutant exhibit defective As(2)O(3) response. Conversely, nonarsenical oxidants elicit PML/RARA multimerization, NB-association, degradation, and leukemia response in vivo, but do not affect PLZF/RARA-driven APLs. Thus, PML oxidation regulates NB-biogenesis, while oxidation-enforced PML/RARA multimerization and direct arsenic-binding cooperate to enforce APL's exquisite As(2)O(3) sensitivity. Copyright (c) 2010 Elsevier Inc. All rights reserved.

Plasticity of an ultrafast interaction between nucleoporins and nuclear transport receptors.

PubMed

Milles, Sigrid; Mercadante, Davide; Aramburu, Iker Valle; Jensen, Malene Ringkjøbing; Banterle, Niccolò; Koehler, Christine; Tyagi, Swati; Clarke, Jane; Shammas, Sarah L; Blackledge, Martin; Gräter, Frauke; Lemke, Edward A

2015-10-22

The mechanisms by which intrinsically disordered proteins engage in rapid and highly selective binding is a subject of considerable interest and represents a central paradigm to nuclear pore complex (NPC) function, where nuclear transport receptors (NTRs) move through the NPC by binding disordered phenylalanine-glycine-rich nucleoporins (FG-Nups). Combining single-molecule fluorescence, molecular simulations, and nuclear magnetic resonance, we show that a rapidly fluctuating FG-Nup populates an ensemble of conformations that are prone to bind NTRs with near diffusion-limited on rates, as shown by stopped-flow kinetic measurements. This is achieved using multiple, minimalistic, low-affinity binding motifs that are in rapid exchange when engaging with the NTR, allowing the FG-Nup to maintain an unexpectedly high plasticity in its bound state. We propose that these exceptional physical characteristics enable a rapid and specific transport mechanism in the physiological context, a notion supported by single molecule in-cell assays on intact NPCs. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.

Depletion of UBC9 Causes Nuclear Defects during the Vegetative and Sexual Life Cycles in Tetrahymena thermophila.

PubMed

Yang, Qianyi; Nasir, Amjad M; Coyne, Robert S; Forney, James D

2015-12-01

Ubc9p is the sole E2-conjugating enzyme for SUMOylation, and its proper function is required for regulating key nuclear events such as transcription, DNA repair, and mitosis. In Tetrahymena thermophila, the genome is separated into a diploid germ line micronucleus (MIC) that divides by mitosis and a polyploid somatic macronucleus (MAC) that divides amitotically. This unusual nuclear organization provides novel opportunities for the study of SUMOylation and Ubc9p function. We identified the UBC9 gene and demonstrated that its complete deletion from both MIC and MAC genomes is lethal. Rescue of the lethal phenotype with a GFP-UBC9 fusion gene driven by a metallothionein promoter generated a cell line with CdCl2-dependent expression of green fluorescent protein (GFP)-Ubc9p. Depletion of Ubc9p in vegetative cells resulted in the loss of MICs, but MACs continued to divide. In contrast, expression of catalytically inactive Ubc9p resulted in the accumulation of multiple MICs. Critical roles for Ubc9p were also identified during the sexual life cycle of Tetrahymena. Cell lines that were depleted for Ubc9p did not form mating pairs and therefore could not complete any of the subsequent stages of conjugation, including meiosis and macronuclear development. Mating between cells expressing catalytically inactive Ubc9p resulted in arrest during macronuclear development, consistent with our observation that Ubc9p accumulates in the developing macronucleus. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Clearance of PML/RARA-bound promoters suffice to initiate APL differentiation.

PubMed

Vitaliano-Prunier, Adeline; Halftermeyer, Juliane; Ablain, Julien; de Reynies, Aurélien; Peres, Laurent; Le Bras, Morgane; Metzger, Daniel; de Thé, Hugues

2014-12-11

PML/RARA, a potent transcriptional inhibitor of nuclear receptor signaling, represses myeloid differentiation genes and drives acute promyelocytic leukemia (APL). Association of the retinoid X receptor-α (RXRA) coreceptor to PML/RARA is required for transformation, with RXRA promoting its efficient DNA binding. APL is exquisitely sensitive to retinoic acid (RA) and arsenic trioxide (arsenic), which both trigger cell differentiation in vivo. Whereas RA elicits transcriptional activation of PML/RARA targets, how arsenic triggers differentiation remains unclear. Here we demonstrate that extinction of PML/RARA triggers terminal differentiation in vivo. Similarly, ablation of retinoid X receptors loosens PML/RARA DNA binding, inducing terminal differentiation of APL cells ex vivo or in vivo. RXRA sumoylation directly contributes to PML/RARA-dependent transformation ex vivo, presumably by enhancing transcriptional repression. Thus, APL differentiation is a default program triggered by clearance of PML/RARA-bound promoters, rather than obligatory active transcriptional activation, explaining how arsenic elicits APL maturation through PML/RARA degradation. © 2014 by The American Society of Hematology.

Trichoplax adhaerens reveals a network of nuclear receptors sensitive to 9-cis-retinoic acid at the base of metazoan evolution

PubMed Central

Novotný, Jan Philipp; Chughtai, Ahmed Ali; Kostrouchová, Markéta; Kostrouchová, Veronika; Kostrouch, David; Kaššák, Filip; Kaňa, Radek; Schierwater, Bernd; Kostrouchová, Marta

2017-01-01

Trichoplax adhaerens, the only known species of Placozoa is likely to be closely related to an early metazoan that preceded branching of Cnidaria and Bilateria. This animal species is surprisingly well adapted to free life in the World Ocean inhabiting tidal costal zones of oceans and seas with warm to moderate temperatures and shallow waters. The genome of T. adhaerens (sp. Grell) includes four nuclear receptors, namely orthologue of RXR (NR2B), HNF4 (NR2A), COUP-TF (NR2F) and ERR (NR3B) that show a high degree of similarity with human orthologues. In the case of RXR, the sequence identity to human RXR alpha reaches 81% in the DNA binding domain and 70% in the ligand binding domain. We show that T. adhaerens RXR (TaRXR) binds 9-cis retinoic acid (9-cis-RA) with high affinity, as well as high specificity and that exposure of T. adhaerens to 9-cis-RA regulates the expression of the putative T. adhaerens orthologue of vertebrate L-malate-NADP+ oxidoreductase (EC 1.1.1.40) which in vertebrates is regulated by a heterodimer of RXR and thyroid hormone receptor. Treatment by 9-cis-RA alters the relative expression profile of T. adhaerens nuclear receptors, suggesting the existence of natural ligands. Keeping with this, algal food composition has a profound effect on T. adhaerens growth and appearance. We show that nanomolar concentrations of 9-cis-RA interfere with T. adhaerens growth response to specific algal food and causes growth arrest. Our results uncover an endocrine-like network of nuclear receptors sensitive to 9-cis-RA in T. adhaerens and support the existence of a ligand-sensitive network of nuclear receptors at the base of metazoan evolution. PMID:28975052

Structure of metabotropic glutamate receptor C-terminal domains in contact with interacting proteins.

PubMed

Enz, Ralf

2012-01-01

Metabotropic glutamate receptors (mGluRs) regulate intracellular signal pathways that control several physiological tasks, including neuronal excitability, learning, and memory. This is achieved by the formation of synaptic signal complexes, in which mGluRs assemble with functionally related proteins such as enzymes, scaffolds, and cytoskeletal anchor proteins. Thus, mGluR associated proteins actively participate in the regulation of glutamatergic neurotransmission. Importantly, dysfunction of mGluRs and interacting proteins may lead to impaired signal transduction and finally result in neurological disorders, e.g., night blindness, addiction, epilepsy, schizophrenia, autism spectrum disorders and Parkinson's disease. In contrast to solved crystal structures of extracellular N-terminal domains of some mGluR types, only a few studies analyzed the conformation of intracellular receptor domains. Intracellular C-termini of most mGluR types are subject to alternative splicing and can be further modified by phosphorylation and SUMOylation. In this way, diverse interaction sites for intracellular proteins that bind to and regulate the glutamate receptors are generated. Indeed, most of the known mGluR binding partners interact with the receptors' C-terminal domains. Within the last years, different laboratories analyzed the structure of these domains and described the geometry of the contact surface between mGluR C-termini and interacting proteins. Here, I will review recent progress in the structure characterization of mGluR C-termini and provide an up-to-date summary of the geometry of these domains in contact with binding partners.

Orphan nuclear receptor oestrogen-related receptor γ (ERRγ) plays a key role in hepatic cannabinoid receptor type 1-mediated induction of CYP7A1 gene expression

PubMed Central

Zhang, Yaochen; Kim, Don-Kyu; Lee, Ji-Min; Park, Seung Bum; Jeong, Won-IL; Kim, Seong Heon; Lee, In-Kyu; Lee, Chul-Ho; Chiang, John Y.L.; Choi, Hueng-Sik

2017-01-01

Bile acids are primarily synthesized from cholesterol in the liver and have important roles in dietary lipid absorption and cholesterol homoeostasis. Detailed roles of the orphan nuclear receptors regulating cholesterol 7α-hydroxylase (CYP7A1), the rate-limiting enzyme in bile acid synthesis, have not yet been fully elucidated. In the present study, we report that oestrogen-related receptor γ (ERRγ) is a novel transcriptional regulator of CYP7A1 expression. Activation of cannabinoid receptor type 1 (CB1 receptor) signalling induced ERRγ-mediated transcription of the CYP7A1 gene. Overexpression of ERRγ increased CYP7A1 expression in vitro and in vivo, whereas knockdown of ERRγ attenuated CYP7A1 expression. Deletion analysis of the CYP7A1 gene promoter and a ChIP assay revealed an ERRγ -binding site on the CYP7A1 gene promoter. Small heterodimer partner (SHP) inhibited the transcriptional activity of ERRγ and thus regulated CYP7A1 expression. Overexpression of ERRγ led to increased bile acid levels, whereas an inverse agonist of ERRγ, GSK5182, reduced CYP7A1 expression and bile acid synthesis. Finally, GSK5182 significantly reduced hepatic CB1 receptor-mediated induction of CYP7A1 expression and bile acid synthesis in alcohol-treated mice. These results provide the molecular mechanism linking ERRγ and bile acid metabolism. PMID:26348907

Application of an in silico liver model to determine nuclear receptor mediated pathways in liver cancer

EPA Science Inventory

Nuclear receptors (NRs) are ligand-activated transcription factors that control diverse cellular processes. Chronic stimulation of some NRs in rodents can result in increased incidence of liver tumors. These are generally thought to develop through a non-genotoxic mechanism with...

The export receptor Crm1 forms a dimer to promote nuclear export of HIV RNA.

PubMed

Booth, David S; Cheng, Yifan; Frankel, Alan D

2014-12-08

The HIV Rev protein routes viral RNAs containing the Rev Response Element (RRE) through the Crm1 nuclear export pathway to the cytoplasm where viral proteins are expressed and genomic RNA is delivered to assembling virions. The RRE assembles a Rev oligomer that displays nuclear export sequences (NESs) for recognition by the Crm1-Ran(GTP) nuclear receptor complex. Here we provide the first view of an assembled HIV-host nuclear export complex using single-particle electron microscopy. Unexpectedly, Crm1 forms a dimer with an extensive interface that enhances association with Rev-RRE and poises NES binding sites to interact with a Rev oligomer. The interface between Crm1 monomers explains differences between Crm1 orthologs that alter nuclear export and determine cellular tropism for viral replication. The arrangement of the export complex identifies a novel binding surface to possibly target an HIV inhibitor and may point to a broader role for Crm1 dimerization in regulating host gene expression.

Protein SUMOylation is Involved in Cell-cycle Progression and Cell Morphology in Giardia lamblia.

PubMed

Di Genova, Bruno M; da Silva, Richard C; da Cunha, Júlia P C; Gargantini, Pablo R; Mortara, Renato A; Tonelli, Renata R

2017-07-01

The unicellular protozoa Giardia lamblia is a food- and waterborne parasite that causes giardiasis. This illness is manifested as acute and self-limited diarrhea and can evolve to long-term complications. Successful establishment of infection by Giardia trophozoites requires adhesion to host cells and colonization of the small intestine, where parasites multiply by mitotic division. The tight binding of trophozoites to host cells occurs by means of the ventral adhesive disc, a spiral array of microtubules and associated proteins such as giardins. In this work we show that knock down of the Small Ubiquitin-like MOdifier (SUMO) results in less adhesive trophzoites, decreased cell proliferation and deep morphological alterations, including at the ventral disc. Consistent with the reduced proliferation, SUMO knocked-down trophozoites were arrested in G1 and in S phases of the cell cycle. Mass spectrometry analysis of anti-SUMO immunoprecipitates was performed to identify SUMO substrates possibly involved in these events. Among the identified SUMOylation targets, α-tubulin was further validated by Western blot and confirmed to be a SUMO target in Giardia trophozoites. © 2016 The Author(s) Journal of Eukaryotic Microbiology © 2016 International Society of Protistologists.

The nuclear receptor PPARγ individually responds to serotonin- and fatty acid-metabolites

PubMed Central

Waku, Tsuyoshi; Shiraki, Takuma; Oyama, Takuji; Maebara, Kanako; Nakamori, Rinna; Morikawa, Kosuke

2010-01-01

The nuclear receptor, peroxisome proliferator-activated receptor γ (PPARγ), recognizes various synthetic and endogenous ligands by the ligand-binding domain. Fatty-acid metabolites reportedly activate PPARγ through conformational changes of the Ω loop. Here, we report that serotonin metabolites act as endogenous agonists for PPARγ to regulate macrophage function and adipogenesis by directly binding to helix H12. A cyclooxygenase inhibitor, indomethacin, is a mimetic agonist of these metabolites. Crystallographic analyses revealed that an indole acetate functions as a common moiety for the recognition by the sub-pocket near helix H12. Intriguingly, a serotonin metabolite and a fatty-acid metabolite each bind to distinct sub-pockets, and the PPARγ antagonist, T0070907, blocked the fatty-acid agonism, but not that of the serotonin metabolites. Mutational analyses on receptor-mediated transcription and coactivator binding revealed that each metabolite individually uses coregulator and/or heterodimer interfaces in a ligand-type-specific manner. Furthermore, the inhibition of the serotonin metabolism reduced the expression of the endogenous PPARγ-target gene. Collectively, these results suggest a novel agonism, in which PPARγ functions as a multiple sensor in response to distinct metabolites. PMID:20717101

A structural perspective on nuclear receptors as targets of environmental compounds

PubMed Central

Delfosse, Vanessa; Maire, Albane le; Balaguer, Patrick; Bourguet, William

2015-01-01

Nuclear receptors (NRs) are members of a large superfamily of evolutionarily related transcription factors that control a plethora of biological processes. NRs orchestrate complex events such as development, organ homeostasis, metabolism, immune function, and reproduction. Approximately one-half of the 48 human NRs have been shown to act as ligand-regulated transcription factors and respond directly to a large variety of endogenous hormones and metabolites that are generally hydrophobic and small in size (eg, retinoic acid or estradiol). The second half of the NR family comprises the so-called orphan receptors, for which regulatory ligands are still unknown or may not exist despite the presence of a C-terminal ligand-binding domain, which is the hallmark of all NRs. Several chemicals released into the environment (eg, bisphenols, phthalates, parabens, etc) share some physicochemical properties with natural ligands, allowing them to bind to NRs and activate or inhibit their action. Collectively referred to as endocrine disruptors or endocrine-disrupting chemicals (EDCs), these environmental pollutants are highly suspected to cause a wide range of developmental, reproductive, neurological, or metabolic defects in humans and wildlife. Crystallographic studies are revealing unanticipated mechanisms by which chemically diverse EDCs interact with the ligand-binding domain of NRs. These studies thereby provide a rational basis for designing novel chemicals with lower impacts on human and animal health. In this review, we provide a structural and mechanistic view of endocrine disrupting action using estrogen receptors α and β, (ERα/β), peroxisome proliferator activated receptor γ (PPARγ), and their respective environmental ligands as representative examples. PMID:25500867

A calreticulin-dependent nuclear export signal is involved in the regulation of liver receptor homologue-1 protein folding.

PubMed

Yang, Feng-Ming; Feng, Shan-Jung; Lai, Tsai-Chun; Hu, Meng-Chun

2015-10-15

As an orphan member of the nuclear receptor family, liver receptor homologue-1 (LRH-1) controls a tremendous range of transcriptional programmes that are essential for metabolism and hormone synthesis. Our previous studies have shown that nuclear localization of the LRH-1 protein is mediated by two nuclear localization signals (NLSs) that are karyopherin/importin-dependent. It is unclear whether LRH-1 can be actively exported from the nucleus to the cytoplasm. In the present study, we describe a nuclear export domain containing two leucine-rich motifs [named nuclear export signal (NES)1 and NES2] within the ligand-binding domain (LBD). Mutation of leucine residues in NES1 or NES2 abolished nuclear export, indicating that both NES1 and NES2 motifs are essential for full nuclear export activity. This NES-mediated nuclear export was insensitive to the chromosomal region maintenance 1 (CRM1) inhibitor leptomycin B (LMB) or to CRM1 knockdown. However, knockdown of calreticulin (CRT) prevented NES-mediated nuclear export. Furthermore, our data show that CRT interacts with LRH-1 and is involved in the nuclear export of LRH-1. With full-length LRH-1, mutation of NES1 led to perinuclear accumulation of the mutant protein. Immunofluorescence analysis showed that these perinuclear aggregates were co-localized with the centrosome marker, microtubule-associated protein 1 light chain 3 (LC3), ubiquitin and heat shock protein 70 (Hsp70), indicating that the mutant was misfolded and sequestered into aggresome-like structures via the autophagic clearance pathway. Our study demonstrates for the first time that LRH-1 has a CRT-dependent NES which is not only required for cytoplasmic trafficking, but also essential for correct protein folding to avoid misfolding-induced aggregation. © 2015 Authors; published by Portland Press Limited.

p35 Regulates the CRM1-Dependent Nucleocytoplasmic Shuttling of Nuclear Hormone Receptor Coregulator-Interacting Factor 1 (NIF-1)

PubMed Central

Zhao, Xiao-Su; Fu, Wing-Yu; Chien, Winnie W. Y.; Li, Zhen; Fu, Amy K. Y.; Ip, Nancy Y.

2014-01-01

Cyclin-dependent kinase 5 (Cdk5) is a proline-directed serine/threonine kinase, which plays critical roles in a wide spectrum of neuronal functions including neuronal survival, neurite outgrowth, and synapse development and plasticity. Cdk5 activity is controlled by its specific activators: p35 or p39. While knockout studies reveal that Cdk5/p35 is critical for neuronal migration during early brain development, functions of Cdk5/p35 have been unraveled through the identification of the interacting proteins of p35, most of which are Cdk5/p35 substrates. However, it remains unclear whether p35 can regulate neuronal functions independent of Cdk5 activity. Here, we report that a nuclear protein, nuclear hormone receptor coregulator (NRC)-interacting factor 1 (NIF-1), is a new interacting partner of p35. Interestingly, p35 regulates the functions of NIF-1 independent of Cdk5 activity. NIF-1 was initially discovered as a transcriptional regulator that enhances the transcriptional activity of nuclear hormone receptors. Our results show that p35 interacts with NIF-1 and regulates its nucleocytoplasmic trafficking via the nuclear export pathway. Furthermore, we identified a nuclear export signal on p35; mutation of this site or blockade of the CRM1/exportin-dependent nuclear export pathway resulted in the nuclear accumulation of p35. Intriguingly, blocking the nuclear export of p35 attenuated the nuclear accumulation of NIF-1. These findings reveal a new p35-dependent mechanism in transcriptional regulation that involves the nucleocytoplasmic shuttling of transcription regulators. PMID:25329792

p35 regulates the CRM1-dependent nucleocytoplasmic shuttling of nuclear hormone receptor coregulator-interacting factor 1 (NIF-1).

PubMed

Zhao, Xiao-Su; Fu, Wing-Yu; Chien, Winnie W Y; Li, Zhen; Fu, Amy K Y; Ip, Nancy Y

2014-01-01

Cyclin-dependent kinase 5 (Cdk5) is a proline-directed serine/threonine kinase, which plays critical roles in a wide spectrum of neuronal functions including neuronal survival, neurite outgrowth, and synapse development and plasticity. Cdk5 activity is controlled by its specific activators: p35 or p39. While knockout studies reveal that Cdk5/p35 is critical for neuronal migration during early brain development, functions of Cdk5/p35 have been unraveled through the identification of the interacting proteins of p35, most of which are Cdk5/p35 substrates. However, it remains unclear whether p35 can regulate neuronal functions independent of Cdk5 activity. Here, we report that a nuclear protein, nuclear hormone receptor coregulator (NRC)-interacting factor 1 (NIF-1), is a new interacting partner of p35. Interestingly, p35 regulates the functions of NIF-1 independent of Cdk5 activity. NIF-1 was initially discovered as a transcriptional regulator that enhances the transcriptional activity of nuclear hormone receptors. Our results show that p35 interacts with NIF-1 and regulates its nucleocytoplasmic trafficking via the nuclear export pathway. Furthermore, we identified a nuclear export signal on p35; mutation of this site or blockade of the CRM1/exportin-dependent nuclear export pathway resulted in the nuclear accumulation of p35. Intriguingly, blocking the nuclear export of p35 attenuated the nuclear accumulation of NIF-1. These findings reveal a new p35-dependent mechanism in transcriptional regulation that involves the nucleocytoplasmic shuttling of transcription regulators.

Variants of Transient Receptor Potential Melastatin Member 4 in Childhood Atrioventricular Block.

PubMed

Syam, Ninda; Chatel, Stéphanie; Ozhathil, Lijo Cherian; Sottas, Valentin; Rougier, Jean-Sébastien; Baruteau, Alban; Baron, Estelle; Amarouch, Mohamed-Yassine; Daumy, Xavier; Probst, Vincent; Schott, Jean-Jacques; Abriel, Hugues

2016-05-20

Transient receptor potential melastatin member 4 (TRPM4) is a nonselective cation channel. TRPM4 mutations have been linked to cardiac conduction disease and Brugada syndrome. The mechanisms underlying TRPM4-dependent conduction slowing are not fully understood. The aim of this study was to characterize TRPM4 genetic variants found in patients with congenital or childhood atrioventricular block. Ninety-one patients with congenital or childhood atrioventricular block were screened for candidate genes. Five rare TRPM4 genetic variants were identified and investigated. The variants were expressed heterologously in HEK293 cells. Two of the variants, A432T and A432T/G582S, showed decreased expression of the protein at the cell membrane; inversely, the G582S variant showed increased expression. Further functional characterization of these variants using whole-cell patch-clamp configuration showed a loss of function and a gain of function, respectively. We hypothesized that the observed decrease in expression was caused by a folding and trafficking defect. This was supported by the observation that incubation of these variants at lower temperature partially rescued their expression and function. Previous studies have suggested that altered SUMOylation of TRPM4 may cause a gain of function; however, we did not find any evidence that supports SUMOylation as being directly involved for the gain-of-function variant. This study underpins the role of TRPM4 in the cardiac conduction system. The loss-of-function variants A432T/G582S found in 2 unrelated patients with atrioventricular block are most likely caused by misfolding-dependent altered trafficking. The ability to rescue this variant with lower temperature may provide a novel use of pharmacological chaperones in treatment strategies. © 2016 The Authors. Published on behalf of the American Heart Association, Inc., by Wiley Blackwell.

Nuclear receptor ERR alpha and coactivator PGC-1 beta are effectors of IFN-gamma-induced host defense.

PubMed

Sonoda, Junichiro; Laganière, Josée; Mehl, Isaac R; Barish, Grant D; Chong, Ling-Wa; Li, Xiangli; Scheffler, Immo E; Mock, Dennis C; Bataille, Alain R; Robert, Francois; Lee, Chih-Hao; Giguère, Vincent; Evans, Ronald M

2007-08-01

Macrophage activation by the proinflammatory cytokine interferon-gamma (IFN-gamma) is a critical component of the host innate response to bacterial pathogenesis. However, the precise nature of the IFN-gamma-induced activation pathway is not known. Here we show using genome-wide expression and chromatin-binding profiling that IFN-gamma induces the expression of many nuclear genes encoding mitochondrial respiratory chain machinery via activation of the nuclear receptor ERR alpha (estrogen-related receptor alpha, NR3B1). Studies with macrophages lacking ERR alpha demonstrate that it is required for induction of mitochondrial reactive oxygen species (ROS) production and efficient clearance of Listeria monocytogenes (LM) in response to IFN-gamma. As a result, mice lacking ERR alpha are susceptible to LM infection, a phenotype that is localized to bone marrow-derived cells. Furthermore, we found that IFN-gamma-induced activation of ERR alpha depends on coactivator PGC-1 beta (peroxisome proliferator-activated receptor gamma coactivator-1 beta), which appears to be a direct target for the IFN-gamma/STAT-1 signaling cascade. Thus, ERR alpha and PGC-1 beta act together as a key effector of IFN-gamma-induced mitochondrial ROS production and host defense.

Expression and Functional Pathway Analysis of Nuclear Receptor NR2F2 in Ovarian Cancer

PubMed Central

Hawkins, Shannon M.; Loomans, Holli A.; Wan, Ying-Wooi; Ghosh-Choudhury, Triparna; Coffey, Donna; Xiao, Weimin; Liu, Zhandong; Sangi-Haghpeykar, Haleh

2013-01-01

Context: Recent evidence implicates the orphan nuclear receptor, nuclear receptor subfamily 2, group F, member 2 (NR2F2; chicken ovalbumin upstream promoter-transcription factor II) as both a master regulator of angiogenesis and an oncogene in prostate and other human cancers. Objective: The objective of the study was to determine whether NR2F2 plays a role in ovarian cancer and dissect its potential mechanisms of action. Design, Setting, and Patients: We examined NR2F2 expression in healthy ovary and ovarian cancers using quantitative PCR and immunohistochemistry. NR2F2 expression was targeted in established ovarian cancer cell lines to assess the impact of dysregulated NR2F2 expression in the epithelial compartment of ovarian cancers. Results: Our results indicate that NR2F2 is robustly expressed in the stroma of healthy ovary with little or no expression in epithelia lining the ovarian surface, clefts, or crypts. This pattern of NR2F2 expression was markedly disrupted in ovarian cancers, in which decreased levels of stromal expression and ectopic epithelial expression were frequently observed. Ovarian cancers with the most disrupted patterns of NR2F2 were associated with significantly shorter disease-free interval by Kaplan-Meier analysis. Targeting NR2F2 expression in established ovarian cancer cell lines enhanced apoptosis and increased proliferation. In addition, we found that NR2F2 regulates the expression of NEK2, RAI14, and multiple other genes involved in the cell cycle, suggesting potential pathways by which dysregulated expression of NR2F2 impacts ovarian cancer. Conclusions: These results uncover novel roles for NR2F2 in ovarian cancer and point to a unique scenario in which a single nuclear receptor plays potentially distinct roles in the stromal and epithelial compartments of the same tissue. PMID:23690307

Proteomics Analysis of Nucleolar SUMO-1 Target Proteins upon Proteasome Inhibition*

PubMed Central

Matafora, Vittoria; D'Amato, Alfonsina; Mori, Silvia; Blasi, Francesco; Bachi, Angela

2009-01-01

Many cellular processes are regulated by the coordination of several post-translational modifications that allow a very fine modulation of substrates. Recently it has been reported that there is a relationship between sumoylation and ubiquitination. Here we propose that the nucleolus is the key organelle in which SUMO-1 conjugates accumulate in response to proteasome inhibition. We demonstrated that, upon proteasome inhibition, the SUMO-1 nuclear dot localization is redirected to nucleolar structures. To better understand this process we investigated, by quantitative proteomics, the effect of proteasome activity on endogenous nucleolar SUMO-1 targets. 193 potential SUMO-1 substrates were identified, and interestingly in several purified SUMO-1 conjugates ubiquitin chains were found to be present, confirming the coordination of these two modifications. 23 SUMO-1 targets were confirmed by an in vitro sumoylation reaction performed on nuclear substrates. They belong to protein families such as small nuclear ribonucleoproteins, heterogeneous nuclear ribonucleoproteins, ribosomal proteins, histones, RNA-binding proteins, and transcription factor regulators. Among these, histone H1, histone H3, and p160 Myb-binding protein 1A were further characterized as novel SUMO-1 substrates. The analysis of the nature of the SUMO-1 targets identified in this study strongly indicates that sumoylation, acting in coordination with the ubiquitin-proteasome system, regulates the maintenance of nucleolar integrity. PMID:19596686

The yeast SUMO isopeptidase Smt4/Ulp2 and the polo kinase Cdc5 act in an opposing fashion to regulate sumoylation in mitosis and cohesion at centromeres.

PubMed

Baldwin, Melissa L; Julius, Jeffrey A; Tang, Xianying; Wang, Yanchang; Bachant, Jeff

2009-10-15

Post-translation modification through the SUMO pathway is cell cycle regulated, with specific SUMO conjugates accumulating in mitotic cells. The basis for this regulation, however, and its functional significance remain poorly understood. We present evidence that in budding yeast sumoylation during mitosis may be controlled through the SUMO deconjugating enzyme Smt4/Ulp2. We isolated the polo kinase Cdc5 as an Ulp2-interacting protein, and find a C-terminal region of Ulp2 is phosphorylated during mitosis in a Cdc5-dependent manner. cdc5 mutants display reduced levels of mitotic SUMO conjugates, suggesting Cdc5 may negatively regulate Ulp2 to promote sumoylation. Previously, we found one phenotype associated with ulp2 mutants is an inability to maintain chromatid cohesion at centromere-proximal chromosomal regions. We now show this defect is rescued by inactivating Cdc5, indicating Ulp2 maintains cohesion by counter-acting Cdc5 activity. The cohesinregulator Pds5 is a likely target of this pathway, as Cdc5 overproduction forces Pds5 dissociation from chromosomes and Pds5 overproduction restores cohesion in ulp2 mutants. Overall, these observations reveal Cdc5 is a novel regulator of the SUMO pathway and suggest the outlines of a broader circuitry in which Ulp2 and Cdc5 act in a mutually antagonistic fashion to modulate maintenance and dissolution of cohesion at centromeres.

Activation of orphan nuclear constitutive androstane receptor requires subnuclear targeting by peroxisome proliferator-activated receptor gamma coactivator-1 alpha. A possible link between xenobiotic response and nutritional state.

PubMed

Shiraki, Takuma; Sakai, Noriko; Kanaya, Eiko; Jingami, Hisato

2003-03-28

In contrast to the classical nuclear receptors, the constitutive androstane receptor (CAR) is transcriptionally active in the absence of ligand. In the course of searching for the mediator of CAR activation, we found that ligand-independent activation of CAR was achieved in cooperation with the peroxisome proliferator-activated receptor gamma coactivator-1 alpha (PGC-1 alpha). PGC-1 beta, a PGC-1 alpha homologue, also activated CAR to less of an extent than PGC-1 alpha. Coexpression of the ligand-binding domain of a heterodimerization partner, retinoid X receptor alpha, enhanced the PGC-1 alpha-mediated activation of CAR, although it had a weak effect on the basal activity of CAR in the absence of PGC-1 alpha. Both the N-terminal region, with the LXXLL motif, and the C-terminal region, with a serine/arginine-rich domain (RS domain), in PGC-1 alpha were required for full activation of CAR. Pull-down experiments using recombinant proteins revealed that CAR directly interacted with both the LXXLL motif and the RS domain. Furthermore, we demonstrated that the RS domain of PGC-1 alpha was required for CAR localization at nuclear speckles. These results indicate that PGC-1 alpha mediates the ligand-independent activation of CAR by means of subnuclear targeting through the RS domain of PGC-1 alpha.

Regulation of antibacterial defense in the small intestine by the nuclear bile acid receptor.

PubMed

Inagaki, Takeshi; Moschetta, Antonio; Lee, Youn-Kyoung; Peng, Li; Zhao, Guixiang; Downes, Michael; Yu, Ruth T; Shelton, John M; Richardson, James A; Repa, Joyce J; Mangelsdorf, David J; Kliewer, Steven A

2006-03-07

Obstruction of bile flow results in bacterial proliferation and mucosal injury in the small intestine that can lead to the translocation of bacteria across the epithelial barrier and systemic infection. These adverse effects of biliary obstruction can be inhibited by administration of bile acids. Here we show that the farnesoid X receptor (FXR), a nuclear receptor for bile acids, induces genes involved in enteroprotection and inhibits bacterial overgrowth and mucosal injury in ileum caused by bile duct ligation. Mice lacking FXR have increased ileal levels of bacteria and a compromised epithelial barrier. These findings reveal a central role for FXR in protecting the distal small intestine from bacterial invasion and suggest that FXR agonists may prevent epithelial deterioration and bacterial translocation in patients with impaired bile flow.

Tributyltin engages multiple nuclear receptor pathways and suppresses osteogenesis in bone marrow multipotent stromal cells.

PubMed

Baker, Amelia H; Watt, James; Huang, Cassie K; Gerstenfeld, Louis C; Schlezinger, Jennifer J

2015-06-15

Organotins are members of the environmental obesogen class of contaminants because they activate peroxisome proliferator-activated receptor γ (PPARγ), the essential regulator of adipogenesis. Exposure to thiazolidinediones (PPARγ ligands used to treat type 2 diabetes) is associated with increased fractures. Diminished bone quality likely results from PPARγ's role in promoting adipogenesis while suppressing osteogenesis of bone marrow multipotent mesenchymal stromal cells (BM-MSC). We hypothesized that tributyltin (TBT) would be a potent modifier of BM-MSC differentiation and a negative regulator of bone formation. Organotins interact with both PPARγ and retinoid X receptors (RXR), suggesting that they activate multiple nuclear receptor pathways. To investigate the role of RXR in the actions of TBT, the effects of PPARγ (rosiglitazone) and RXR (bexarotene, LG100268) agonists were compared to the effects of TBT in BMS2 cells and primary mouse BM-MSC cultures. In BMS2 cells, TBT induced the expression of Fabp4, Abca1, and Tgm2 in an RXR-dependent manner. All agonists suppressed osteogenesis in primary mouse BM-MSC cultures, based on decreased alkaline phosphatase activity, mineralization, and expression of osteoblast-related genes. While rosiglitazone and TBT strongly activated adipogenesis, based on lipid accumulation and expression of adipocyte-related genes, the RXR agonists did not. Extending these analyses to other RXR heterodimers showed that TBT and the RXR agonists activated the liver X receptor pathway, whereas rosiglitazone did not. Application of either a PPARγ antagonist (T0070907) or an RXR antagonist (HX531) significantly reduced rosiglitazone-induced suppression of bone nodule formation. Only the RXR antagonist significantly reduced LG100268- and TBT-induced bone suppression. The RXR antagonist also inhibited LG100268- and TBT-induced expression of Abca1, an LXR target gene, in primary BM-MSC cultures. These results provide novel evidence that

Action mechanisms of Liver X Receptors

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gabbi, Chiara; Warner, Margaret; Gustafsson, Jan-Åke, E-mail: jgustafs@central.uh.edu

2014-04-11

Highlights: • LXRα and LXRβ are ligand-activated nuclear receptors. • They share oxysterol ligands and the same heterodimerization partner, RXR. • LXRs regulate lipid and glucose metabolism, CNS and immune functions, and water transport. - Abstract: The two Liver X Receptors, LXRα and LXRβ, are nuclear receptors belonging to the superfamily of ligand-activated transcription factors. They share more than 78% homology in amino acid sequence, a common profile of oxysterol ligands and the same heterodimerization partner, Retinoid X Receptor. LXRs play crucial roles in several metabolic pathways: lipid metabolism, in particular in preventing cellular cholesterol accumulation; glucose homeostasis; inflammation; centralmore » nervous system functions and water transport. As with all nuclear receptors, the transcriptional activity of LXR is the result of an orchestration of numerous cellular factors including ligand bioavailability, presence of corepressors and coactivators and cellular context i.e., what other pathways are activated in the cell at the time the receptor recognizes its ligand. In this mini-review we summarize the factors regulating the transcriptional activity and the mechanisms of action of these two receptors.« less

Phytol metabolites are circulating dietary factors that activate the nuclear receptor RXR.

PubMed Central

Kitareewan, S; Burka, L T; Tomer, K B; Parker, C E; Deterding, L J; Stevens, R D; Forman, B M; Mais, D E; Heyman, R A; McMorris, T; Weinberger, C

1996-01-01

RXR is a nuclear receptor that plays a central role in cell signaling by pairing with a host of other receptors. Previously, 9-cis-retinoic acid (9cRA) was defined as a potent RXR activator. Here we describe a unique RXR effector identified from organic extracts of bovine serum by following RXR-dependent transcriptional activity. Structural analyses of material in active fractions pointed to the saturated diterpenoid phytanic acid, which induced RXR-dependent transcription at concentrations between 4 and 64 microM. Although 200 times more potent than phytanic acid, 9cRA was undetectable in equivalent amounts of extract and cannot be present at a concentration that could account for the activity. Phytanic acid, another phytol metabolite, was synthesized and stimulated RXR with a potency and efficacy similar to phytanic acid. These metabolites specifically displaced [3H]-9cRA from RXR with Ki values of 4 microM, indicating that their transcriptional effects are mediated by direct receptor interactions. Phytol metabolites are compelling candidates for physiological effectors, because their RXR binding affinities and activation potencies match their micromolar circulating concentrations. Given their exclusive dietary origin, these chlorophyll metabolites may represent essential nutrients that coordinate cellular metabolism through RXR-dependent signaling pathways. PMID:8856661

A Transcriptional Regulatory Network Containing Nuclear Receptors and Long Noncoding RNAs Controls Basal and Drug-Induced Expression of Cytochrome P450s in HepaRG Cells.

PubMed

Chen, Liming; Bao, Yifan; Piekos, Stephanie C; Zhu, Kexin; Zhang, Lirong; Zhong, Xiao-Bo

2018-07-01

Cytochrome P450 (P450) enzymes are responsible for metabolizing drugs. Expression of P450s can directly affect drug metabolism, resulting in various outcomes in therapeutic efficacy and adverse effects. Several nuclear receptors are transcription factors that can regulate expression of P450s at both basal and drug-induced levels. Some long noncoding RNAs (lncRNAs) near a transcription factor are found to participate in the regulatory functions of the transcription factors. The aim of this study is to determine whether there is a transcriptional regulatory network containing nuclear receptors and lncRNAs controlling both basal and drug-induced expression of P450s in HepaRG cells. Small interfering RNAs or small hairpin RNAs were applied to knock down four nuclear receptors [hepatocyte nuclear factor 1 α (HNF1 α ), hepatocyte nuclear factor 4 α (HNF4 α ), pregnane X receptor (PXR), and constitutive androstane receptor (CAR)] as well as two lncRNAs [HNF1 α antisense RNA 1 (HNF1 α -AS1) and HNF4 α antisense RNA 1 (HNF4 α -AS1)] in HepaRG cells with or without treatment of phenobarbital or rifampicin. Expression of eight P450 enzymes was examined in both basal and drug-induced levels. CAR and PXR mainly regulated expression of specific P450s. HNF1 α and HNF4 α affected expression of a wide range of P450s as well as other transcription factors. HNF1 α and HNF4 α controlled the expression of their neighborhood lncRNAs, HNF1 α -AS1 and HNF4 α -AS1, respectively. HNF1 α -AS1 and HNF4 α -AS1 was also involved in the regulation of P450s and transcription factors in diverse manners. Altogether, our study concludes that a transcription regulatory network containing the nuclear receptors and lncRNAs controls both basal and drug-induced expression of P450s in HepaRG cells. Copyright © 2018 by The American Society for Pharmacology and Experimental Therapeutics.

Metabotropic glutamate receptor 5 mediates phosphorylation of vascular endothelial cadherin and nuclear localization of β-catenin in response to homocysteine.

PubMed

Beard, Richard S; Reynolds, Jason J; Bearden, Shawn E

2012-01-01

Elevated plasma homocysteine (Hcy) is an independent risk factor for vascular disease and stroke in part by causing generalized endothelial dysfunction. A receptor that is sensitive to Hcy and its intracellular signaling systems has not been identified. β-catenin is a pleiotropic regulator of transcription and cell function. Using a brain microvascular endothelial cell line (bEnd.3), we tested the hypothesis that Hcy causes receptor-dependent nuclear translocation of β-catenin. Hcy increased phosphorylation of Y731 on vascular endothelial cadherin (VE-cadherin), a site involved in coupling β-catenin to VE-cadherin. This was blocked by inhibition of either metabotropic glutamate receptor 5 (mGluR5) or ionotropic glutamate receptor (NMDAr) and by shRNA knockdown of mGluR5. Expression of these receptors was confirmed by flow cytometry, immunohistochemistry, and western blotting. Directed pharmacology with specific agonists elucidated a signaling cascade where Hcy activates mGluR5 which activates NMDAr with subsequent PKC activation and uncoupling of the VE-cadherin/β-catenin complex. Moreover, Hcy caused a shift in localization of β-catenin from membrane-bound VE-cadherin to the cell nucleus, where it bound DNA, including a regulatory region of the gene for claudin-5, leading to reduced expression of claudin-5. Nuclear localization, DNA binding of β-catenin, and reduced claudin-5 expression were blocked by inhibition of mGluR5. Knockdown of mGluR5 expression with shRNA also rescued claudin-5 expression from the effects of Hcy treatment. These data uniquely identify mGluR5 as a master switch that drives β-catenin nuclear localization in vascular endothelium and regulates cell-cell coupling in response to elevated Hcy levels. These studies dissect a pharmacological opportunity for developing new therapeutic strategies in HHcy. Copyright © 2012 Elsevier Inc. All rights reserved.

Orphan nuclear receptor ERRγ is a key regulator of human fibrinogen gene expression

PubMed Central

Zhang, Yaochen; Kim, Don-Kyu; Lu, Yan; Jung, Yoon Seok; Lee, Ji-min; Kim, Young-Hoon; Lee, Yong Soo; Kim, Jina; Dewidar, Bedair; Jeong, Won-IL; Lee, In-Kyu; Cho, Sung Jin; Dooley, Steven; Lee, Chul-Ho; Li, Xiaoying

2017-01-01

Fibrinogen, 1 of 13 coagulation factors responsible for normal blood clotting, is synthesized by hepatocytes. Detailed roles of the orphan nuclear receptors regulating fibrinogen gene expression have not yet been fully elucidated. Here, we identified estrogen-related receptor gamma (ERRγ) as a novel transcriptional regulator of human fibrinogen gene expression. Overexpression of ERRγ specially increased fibrinogen expression in human hepatoma cell line. Cannabinoid receptor types 1(CB1R) agonist arachidonyl-2'-chloroethylamide (ACEA) up-regulated transcription of fibrinogen via induction of ERRγ, whereas knockdown of ERRγ attenuated fibrinogen expression. Deletion analyses of the fibrinogen γ (FGG) gene promoter and ChIP assays revealed binding sites of ERRγ on human fibrinogen γ gene promoter. Moreover, overexpression of ERRγ was sufficient to increase fibrinogen gene expression, whereas treatment with GSK5182, a selective inverse agonist of ERRγ led to its attenuation in cell culture. Finally, fibrinogen and ERRγ gene expression were elevated in liver tissue of obese patients suggesting a conservation of this mechanism. Overall, this study elucidates a molecular mechanism linking CB1R signaling, ERRγ expression and fibrinogen gene transcription. GSK5182 may have therapeutic potential to treat hyperfibrinogenemia. PMID:28750085

Transcriptional targets shared by estrogen receptor- related receptors (ERRs) and estrogen receptor (ER) alpha, but not by ERbeta.

PubMed Central

Vanacker, J M; Pettersson, K; Gustafsson, J A; Laudet, V

1999-01-01

The physiological activities of estrogens are thought to be mediated by specific nuclear receptors, ERalpha and ERbeta. However, certain tissues, such as the bone, that are highly responsive to estrogens only express a low level of these receptors. Starting from this apparent contradiction, we have evaluated the potentials of two related receptors ERRalpha and ERRbeta to intervene in estrogen signaling. ERalpha, ERRalpha and ERRbeta bind to and activate transcription through both the classical estrogen response element (ERE) and the SF-1 response element (SFRE). In contrast, ERbeta DNA-binding and transcriptional activity is restricted to the ERE. Accordingly, the osteopontin gene promoter is stimulated through SFRE sequences, by ERRalpha as well as by ERalpha, but not by ERbeta. Analysis of the cross-talk within the ER/ERR subgroup of nuclear receptors thus revealed common targets but also functional differences between the two ERs. PMID:10428965

A live zebrafish-based screening system for human nuclear receptor ligand and cofactor discovery.

PubMed

Tiefenbach, Jens; Moll, Pamela R; Nelson, Meryl R; Hu, Chun; Baev, Lilia; Kislinger, Thomas; Krause, Henry M

2010-03-22

Nuclear receptors (NRs) belong to a superfamily of transcription factors that regulate numerous homeostatic, metabolic and reproductive processes. Taken together with their modulation by small lipophilic molecules, they also represent an important and successful class of drug targets. Although many NRs have been targeted successfully, the majority have not, and one third are still orphans. Here we report the development of an in vivo GFP-based reporter system suitable for monitoring NR activities in all cells and tissues using live zebrafish (Danio rerio). The human NR fusion proteins used also contain a new affinity tag cassette allowing the purification of receptors with bound molecules from responsive tissues. We show that these constructs 1) respond as expected to endogenous zebrafish hormones and cofactors, 2) facilitate efficient receptor and cofactor purification, 3) respond robustly to NR hormones and drugs and 4) yield readily quantifiable signals. Transgenic lines representing the majority of human NRs have been established and are available for the investigation of tissue- and isoform-specific ligands and cofactors.

Orphan nuclear receptor small heterodimer partner inhibits transforming growth factor-beta signaling by repressing SMAD3 transactivation.

PubMed

Suh, Ji Ho; Huang, Jiansheng; Park, Yun-Yong; Seong, Hyun-A; Kim, Dongwook; Shong, Minho; Ha, Hyunjung; Lee, In-Kyu; Lee, Keesook; Wang, Li; Choi, Hueng-Sik

2006-12-22

Orphan nuclear receptor small heterodimer partner (SHP) is an atypical member of the nuclear receptor superfamily; SHP regulates the nuclear receptor-mediated transcription of target genes but lacks a conventional DNA binding domain. In this study, we demonstrate that SHP represses transforming growth factor-beta (TGF-beta)-induced gene expression through a direct interaction with Smad, a transducer of TGF-beta signaling. Transient transfection studies demonstrate that SHP represses Smad3-induced transcription. In vivo and in vitro protein interaction assays revealed that SHP directly interacts with Smad2 and Smad3 but not with Smad4. Mapping of domains mediating the interaction between SHP and Smad3 showed that the entire N-terminal domain (1-159 amino acids) of SHP and the linker domain of Smad3 are involved in this interaction. In vitro glutathione S-transferase pulldown competition experiments revealed the SHP-mediated repression of Smad3 transactivation through competition with its co-activator p300. SHP also inhibits the activation of endogenous TGF-beta-responsive gene promoters, the p21, Smad7, and plasminogen activator inhibitor-1 (PAI-1) promoters. Moreover, adenovirus-mediated overexpression of SHP decreases PAI-1 mRNA levels, and down-regulation of SHP by a small interfering RNA increases both the transactivation of Smad3 and the PAI-1 mRNA levels. Finally, the PAI-1 gene is expressed in SHP(-/-) mouse hepatocytes at a higher level than in normal hepatocytes. Taken together, these data indicate that SHP is a novel co-regulator of Smad3, and this study provides new insights into regulation of TGF-beta signaling.

Evidence for triclosan-induced activation of human and rodent xenobiotic nuclear receptors.

PubMed

Paul, Katie B; Thompson, Jerry T; Simmons, Steven O; Vanden Heuvel, John P; Crofton, Kevin M

2013-10-01

The bacteriostat triclosan (2,4,4'-trichloro-2'-hydroxydiphenylether) (TCS) decreases rat serum thyroxine via putative nuclear receptor (NR) interaction(s) and subsequent transcriptional up-regulation of hepatic catabolism and clearance. However, due to the evolutionary divergence of the constitutive androstane and pregnane-X receptors (CAR, PXR), TCS-mediated downstream effects may be species-dependent. To test the hypothesis that TCS activates xenobiotic NRs across species, cell-based NR reporter assays were employed to assess potential activation of rat, mouse, and human PXR, and rat, mouse, and three splice variants of human CAR. TCS activated hPXR, acted as an inverse agonist of hCAR1, and as a weak agonist of hCAR3. TCS failed to activate rPXR in full-length receptor reporter assays, and instead acted as a modest inverse agonist of rCAR. Consistent with the rat data, TCS also failed to activate mPXR and was a modest inverse agonist of mCAR. These data suggest that TCS may interact with multiple NRs, including hPXR, hCAR1, hCAR3, and rCAR in order to potentially affect hepatic catabolism. Overall these data support the conclusion that TCS may interact with NRs to regulate hepatic catabolism and downstream thyroid hormone homeostasis in both rat and human models, though perhaps by divergent mechanisms. Published by Elsevier Ltd.

Phosphorylation and Intramolecular Stabilization of the Ligand Binding Domain in the Nuclear Receptor Steroidogenic Factor 1

PubMed Central

Desclozeaux, Marion; Krylova, Irina N.; Horn, Florence; Fletterick, Robert J.; Ingraham, Holly A.

2002-01-01

Steroidogenic factor 1 (SF-1) is an orphan nuclear receptor with no known ligand. We showed previously that phosphorylation at serine 203 located N′-terminal to the ligand binding domain (LBD) enhanced cofactor recruitment, analogous to the ligand-mediated recruitment in ligand-dependent receptors. In this study, results of biochemical analyses and an LBD helix assembly assay suggest that the SF-1 LBD adopts an active conformation, with helices 1 and 12 packed against the predicted alpha-helical bundle, in the apparent absence of ligand. Fine mapping of the previously defined proximal activation function in SF-1 showed that the activation function mapped fully to helix 1 of the LBD. Limited proteolyses demonstrate that phosphorylation of S203 in the hinge region mimics the stabilizing effects of ligand on the LBD. Moreover, similar effects were observed in an SF-1/thyroid hormone LBD chimera receptor, illustrating that the S203 phosphorylation effects are transferable to a heterologous ligand-dependent receptor. Our collective data suggest that the hinge together with helix 1 is an individualized specific motif, which is tightly associated with its cognate LBD. For SF-1, we find that this intramolecular association and hence receptor activity are further enhanced by mitogen-activated protein kinase phosphorylation, thus mimicking many of the ligand-induced changes observed for ligand-dependent receptors. PMID:12242296

Both conditional ablation and overexpression of E2 SUMO-conjugating enzyme (UBC9) in mouse pancreatic beta cells result in impaired beta cell function.

PubMed

He, Xiaoyu; Lai, Qiaohong; Chen, Cai; Li, Na; Sun, Fei; Huang, Wenting; Zhang, Shu; Yu, Qilin; Yang, Ping; Xiong, Fei; Chen, Zhishui; Gong, Quan; Ren, Boxu; Weng, Jianping; Eizirik, Décio L; Zhou, Zhiguang; Wang, Cong-Yi

2018-04-01

Post-translational attachment of a small ubiquitin-like modifier (SUMO) to the lysine (K) residue(s) of target proteins (SUMOylation) is an evolutionary conserved regulatory mechanism. This modification has previously been demonstrated to be implicated in the control of a remarkably versatile regulatory mechanism of cellular processes. However, the exact regulatory role and biological actions of the E2 SUMO-conjugating enzyme (UBC9)-mediated SUMOylation function in pancreatic beta cells has remained elusive. Inducible beta cell-specific Ubc9 (also known as Ube2i) knockout (KO; Ubc9 Δbeta ) and transgenic (Ubc9 Tg ) mice were employed to address the impact of SUMOylation on beta cell viability and functionality. Ubc9 deficiency or overexpression was induced at 8 weeks of age using tamoxifen. To study the mechanism involved, we closely examined the regulation of the transcription factor nuclear factor erythroid 2-related factor 2 (NRF2) through SUMOylation in beta cells. Upon induction of Ubc9 deficiency, Ubc9 Δbeta islets exhibited a 3.5-fold higher accumulation of reactive oxygen species (ROS) than Ubc9 f/f control islets. Islets from Ubc9 Δbeta mice also had decreased insulin content and loss of beta cell mass after tamoxifen treatment. Specifically, at day 45 after Ubc9 deletion only 40% of beta cell mass remained in Ubc9 Δbeta mice, while 90% of beta cell mass was lost by day 75. Diabetes onset was noted in some Ubc9 Δbeta mice 8 weeks after induction of Ubc9 deficiency and all mice developed diabetes by 10 weeks following tamoxifen treatment. In contrast, Ubc9 Tg beta cells displayed an increased antioxidant ability but impaired insulin secretion. Unlike Ubc9 Δbeta mice, which spontaneously developed diabetes, Ubc9 Tg mice preserved normal non-fasting blood glucose levels without developing diabetes. It was noted that SUMOylation of NRF2 promoted its nuclear expression along with enhanced transcriptional activity, thereby preventing ROS accumulation in

Hormonally active phytochemicals from macroalgae: A largely untapped source of ligands to deorphanize nuclear receptors in emerging marine animal models.

PubMed

Markov, Gabriel V; Girard, Jean; Laudet, Vincent; Leblanc, Catherine

2018-06-15

Hormonally active phytochemicals (HAPs) are signaling molecules produced by plants that alter hormonal signaling in animals, due to consumption or environmental exposure. To date, HAPs have been investigated mainly in terrestrial ecosystems. To gain a full understanding of the origin and evolution of plant-animal interactions, it is necessary also to study these interactions in the marine environment, where the major photosynthetic lineages are very distant from the terrestrial plants. Here we focus on chemicals from red and brown macroalgae and point out their potential role as modulators of the endocrine system of aquatic animals through nuclear hormone receptors. We show that, regarding steroids and oxylipins, there are already some candidates available for further functional investigations of ligand-receptor interactions. Furthermore, several carotenoids, produced by cyanobacteria provide candidates that could be investigated with respect to their presence in macroalgae. Finally, regarding halogenated compounds, it is not clear yet which molecules could bridge the gap to explain the transition from lipid sensing to thyroid hormone high affinity binding among nuclear receptors. Copyright © 2018 Elsevier Inc. All rights reserved.

Nuclear Receptors and AMPK: Can Exercise Mimetics Cure Diabetes?

PubMed Central

Wall, Christopher E.; Yu, Ruth T.; Atkins, Anne R.; Downes, Michael; Evans, Ronald M.

2016-01-01

Endurance exercise can lead to systemic improvements in insulin sensitivity and metabolic homeostasis, and is an effective approach to combat metabolic diseases. Pharmacological compounds that recapitulate the beneficial effects of exercise, also known as “exercise mimetics,” have the potential to improve disease symptoms of metabolic syndrome. These drugs, which can increase energy expenditure, suppress hepatic gluconeogenesis, and induce insulin sensitization, have accordingly been highly scrutinized for their utility in treating metabolic diseases including diabetes. Nevertheless, the identity of an efficacious exercise mimetic still remains elusive. In this article, we will highlight several nuclear receptors and cofactors that are putative molecular targets for exercise mimetics, and review recent studies that provide advancements in our mechanistic understanding of how exercise mimetics exert their beneficial effects. We will also discuss evidence from clinical trials utilizing these compounds in human subjects to evaluate their efficacy in treating diabetes. PMID:27106806

Growth hormone-specific induction of the nuclear localization of porcine growth hormone receptor in porcine hepatocytes.

PubMed

Lan, H N; Hong, P; Li, R N; Shan, A S; Zheng, X

2017-10-01

The phenomenon of nuclear translocation of growth hormone receptor (GHR) in human, rat, and fish has been reported. To date, this phenomenon has not been described in a domestic animal (such as pig). In addition, the molecular mechanisms of GHR nuclear translocation have not been thoroughly elucidated. To this end, porcine hepatocytes were isolated and used as a cell model. We observed that porcine growth hormone (pGH) can induce porcine GHR's nuclear localization in porcine hepatocytes. Subsequently, the dynamics of pGH-induced pGHR's nuclear localization were analyzed and demonstrated that pGHR's nuclear localization occurs in a time-dependent manner. Next, we explored the mechanism of pGHR nuclear localization using different pGHR ligands, and we demonstrated that pGHR's nuclear translocation is GH(s)-dependent. We also observed that pGHR translocates into cell nuclei in a pGH dimerization-dependent fashion, whereas further experiments indicated that IMPα/β is involved in the nuclear translocation of the pGH-pGHR dimer. The pGH-pGHR dimer may form a pGH-GHR-JAK2 multiple complex in cell nuclei, which would suggest that similar to its function in the cell membrane, the nuclear-localized pGH-pGHR dimer might still have the ability to signal. Copyright © 2017 Elsevier Inc. All rights reserved.

Nuclear accumulation of the Arabidopsis immune receptor RPS4 is necessary for triggering EDS1-dependent defense.

PubMed

Wirthmueller, Lennart; Zhang, Yan; Jones, Jonathan D G; Parker, Jane E

2007-12-04

Recognition of specific pathogen molecules inside the cell by nucleotide-binding domain and leucine-rich repeat (NB-LRR) receptors constitutes an important layer of innate immunity in plants. Receptor activation triggers host cellular reprogramming involving transcriptional potentiation of basal defenses and localized programmed cell death. The sites and modes of action of NB-LRR receptors are, however, poorly understood. Arabidopsis Toll/Interleukin-1 (TIR) type NB-LRR receptor RPS4 recognizes the bacterial type III effector AvrRps4. We show that epitope-tagged RPS4 expressed under its native regulatory sequences distributes between endomembranes and nuclei in healthy and AvrRps4-triggered tissues. RPS4 accumulation in the nucleus, mediated by a bipartite nuclear localization sequence (NLS) at its C terminus, is necessary for triggering immunity through authentic activation by AvrRps4 in Arabidopsis or as an effector-independent "deregulated" receptor in tobacco. A strikingly conserved feature of TIR-NB-LRR receptors is their recruitment of the nucleocytoplasmic basal-defense regulator EDS1 in resistance to diverse pathogens. We find that EDS1 is an indispensable component of RPS4 signaling and that it functions downstream of RPS4 activation but upstream of RPS4-mediated transcriptional reprogramming in the nucleus.

Mechanism regulating nuclear calcium signaling.

PubMed

Malviya, Anant N; Klein, Christian

2006-01-01

Although the outer nuclear membrane is continuous with the endoplasmic reticulum, it is possible to isolate nuclei both intact and free from endoplasmic reticulum contaminants. The outer and the inner nuclear membranes can be purified free from cross-contamination. Evidence in support of autonomous regulation of nuclear calcium signaling relies upon the investigations with isolated nuclei. Mechanisms for generating calcium signaling in the nucleus have been identified. Two calcium transporting systems, an ATP-dependant nuclear Ca(2+)-ATPase and an IP4-mediated inositol 1,3,4,5-tetrakisphosphate receptor, are located on the outer nuclear membrane. Thus, ATP and IP4, depending on external free calcium concentrations, are responsible for filling the nuclear envelope calcium pool. The inositol 1,4,5-trisphosphate receptor is located on the inner nuclear membrane with its ligand binding domain facing toward the nucleoplasm. Likewise, the ryanodine receptor is located on the inner nuclear membrane and its ligand cADP-ribose is generated within the nucleus. A 120 kDa protein fragment of nuclear PLC-gamma1 is stimulated in vivo by epidermal growth factor nuclear signaling coincident with the time course of nuclear membrane epidermal growth factor receptor activation. Stimulated 120 kDa protein fragment interacts with PIKE, a nuclear GTPase, and together they form a complex with PI[3]kinase serving as a module for nuclear PI[3]K stimulation. Thus, the nucleus has its own IP(3) generating system.

The Drosophila Juvenile Hormone Receptor Candidates Methoprene-tolerant (MET) and Germ Cell-expressed (GCE) Utilize a Conserved LIXXL Motif to Bind the FTZ-F1 Nuclear Receptor*

PubMed Central

Bernardo, Travis J.; Dubrovsky, Edward B.

2012-01-01

Juvenile hormone (JH) has been implicated in many developmental processes in holometabolous insects, but its mechanism of signaling remains controversial. We previously found that in Drosophila Schneider 2 cells, the nuclear receptor FTZ-F1 is required for activation of the E75A gene by JH. Here, we utilized insect two-hybrid assays to show that FTZ-F1 interacts with two JH receptor candidates, the bHLH-PAS paralogs MET and GCE, in a JH-dependent manner. These interactions are severely reduced when helix 12 of the FTZ-F1 activation function 2 (AF2) is removed, implicating AF2 as an interacting site. Through homology modeling, we found that MET and GCE possess a C-terminal α-helix featuring a conserved motif LIXXL that represents a novel nuclear receptor (NR) box. Docking simulations supported by two-hybrid experiments revealed that FTZ-F1·MET and FTZ-F1·GCE heterodimer formation involves a typical NR box-AF2 interaction but does not require the canonical charge clamp residues of FTZ-F1 and relies primarily on hydrophobic contacts, including a unique interaction with helix 4. Moreover, we identified paralog-specific features, including a secondary interaction site found only in MET. Our findings suggest that a novel NR box enables MET and GCE to interact JH-dependently with the AF2 of FTZ-F1. PMID:22249180

Tight regulation of a timed nuclear import wave of EKLF by PKCθ and FOE during Pro-E to Baso-E transition.

PubMed

Shyu, Yu-Chiau; Lee, Tung-Liang; Chen, Xin; Hsu, Pang-Hung; Wen, Shau-Ching; Liaw, Yi-Wei; Lu, Chi-Huan; Hsu, Po-Yen; Lu, Mu-Jie; Hwang, JauLang; Tsai, Ming-Daw; Hwang, Ming-Jing; Chen, Jim-Ray; Shen, Che-Kun James

2014-02-24

Erythropoiesis is a highly regulated process during which BFU-E are differentiated into RBCs through CFU-E, Pro-E, PolyCh-E, OrthoCh-E, and reticulocyte stages. Uniquely, most erythroid-specific genes are activated during the Pro-E to Baso-E transition. We show that a wave of nuclear import of the erythroid-specific transcription factor EKLF occurs during the Pro-E to Baso-E transition. We further demonstrate that this wave results from a series of finely tuned events, including timed activation of PKCθ, phosphorylation of EKLF at S68 by P-PKCθ(S676), and sumoylation of EKLF at K74. The latter EKLF modifications modulate its interactions with a cytoplasmic ankyrin-repeat-protein FOE and importinβ1, respectively. The role of FOE in the control of EKLF nuclear import is further supported by analysis of the subcellular distribution patterns of EKLF in FOE-knockout mice. This study reveals the regulatory mechanisms of the nuclear import of EKLF, which may also be utilized in the nuclear import of other factors. Copyright © 2014 Elsevier Inc. All rights reserved.

Modification of nuclear PML protein by SUMO-1 regulates Fas-induced apoptosis in rheumatoid arthritis synovial fibroblasts

PubMed Central

Meinecke, Ingmar; Cinski, Antje; Baier, Anja; Peters, Marvin A.; Dankbar, Berno; Wille, Aline; Drynda, Andreas; Mendoza, Heidi; Gay, Renate E.; Hay, Ronald T.; Ink, Barbara; Gay, Steffen; Pap, Thomas

2007-01-01

The small ubiquitin-like modifier (SUMO)-1 is an important posttranslational regulator of different signaling pathways and involved in the formation of promyelocytic leukemia (PML) protein nuclear bodies (NBs). Overexpression of SUMO-1 has been associated with alterations in apoptosis, but the underlying mechanisms and their relevance for human diseases are not clear. Here, we show that the increased expression of SUMO-1 in rheumatoid arthritis (RA) synovial fibroblasts (SFs) contributes to the resistance of these cells against Fas-induced apoptosis through increased SUMOylation of nuclear PML protein and increased recruitment of the transcriptional repressor DAXX to PML NBs. We also show that the nuclear SUMO-protease SENP1, which is found at lower levels in RA SFs, can revert the apoptosis-inhibiting effects of SUMO-1 by releasing DAXX from PML NBs. Our findings indicate that in RA SFs overexpression of SENP1 can alter the SUMO-1-mediated recruitment of DAXX to PML NBs, thus influencing the proapoptotic effects of DAXX. Accumulation of DAXX in PML NBs by SUMO-1 may, therefore, contribute to the pathogenesis of inflammatory disorders. PMID:17360386

The Orphan Nuclear Receptor TLX Is an Enhancer of STAT1-Mediated Transcription and Immunity to Toxoplasma gondii.

PubMed

Beiting, Daniel P; Hidano, Shinya; Baggs, Julie E; Geskes, Jeanne M; Fang, Qun; Wherry, E John; Hunter, Christopher A; Roos, David S; Cherry, Sara

2015-07-01

The protozoan parasite, Toxoplasma, like many intracellular pathogens, suppresses interferon gamma (IFN-γ)-induced signal transducer and activator of transcription 1 (STAT1) activity. We exploited this well-defined host-pathogen interaction as the basis for a high-throughput screen, identifying nine transcription factors that enhance STAT1 function in the nucleus, including the orphan nuclear hormone receptor TLX. Expression profiling revealed that upon IFN-γ treatment TLX enhances the output of a subset of IFN-γ target genes, which we found is dependent on TLX binding at those loci. Moreover, infection of TLX deficient mice with the intracellular parasite Toxoplasma results in impaired production of the STAT1-dependent cytokine interleukin-12 by dendritic cells and increased parasite burden in the brain during chronic infection. These results demonstrate a previously unrecognized role for this orphan nuclear hormone receptor in regulating STAT1 signaling and host defense and reveal that STAT1 activity can be modulated in a context-specific manner by such "modifiers."

Fatty acid amide hydrolase (FAAH) inhibition enhances memory acquisition through activation of PPAR-α nuclear receptors

PubMed Central

Mazzola, Carmen; Medalie, Julie; Scherma, Maria; Panlilio, Leigh V.; Solinas, Marcello; Tanda, Gianluigi; Drago, Filippo; Cadet, Jean Lud; Goldberg, Steven R.; Yasar, Sevil

2009-01-01

Inhibitors of fatty acid amide hydrolase (FAAH) increase endogenous levels of anandamide (a cannabinoid CB1-receptor ligand) and oleoylethanolamide and palmitoylethanolamide (OEA and PEA, ligands for α-type peroxisome proliferator-activated nuclear receptors, PPAR-α) when and where they are naturally released in the brain. Using a passive-avoidance task in rats, we found that memory acquisition was enhanced by the FAAH inhibitor URB597 or by the PPAR-α agonist WY14643, and these enhancements were blocked by the PPAR-α antagonist MK886. These findings demonstrate novel mechanisms for memory enhancement by activation of PPAR-α, either directly by administering a PPAR-α agonist or indirectly by administering a FAAH inhibitor. PMID:19403796

SENP1-mediated NEMO deSUMOylation in adipocytes limits inflammatory responses and type-1 diabetes progression

PubMed Central

Shao, Lan; Zhou, Huanjiao Jenny; Zhang, Haifeng; Qin, Lingfeng; Hwa, John; Yun, Zhong; Ji, Weidong; Min, Wang

2015-01-01

Adipocyte dysfunction correlates with the development of diabetes. Here we show that mice with a adipocyte-specific deletion of the SUMO-specific protease SENP1 gene develop symptoms of type-1 diabetes mellitus (T1DM), including hyperglycaemia and glucose intolerance with mild insulin resistance. Peri-pancreatic adipocytes from SENP1-deficient mice exhibit heightened NF-κB activity and production of proinflammatory cytokines, which induce CCL5 expression in adjacent pancreatic islets and direct cytotoxic effects on pancreatic islets. Mechanistic studies show that SENP1 deletion in adipocytes enhances SUMOylation of the NF-κB essential molecule, NEMO, at lysine 277/309, leading to increased NF-κB activity, cytokine production and pancreatic inflammation. We further show that NF-κB inhibitors could inhibit pre-diabetic cytokine production, β-cell damages and ameliorate the T1DM phenotype in SENP1-deficient mice. Feeding a high-fat diet augments both type-1 and type-2 diabetes phenotypes in SENP1-deficient mice, consistent with the effects on adipocyte-derived NF-κB and cytokine signalling. Our study reveals previously unrecognized mechanism regulating the onset and progression of T1DM associated with adipocyte dysfunction. PMID:26596471

SENP1-mediated NEMO deSUMOylation in adipocytes limits inflammatory responses and type-1 diabetes progression.

PubMed

Shao, Lan; Zhou, Huanjiao Jenny; Zhang, Haifeng; Qin, Lingfeng; Hwa, John; Yun, Zhong; Ji, Weidong; Min, Wang

2015-11-24

Adipocyte dysfunction correlates with the development of diabetes. Here we show that mice with a adipocyte-specific deletion of the SUMO-specific protease SENP1 gene develop symptoms of type-1 diabetes mellitus (T1DM), including hyperglycaemia and glucose intolerance with mild insulin resistance. Peri-pancreatic adipocytes from SENP1-deficient mice exhibit heightened NF-κB activity and production of proinflammatory cytokines, which induce CCL5 expression in adjacent pancreatic islets and direct cytotoxic effects on pancreatic islets. Mechanistic studies show that SENP1 deletion in adipocytes enhances SUMOylation of the NF-κB essential molecule, NEMO, at lysine 277/309, leading to increased NF-κB activity, cytokine production and pancreatic inflammation. We further show that NF-κB inhibitors could inhibit pre-diabetic cytokine production, β-cell damages and ameliorate the T1DM phenotype in SENP1-deficient mice. Feeding a high-fat diet augments both type-1 and type-2 diabetes phenotypes in SENP1-deficient mice, consistent with the effects on adipocyte-derived NF-κB and cytokine signalling. Our study reveals previously unrecognized mechanism regulating the onset and progression of T1DM associated with adipocyte dysfunction.

Deficiency of the NR4A Orphan Nuclear Receptor NOR1 attenuates Neointima Formation Following Vascular Injury

PubMed Central

Nomiyama, Takashi; Zhao, Yue; Gizard, Florence; Findeisen, Hannes M.; Heywood, Elizabeth B.; Jones, Karrie L.; Conneely, Orla M.; Bruemmer, Dennis

2009-01-01

Background The neuron-derived orphan receptor-1 (NOR1) belongs to the evolutionary highly conserved and most ancient NR4A subfamily of the nuclear hormone receptor superfamily. Members of this subfamily function as early response genes regulating key cellular processes including proliferation, differentiation, and survival. Although NOR1 has previously been demonstrated to be required for smooth muscle cell (SMC) proliferation in vitro, the role of this nuclear receptor for the proliferative response underlying neointima formation and target genes trans-activated by NOR1 remain to be defined. Methods and Results Using a model of guide wire-induced arterial injury, we demonstrate decreased neointima formation in NOR1-/- mice compared to wildtype mice. In vitro, NOR1-deficient SMC exhibit decreased proliferation due to a G1→S phase arrest of the cell cycle and increased apoptosis in response to serum deprivation. NOR1-deficiency alters phosphorylation of the retinoblastoma protein by preventing mitogen-induced cyclin D1 and D2 expression. Conversely, overexpression of NOR1 induces cyclin D1 expression and the transcriptional activity of the cyclin D1 promoter in transient reporter assays. Gel shift and chromatin immunoprecipitation assays identified a putative response element for NR4A receptors in the cyclin D1 promoter, to which NOR1 is recruited in response to mitogenic stimulation. Finally, we provide evidence that these observations are applicable in vivo by demonstrating decreased cyclin D1 expression during neointima formation in NOR1-deficient mice. Conclusions These experiments characterize cyclin D1 as a NOR1-regulated target gene in SMC and demonstrate that NOR1 deficiency decreases neointima formation in response to vascular injury. PMID:19153266

Research resource: Tissue-specific transcriptomics and cistromics of nuclear receptor signaling: a web research resource.

PubMed

Ochsner, Scott A; Watkins, Christopher M; LaGrone, Benjamin S; Steffen, David L; McKenna, Neil J

2010-10-01

Nuclear receptors (NRs) are ligand-regulated transcription factors that recruit coregulators and other transcription factors to gene promoters to effect regulation of tissue-specific transcriptomes. The prodigious rate at which the NR signaling field has generated high content gene expression and, more recently, genome-wide location analysis datasets has not been matched by a committed effort to archiving this information for routine access by bench and clinical scientists. As a first step towards this goal, we searched the MEDLINE database for studies, which referenced either expression microarray and/or genome-wide location analysis datasets in which a NR or NR ligand was an experimental variable. A total of 1122 studies encompassing 325 unique organs, tissues, primary cells, and cell lines, 35 NRs, and 91 NR ligands were retrieved and annotated. The data were incorporated into a new section of the Nuclear Receptor Signaling Atlas Molecule Pages, Transcriptomics and Cistromics, for which we designed an intuitive, freely accessible user interface to browse the studies. Each study links to an abstract, the MEDLINE record, and, where available, Gene Expression Omnibus and ArrayExpress records. The resource will be updated on a regular basis to provide a current and comprehensive entrez into the sum of transcriptomic and cistromic research in this field.

Nuclear Receptor Rev-erb Alpha (Nr1d1) Functions in Concert with Nr2e3 to Regulate Transcriptional Networks in the Retina

PubMed Central

Mollema, Nissa J.; Yuan, Yang; Jelcick, Austin S.; Sachs, Andrew J.; von Alpen, Désirée; Schorderet, Daniel; Escher, Pascal; Haider, Neena B.

2011-01-01

The majority of diseases in the retina are caused by genetic mutations affecting the development and function of photoreceptor cells. The transcriptional networks directing these processes are regulated by genes such as nuclear hormone receptors. The nuclear hormone receptor gene Rev-erb alpha/Nr1d1 has been widely studied for its role in the circadian cycle and cell metabolism, however its role in the retina is unknown. In order to understand the role of Rev-erb alpha/Nr1d1 in the retina, we evaluated the effects of loss of Nr1d1 to the developing retina and its co-regulation with the photoreceptor-specific nuclear receptor gene Nr2e3 in the developing and mature retina. Knock-down of Nr1d1 expression in the developing retina results in pan-retinal spotting and reduced retinal function by electroretinogram. Our studies show that NR1D1 protein is co-expressed with NR2E3 in the outer neuroblastic layer of the developing mouse retina. In the adult retina, NR1D1 is expressed in the ganglion cell layer and is co-expressed with NR2E3 in the outer nuclear layer, within rods and cones. Several genes co-targeted by NR2E3 and NR1D1 were identified that include: Nr2c1, Recoverin, Rgr, Rarres2, Pde8a, and Nupr1. We examined the cyclic expression of Nr1d1 and Nr2e3 over a twenty-four hour period and observed that both nuclear receptors cycle in a similar manner. Taken together, these studies reveal a novel role for Nr1d1, in conjunction with its cofactor Nr2e3, in regulating transcriptional networks critical for photoreceptor development and function. PMID:21408158

Targeting of the Nuclear Receptor Coactivator Isoform DELTA3AIB1 in Breast Cancer

DTIC Science & Technology

2007-03-01

lab showed that the downregulation of overall levels of AIB1 plus ∆3AIB1, using a regulatable AIB1 directed ribozyme , resulted in reduced tumor...overall levels of AIB1 plus ∆3AIB1, using a regulatable AIB1 directed ribozyme , resulted in reduced tumor growth in vivo. Overall, these data indicate a...Reiter R, Powers C, Wellstein A, Riegel AT. Ribozyme targeting shows that the nuclear receptor coactivator AIB1 is a rate-limiting factor for estrogen

Nuclear receptor co-activators and HER-2/neu are upregulated in breast cancer patients during neo-adjuvant treatment with aromatase inhibitors

PubMed Central

Flågeng, M Hauglid; Haugan Moi, L L; Dixon, J M; Geisler, J; Lien, E A; Miller, W R; Lønning, P E; Mellgren, G

2009-01-01

Background: Acquired resistance to endocrine therapy in breast cancer is poorly understood. Characterisation of the molecular response to aromatase inhibitors in breast cancer tissue may provide important information regarding development of oestrogen hypersensitivity. Methods: We examined the expression levels of nuclear receptor co-regulators, the orphan nuclear receptor liver receptor homologue-1 and HER-2/neu growth factor receptor using real-time RT-PCR before and after 13–16 weeks of primary medical treatment with the aromatase inhibitors anastrozole or letrozole. Results: mRNA expression of the steroid receptor co-activator 1 (SRC-1) and peroxisome-proliferator-activated receptor γ co-activator-1α (PGC-1α) was correlated (P=0.002), and both co-activators increased during treatment in the patient group as a whole (P=0.008 and P=0.032, respectively), as well as in the subgroup of patients achieving an objective treatment response (P=0.002 and P=0.006). Although we recorded no significant change in SRC-3/amplified in breast cancer 1 level, the expression correlated positively to the change of SRC-1 (P=0.002). Notably, we recorded an increase in HER-2/neu levels during therapy in the total patient group (18 out of 26; P=0.016), but in particular among responders (15 out of 21; P=0.008). Conclusion: Our results show an upregulation of co-activator mRNA and HER-2/neu during treatment with aromatase inhibitors. These mechanisms may represent an early adaption of the breast cancer cells to oestrogen deprivation in vivo. PMID:19755984

Nuclear Receptors in Drug Metabolism, Drug Response and Drug Interactions

PubMed Central

Prakash, Chandra; Zuniga, Baltazar; Song, Chung Seog; Jiang, Shoulei; Cropper, Jodie; Park, Sulgi; Chatterjee, Bandana

2016-01-01

Orally delivered small-molecule therapeutics are metabolized in the liver and intestine by phase I and phase II drug-metabolizing enzymes (DMEs), and transport proteins coordinate drug influx (phase 0) and drug/drug-metabolite efflux (phase III). Genes involved in drug metabolism and disposition are induced by xenobiotic-activated nuclear receptors (NRs), i.e. PXR (pregnane X receptor) and CAR (constitutive androstane receptor), and by the 1α, 25-dihydroxy vitamin D3-activated vitamin D receptor (VDR), due to transactivation of xenobiotic-response elements (XREs) present in phase 0-III genes. Additional NRs, like HNF4-α, FXR, LXR-α play important roles in drug metabolism in certain settings, such as in relation to cholesterol and bile acid metabolism. The phase I enzymes CYP3A4/A5, CYP2D6, CYP2B6, CYP2C9, CYP2C19, CYP1A2, CYP2C8, CYP2A6, CYP2J2, and CYP2E1 metabolize >90% of all prescription drugs, and phase II conjugation of hydrophilic functional groups (with/without phase I modification) facilitates drug clearance. The conjugation step is mediated by broad-specificity transferases like UGTs, SULTs, GSTs. This review delves into our current understanding of PXR/CAR/VDR-mediated regulation of DME and transporter expression, as well as effects of single nucleotide polymorphism (SNP) and epigenome (specified by promoter methylation, histone modification, microRNAs, long non coding RNAs) on the expression of PXR/CAR/VDR and phase 0-III mediators, and their impacts on variable drug response. Therapeutic agents that target epigenetic regulation and the molecular basis and consequences (overdosing, underdosing, or beneficial outcome) of drug-drug/drug-food/drug-herb interactions are also discussed. Precision medicine requires understanding of a drug’s impact on DME and transporter activity and their NR-regulated expression in order to achieve optimal drug efficacy without adverse drug reactions. In future drug screening, new tools such as humanized mouse models and

Expression profiling of nuclear receptors in breast cancer identifies TLX as a mediator of growth and invasion in triple-negative breast cancer

PubMed Central

Remenyi, Judit; Banerji, Christopher R.S.; Lai, Chun-Fui; Periyasamy, Manikandan; Lombardo, Ylenia; Busonero, Claudia; Ottaviani, Silvia; Passey, Alun; Quinlan, Philip R.; Purdie, Colin A.; Jordan, Lee B.; Thompson, Alastair M.; Finn, Richard S.; Rueda, Oscar M.; Caldas, Carlos; Gil, Jesus; Coombes, R. Charles; Fuller-Pace, Frances V.; Teschendorff, Andrew E.; Buluwela, Laki; Ali, Simak

2015-01-01

The Nuclear Receptor (NR) superfamily of transcription factors comprises 48 members, several of which have been implicated in breast cancer. Most important is estrogen receptor-α (ERα), which is a key therapeutic target. ERα action is facilitated by co-operativity with other NR and there is evidence that ERα function may be recapitulated by other NRs in ERα-negative breast cancer. In order to examine the inter-relationships between nuclear receptors, and to obtain evidence for previously unsuspected roles for any NRs, we undertook quantitative RT-PCR and bioinformatics analysis to examine their expression in breast cancer. While most NRs were expressed, bioinformatic analyses differentiated tumours into distinct prognostic groups that were validated by analyzing public microarray data sets. Although ERα and progesterone receptor were dominant in distinguishing prognostic groups, other NR strengthened these groups. Clustering analysis identified several family members with potential importance in breast cancer. Specifically, RORγ is identified as being co-expressed with ERα, whilst several NRs are preferentially expressed in ERα-negative disease, with TLX expression being prognostic in this subtype. Functional studies demonstrated the importance of TLX in regulating growth and invasion in ERα-negative breast cancer cells. PMID:26280373

Expression profiling of nuclear receptors in breast cancer identifies TLX as a mediator of growth and invasion in triple-negative breast cancer.

PubMed

Lin, Meng-Lay; Patel, Hetal; Remenyi, Judit; Banerji, Christopher R S; Lai, Chun-Fui; Periyasamy, Manikandan; Lombardo, Ylenia; Busonero, Claudia; Ottaviani, Silvia; Passey, Alun; Quinlan, Philip R; Purdie, Colin A; Jordan, Lee B; Thompson, Alastair M; Finn, Richard S; Rueda, Oscar M; Caldas, Carlos; Gil, Jesus; Coombes, R Charles; Fuller-Pace, Frances V; Teschendorff, Andrew E; Buluwela, Laki; Ali, Simak

2015-08-28

The Nuclear Receptor (NR) superfamily of transcription factors comprises 48 members, several of which have been implicated in breast cancer. Most important is estrogen receptor-α (ERα), which is a key therapeutic target. ERα action is facilitated by co-operativity with other NR and there is evidence that ERα function may be recapitulated by other NRs in ERα-negative breast cancer. In order to examine the inter-relationships between nuclear receptors, and to obtain evidence for previously unsuspected roles for any NRs, we undertook quantitative RT-PCR and bioinformatics analysis to examine their expression in breast cancer. While most NRs were expressed, bioinformatic analyses differentiated tumours into distinct prognostic groups that were validated by analyzing public microarray data sets. Although ERα and progesterone receptor were dominant in distinguishing prognostic groups, other NR strengthened these groups. Clustering analysis identified several family members with potential importance in breast cancer. Specifically, RORγ is identified as being co-expressed with ERα, whilst several NRs are preferentially expressed in ERα-negative disease, with TLX expression being prognostic in this subtype. Functional studies demonstrated the importance of TLX in regulating growth and invasion in ERα-negative breast cancer cells.

Structural and functional evidences for the interactions between nuclear hormone receptors and endocrine disruptors at low doses.

PubMed

Balaguer, Patrick; Delfosse, Vanessa; Grimaldi, Marina; Bourguet, William

Endocrine-disrupting chemicals (EDCs) represent a broad class of exogenous substances that cause adverse effects in the endocrine system mainly by interacting with nuclear hormone receptors (NRs). Humans are generally exposed to low doses of pollutants, and current researches aim at deciphering the mechanisms accounting for the health impact of EDCs at environmental concentrations. Our correlative analysis of structural, interaction and cell-based data has revealed a variety of, sometimes unexpected, binding modes, reflecting a wide range of EDC affinities and specificities. Here, we present a few representative examples to illustrate various means by which EDCs achieve high-affinity binding to NRs. These examples include the binding of the mycoestrogen α-zearalanol to estrogen receptors, the covalent interaction of organotins with the retinoid X- and peroxisome proliferator-activated receptors, and the cooperative binding of two chemicals to the pregnane X receptor. We also discuss some hypotheses that could further explain low-concentration effects of EDCs with weaker affinity towards NRs. Copyright © 2017. Published by Elsevier Masson SAS.

SMILE, a new orphan nuclear receptor SHP-interacting protein, regulates SHP-repressed estrogen receptor transactivation.

PubMed

Xie, Yuan-Bin; Lee, Ok-Hee; Nedumaran, Balachandar; Seong, Hyun-A; Lee, Kyeong-Min; Ha, Hyunjung; Lee, In-Kyu; Yun, Yungdae; Choi, Hueng-Sik

2008-12-15

SHP (small heterodimer partner) is a well-known NR (nuclear receptor) co-regulator. In the present study, we have identified a new SHP-interacting protein, termed SMILE (SHP-interacting leucine zipper protein), which was previously designated as ZF (Zhangfei) via a yeast two-hybrid system. We have determined that the SMILE gene generates two isoforms [SMILE-L (long isoform of SMILE) and SMILE-S (short isoform of SMILE)]. Mutational analysis has demonstrated that the SMILE isoforms arise from the alternative usage of initiation codons. We have confirmed the in vivo interaction and co-localization of the SMILE isoforms and SHP. Domain-mapping analysis indicates that the entire N-terminus of SHP and the middle region of SMILE-L are involved in this interaction. Interestingly, the SMILE isoforms counteract the SHP repressive effect on the transactivation of ERs (estrogen receptors) in HEK-293T cells (human embryonic kidney cells expressing the large T-antigen of simian virus 40), but enhance the SHP-repressive effect in MCF-7, T47D and MDA-MB-435 cells. Knockdown of SMILE gene expression using siRNA (small interfering RNA) in MCF-7 cells increases ER-mediated transcriptional activity. Moreover, adenovirus-mediated overexpression of SMILE and SHP down-regulates estrogen-induced mRNA expression of the critical cell-cycle regulator E2F1. Collectively, these results indicate that SMILE isoforms regulate the inhibition of ER transactivation by SHP in a cell-type-specific manner and act as a novel transcriptional co-regulator in ER signalling.

Molecular adaptation and resilience of the insect’s nuclear receptor USP

PubMed Central

2012-01-01

Background The maintenance of biological systems requires plasticity and robustness. The function of the ecdysone receptor, a heterodimer composed of the nuclear receptors ECR (NR1H1) and USP (NR2B4), was maintained in insects despite a dramatic divergence that occurred during the emergence of Mecopterida. This receptor is therefore a good model to study the evolution of plasticity. We tested the hypothesis that selection has shaped the Ligand-Binding Domain (LBD) of USP during evolution of Mecopterida. Results We isolated usp and cox1 in several species of Drosophilidae, Tenebrionidae and Blattaria and estimated non-synonymous/synonymous rate ratios using maximum-likelihood methods and codon-based substitution models. Although the usp sequences were mainly under negative selection, we detected relaxation at residues located on the surface of the LBD within Mecopterida families. Using branch-site models, we also detected changes in selective constraints along three successive branches of the Mecopterida evolution. Residues located at the bottom of the ligand-binding pocket (LBP) underwent strong positive selection during the emergence of Mecopterida. This change is correlated with the acquisition of a large LBP filled by phospholipids that probably allowed the stabilisation of the new Mecopterida structure. Later, when the two subgroups of Mecopterida (Amphiesmenoptera: Lepidoptera, Trichoptera; Antliophora: Diptera, Mecoptera, Siphonaptera) diverged, the same positions became under purifying selection. Similarly, several positions of the heterodimerisation interface experienced positive selection during the emergence of Mecopterida, rapidly followed by a phase of constrained evolution. An enlargement of the heterodimerisation surface is specific for Mecopterida and was associated with a reinforcement of the obligatory partnership between ECR and USP, at the expense of homodimerisation. Conclusions In order to explain the episodic mode of evolution of USP, we

A Review About Lycopene-Induced Nuclear Hormone Receptor Signalling in Inflammation and Lipid Metabolism via still Unknown Endogenous Apo-10´-Lycopenoids.

PubMed

Caris-Veyrat, Catherine; Garcia, Ada L; Reynaud, Eric; Lucas, Renata; Aydemir, Gamze; Rühl, Ralph

2017-10-20

Lycopene is the red pigment in tomatoes and tomato products and is an important dietary carotenoid found in the human organism. Lycopene-isomers, oxidative lycopene metabolites and apo-lycopenoids are found in the food matrix. Lycopene intake derived from tomato consumption is associated with alteration of lipid metabolism and a lower incidence of cardiovascular diseases (CVD). Lycopene is mainly described as a potent antioxidant but novel studies are shifting towards its metabolites and their capacity to mediate nuclear receptor signalling. Di-/tetra-hydro-derivatives of apo-10´-lycopenoic acid and apo-15´-lycopenoic acids are potential novel endogenous mammalian lycopene metabolites which may act as ligands for nuclear hormone mediated activation and signalling. In this review, we postulate that complex lycopene metabolism results in various lycopene metabolites which have the ability to mediate transactivation of various nuclear hormone receptors like RARs, RXRs and PPARs. A new mechanistic explanation of how tomato consumption could positively modulate inflammation and lipid metabolism is discussed.

Therapeutic androgen receptor ligands

PubMed Central

Allan, George F.; Sui, Zhihua

2003-01-01

In the past several years, the concept of tissue-selective nuclear receptor ligands has emerged. This concept has come to fruition with estrogens, with the successful marketing of drugs such as raloxifene. The discovery of raloxifene and other selective estrogen receptor modulators (SERMs) has raised the possibility of generating selective compounds for other pathways, including androgens (that is, selective androgen receptor modulators, or SARMs). PMID:16604181

Possible Involvement of Tight Junctions, Extracellular Matrix and Nuclear Receptors in Epithelial Differentiation

PubMed Central

Ichikawa-Tomikawa, Naoki; Sugimoto, Kotaro; Satohisa, Seiro; Nishiura, Keisuke; Chiba, Hideki

2011-01-01

Tight junctions are intercellular junctions localized at the most apical end of the lateral plasma membrane. They consist of four kinds of transmembrane proteins (occludin, claudins, junctional adhesion molecules, and tricellulin) and huge numbers of scaffolding proteins and contribute to the paracellular barrier and fence function. The mutation and deletion of these proteins impair the functions of tight junctions and cause various human diseases. In this paper, we provide an overview of recent studies on transmembrane proteins of tight junctions and highlight the functional significance of tight junctions, extracellular matrix, and nuclear receptors in epithelial differentiation. PMID:22162632

Does bilirubin prevent hepatic steatosis through activation of the PPARα nuclear receptor?

PubMed

Hinds, Terry D; Adeosun, Samuel O; Alamodi, Abdulhadi A; Stec, David E

2016-10-01

Several large population studies have demonstrated a negative correlation between serum bilirubin levels and the development of obesity, hepatic steatosis, and cardiovascular disease. Despite the strong correlative data demonstrating the protective role of bilirubin, the mechanism by which bilirubin can protect against these pathologies remains unknown. Bilirubin has long been known as a powerful antioxidant and also has anti-inflammatory actions, each of which may contribute to the protection afforded by increased levels. We have recently described a novel function of bilirubin as a ligand for the peroxisome proliferator-activated receptor-alpha (PPARα), which we show specifically binds to the nuclear receptor. Bilirubin may function as a selective PPAR modulator (SPPARM) to control lipid accumulation and blood glucose. However, it is not known to what degree bilirubin activation of PPARα is responsible for the protection afforded to reduce hepatic steatosis. We hypothesize that bilirubin, acting as a novel SPPARM, increases hepatic fatty acid metabolism through a PPARα-dependent mechanism which reduces hepatic lipid accumulation and protects against hepatic steatosis and non-alcoholic fatty liver disease (NAFLD). Copyright © 2016 Elsevier Ltd. All rights reserved.

Regulation of the ovarian inflammatory response at ovulation by nuclear progesterone receptor.

PubMed

Akison, Lisa K; Robertson, Sarah A; Gonzalez, Macarena B; Richards, JoAnne S; Smith, C Wayne; Russell, Darryl L; Robker, Rebecca L

2018-06-01

The nuclear progesterone receptor (PGR) transcription factor is essential for ovulation; however, the exact mechanisms by which PGR controls ovulation are not known. The aim of this study was to determine whether PGR regulates inflammatory mediators in the ovary. Ovaries from mice lacking PGR (PRKO) and heterozygous PR+/- littermates were subjected to microarray analysis of a large panel of inflammatory genes. Immune cell subsets were detected by gene expression; and neutrophils by immunohistochemistry and chemotaxis assay. PRKO ovaries exhibited dysregulated expression of vasodilator (Edn1), cytokine (Il-6, Tgfb1), adhesion receptor (Cd34), apoptotic factor (Bax) and transcription factors (Nfkb2, Socs1, Stat3). Ptgs2 was also reduced in PRKO ovaries, but mRNA and protein were not different in granulosa cells. There were reduced neutrophils in ovaries of PRKO mice at ovulation; however, chemotaxis assays showed PRKO neutrophils migrate normally and that PRKO ovarian extracts exhibit chemotactic properties in vitro. Specific inflammatory mediators are altered in the ovaries of PRKO mice indicating that progesterone regulates features of inflammation at ovulation. © 2018 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Regulation of behaviour by the nuclear receptor TLX.

PubMed

O'Leary, J D; O'Leary, O F; Cryan, J F; Nolan, Y M

2018-03-01

The orphan nuclear receptor Tlx (Nr2e1) is a key regulator of both embryonic and adult hippocampal neurogenesis. Several different mouse models have been developed which target Tlx in vivo including spontaneous deletion models (from birth) and targeted and conditional knockouts. Although some conflicting findings have been reported, for the most part studies have demonstrated that Tlx is important in regulating processes that underlie neurogenesis, spatial learning, anxiety-like behaviour and interestingly, aggression. More recent data have demonstrated that disrupting Tlx during early life induces hyperactivity and that Tlx plays a role in emotional regulation. Moreover, there are sex- and age-related differences in some behaviours in Tlx knockout mice during adolescence and adulthood. Here, we discuss the role of Tlx in motor-, cognitive-, aggressive- and anxiety-related behaviours during adolescence and adulthood. We examine current evidence which provides insight into Tlx during neurodevelopment, and offer our thoughts on the function of Tlx in brain and behaviour. We further hypothesize that Tlx is a key target in understanding the emergence of neurobiological disorders during adolescence and early adulthood. © 2016 John Wiley & Sons Ltd and International Behavioural and Neural Genetics Society.

Effect of marine collagen peptides on markers of metabolic nuclear receptors in type 2 diabetic patients with/without hypertension.

PubMed

Zhu, Cui-Feng; Li, Guan-Zhi; Peng, Hong-Bin; Zhang, Fan; Chen, Yun; Li, Yong

2010-04-01

To explore Effects of marine collagen peptides (MCPs) on markers of metablic nuclear receptors, i.e peroxisome proliferator-activated receptor (PPARs), liver X receptor (LXRs) and farnesoid X receptor (FXRs) in type 2 diabetic patients with/without hypertension. METHOD Study population consisted of 200 type 2 diabetic patients with/without hypertension and 50 healthy subjects, all of whom were randomly assigned to MCPs-treated diabetics (n = 50), placebo-treated diabetics (n = 50), MCPs-treated diabetics with hypertension (n=50), placebo-treated diabetics with hypertension (n = 50), and healthy controls (n = 50). MCPs or placebo (water-soluble starch) were given daily before breakfast and bedtime over three months. Levels of free fatty acid, cytochrome P450, leptin, resistin, adiponectin, bradykinin, NO, and Prostacyclin were determined before intervention, and 1.5 months, and 3 months after intervention. Hypoglycemia and the endpoint events during the study were recorded and compared among the study groups. At the end of the study period, MCPs-treated patients showed marked improvement compared with patients receiving placebo. The protection exerted by MCPs seemed more profound in diabetics than in diabetics with hypertension. In particular, after MCPs intervention, levels of free fatty acid, hs-CRP, resistin, Prostacyclin decreased significantly in diabetics and tended to decrease in diabetic and hypertensive patients whereas levels of cytochrome P450, leptin, NO tended to decrease in diabetics with/without hypertension. Meanwhile, levels of adiponectin and bradykinin rose markedly in diabetics following MCPs administration. MCPs could offer protection against diabetes and hypertension by affecting levels of molecules involved in diabetic and hypertensive pathogenesis. Regulation on metabolic nuclear receptors by MCPs may be the possible underlying mechanism for its observed effects in the study. Further study into its action may shed light on development of new

The Orphan Nuclear Receptor TLX Is an Enhancer of STAT1-Mediated Transcription and Immunity to Toxoplasma gondii

PubMed Central

Beiting, Daniel P.; Hidano, Shinya; Baggs, Julie E.; Geskes, Jeanne M.; Fang, Qun; Wherry, E. John; Hunter, Christopher A.; Roos, David S.; Cherry, Sara

2015-01-01

The protozoan parasite, Toxoplasma, like many intracellular pathogens, suppresses interferon gamma (IFN-γ)-induced signal transducer and activator of transcription 1 (STAT1) activity. We exploited this well-defined host–pathogen interaction as the basis for a high-throughput screen, identifying nine transcription factors that enhance STAT1 function in the nucleus, including the orphan nuclear hormone receptor TLX. Expression profiling revealed that upon IFN-γ treatment TLX enhances the output of a subset of IFN-γ target genes, which we found is dependent on TLX binding at those loci. Moreover, infection of TLX deficient mice with the intracellular parasite Toxoplasma results in impaired production of the STAT1-dependent cytokine interleukin-12 by dendritic cells and increased parasite burden in the brain during chronic infection. These results demonstrate a previously unrecognized role for this orphan nuclear hormone receptor in regulating STAT1 signaling and host defense and reveal that STAT1 activity can be modulated in a context-specific manner by such “modifiers.” PMID:26196739

The nuclear hormone receptor SEX-1 is an X-chromosome signal that determines nematode sex.

PubMed

Carmi, I; Kopczynski, J B; Meyer, B J

1998-11-12

Organisms in many phyla determine sexual fate by distinguishing one X chromosome from two. Here we use the model organism Caenorhabditis elegans to dissect such an X-chromosome-counting mechanism in molecular detail. In this nematode, several genes on the X chromosome called X signal elements communicate X-chromosome dose by controlling the activity of the sex-determination gene xol-1. xol-1 specifies male (XO) fate when active and hermaphrodite (XX) fate when inactive. The only X signal element described so far represses xol-1 post-transcriptionally, but xol-1 is repressed in XX animals by transcriptional and post-transcriptional mechanisms. Here we identify a nuclear-hormone-receptor homologue, SEX-1, that regulates the transcription of xol-1. We show that sex-1 is vital to X-chromosome counting: changing sex-1 gene dose in XX or XO embryos causes sexual transformation and death from inadequate dosage compensation (the hermaphrodite-specific process that equalizes X-gene expression between the sexes). The SEX-1 protein acts directly on xol-1, associating with its promoter in vivo and repressing xol-1 transcription in XX embryos. Thus, xol-1 is the direct molecular target of the primary sex-determination signal, and the dose of a nuclear hormone receptor helps to communicate X-chromosome number to determine nematode sex.

Coexpression of Nuclear Receptors and Histone Methylation Modifying Genes in the Testis: Implications for Endocrine Disruptor Modes of Action

PubMed Central

Anderson, Alison M.; Carter, Kim W.; Anderson, Denise; Wise, Michael J.

2012-01-01

Background Endocrine disruptor chemicals elicit adverse health effects by perturbing nuclear receptor signalling systems. It has been speculated that these compounds may also perturb epigenetic mechanisms and thus contribute to the early origin of adult onset disease. We hypothesised that histone methylation may be a component of the epigenome that is susceptible to perturbation. We used coexpression analysis of publicly available data to investigate the combinatorial actions of nuclear receptors and genes involved in histone methylation in normal testis and when faced with endocrine disruptor compounds. Methodology/Principal Findings The expression patterns of a set of genes were profiled across testis tissue in human, rat and mouse, plus control and exposed samples from four toxicity experiments in the rat. Our results indicate that histone methylation events are a more general component of nuclear receptor mediated transcriptional regulation in the testis than previously appreciated. Coexpression patterns support the role of a gatekeeper mechanism involving the histone methylation modifiers Kdm1, Prdm2, and Ehmt1 and indicate that this mechanism is a common determinant of transcriptional integrity for genes critical to diverse physiological endpoints relevant to endocrine disruption. Coexpression patterns following exposure to vinclozolin and dibutyl phthalate suggest that coactivity of the demethylase Kdm1 in particular warrants further investigation in relation to endocrine disruptor mode of action. Conclusions/Significance This study provides proof of concept that a bioinformatics approach that profiles genes related to a specific hypothesis across multiple biological settings can provide powerful insight into coregulatory activity that would be difficult to discern at an individual experiment level or by traditional differential expression analysis methods. PMID:22496781

Estetrol, a Fetal Selective Estrogen Receptor Modulator, Acts on the Vagina of Mice through Nuclear Estrogen Receptor α Activation.

PubMed

Benoit, Thibaut; Valera, Marie-Cecile; Fontaine, Coralie; Buscato, Melissa; Lenfant, Francoise; Raymond-Letron, Isabelle; Tremollieres, Florence; Soulie, Michel; Foidart, Jean-Michel; Game, Xavier; Arnal, Jean-Francois

2017-11-01

The genitourinary syndrome of menopause has a negative impact on quality of life of postmenopausal women. The treatment of vulvovaginal atrophy includes administration of estrogens. However, oral estrogen treatment is controversial because of its potential risks on venous thrombosis and breast cancer. Estetrol (E4) is a natural estrogen synthesized exclusively during pregnancy by the human fetal liver and initially considered as a weak estrogen. However, E4 was recently evaluated in phase 1 to 2 clinical studies and found to act as an oral contraceptive in combination with a progestin, without increasing the level of coagulation factors. We recently showed that E4 stimulates uterine epithelial proliferation through nuclear estrogen receptor (ER) α, but failed to elicit endothelial responses. Herein, we first evaluated the morphological and functional impacts of E4 on the vagina of ovariectomized mice, and we determined the molecular mechanism mediating these effects. Vaginal epithelial proliferation and lubrication after stimulation were found to increase after E4 chronic treatment. Using a combination of pharmacological and genetic approaches, we demonstrated that these E4 effects on the vagina are mediated by nuclear ERα activation. Altogether, we demonstrate that the selective activation of nuclear ERα is both necessary and sufficient to elicit functional and structural effects on the vagina, and therefore E4 appears promising as a therapeutic option to improve vulvovaginal atrophy. Copyright © 2017 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

Rho-kinase signaling controls nucleocytoplasmic shuttling of class IIa Histone Deacetylase (HDAC7) and transcriptional activation of orphan nuclear receptor NR4A1

DOE Office of Scientific and Technical Information (OSTI.GOV)

Compagnucci, Claudia; Barresi, Sabina; Petrini, Stefania

2015-04-03

Rho-kinase (ROCK) has been well documented to play a key role in RhoA-induced actin remodeling. ROCK activation results in myosin light chain (MLC) phosphorylation either by direct action on MLC kinase (MLCK) or by inhibition of MLC phosphatase (MLCP), modulating actin–myosin contraction. We found that inhibition of the ROCK pathway in induced pluripotent stem cells, leads to nuclear export of HDAC7 and transcriptional activation of the orphan nuclear receptor NR4A1 while in cells with constitutive ROCK hyperactivity due to loss of function of the RhoGTPase activating protein Oligophrenin-1 (OPHN1), the orphan nuclear receptor NR4A1 is downregulated. Our study identify amore » new target of ROCK signaling via myosin phosphatase subunit (MYPT1) and Histone Deacetylase (HDAC7) at the nuclear level and provide new insights in the cellular functions of ROCK. - Highlights: • ROCK regulates nucleocytoplasmic shuttling of HDAC7 via phosphorylation of MYPT1. • Nuclear export of HDAC7 and upregulation of NR4A1 occurs with low ROCK activity. • High levels of ROCK activity due to OPHN1 loss of function downregulate NR4A1.« less

Changes in nuclear receptor corepressor RIP140 do not influence mitochondrial content in the cortex.

PubMed

Herbst, Eric A F; Bonen, Arend; Holloway, Graham P

2015-10-01

Changes in nuclear receptor interacting protein 140 (RIP140) influences mitochondrial content in skeletal muscle; however, the translation of these findings to the brain has not been investigated. The present study examined the impact of overexpressing and ablating RIP140 on mitochondrial content in muscle and the cortex through examining mRNA, mtDNA, and mitochondrial protein content. Our results show that changes in RIP140 expression significantly alters markers of mitochondrial content in skeletal muscle but not the brain.

Molecular and Structural Traits of Insulin Receptor Substrate 1/LC3 Nuclear Structures and Their Role in Autophagy Control and Tumor Cell Survival.

PubMed

Lassak, Adam; Dean, Mathew; Wyczechowska, Dorota; Wilk, Anna; Marrero, Luis; Trillo-Tinoco, Jimena; Boulares, A Hamid; Sarkaria, Jann N; Del Valle, Luis; Peruzzi, Francesca; Ochoa, Augusto; Reiss, Krzysztof

2018-05-15

Insulin receptor substrate 1 (IRS-1) is a common cytosolic adaptor molecule involved in signal transduction from insulin and insulin-like growth factor I (IGF-I) receptors. IRS-1 can also be found in the nucleus. We report here a new finding of unique IRS-1 nuclear structures, which we observed initially in glioblastoma biopsy specimens and glioblastoma xenografts. These nuclear structures can be reproduced in vitro by the ectopic expression of IRS-1 cDNA cloned in frame with the nuclear localization signal (NLS-IRS-1). In these structures, IRS-1 localizes at the periphery, while the center harbors a key autophagy protein, LC3. These new nuclear structures are highly dynamic, rapidly exchange IRS-1 molecules with the surrounding nucleoplasm, disassemble during mitosis, and require a growth stimulus for their reassembly and maintenance. In tumor cells engineered to express NLS-IRS-1, the IRS-1/LC3 nuclear structures repress autophagy induced by either amino acid starvation or rapamycin treatment. In this process, IRS-1 nuclear structures sequester LC3 inside the nucleus, possibly preventing its cytosolic translocation and the formation of new autophagosomes. This novel mechanism provides a quick and reversible way of inhibiting autophagy, which could counteract autophagy-induced cancer cell death under severe stress, including anticancer therapies. Copyright © 2018 American Society for Microbiology.

Cross-species extrapolation of an adverse outcome pathway for ecdysone receptor activation

EPA Science Inventory

Different invertebrate nuclear receptors serve as targets for a variety of environmental contaminants. One of these is the ecdysteroid receptor (EcR). Due to the important role of this nuclear receptor in regulating development and reproduction in invertebrates, particularly duri...

Cross-species extrapolation of an adverse outcome pathway for ecdysteroid receptor activation

EPA Science Inventory

Different invertebrate nuclear receptors serve as targets for a variety of environmental contaminants. One of these is the ecdysteroid receptor (EcR). Due to the important role of this nuclear receptor in regulating development and reproduction in invertebrates, particularly duri...

Food Components Modulate Obesity and Energy Metabolism via the Transcriptional Regulation of Lipid-Sensing Nuclear Receptors.

PubMed

Goto, Tsuyoshi; Takahashi, Nobuyuki; Kawada, Teruo

2015-01-01

Obesity is a major risk factor for chronic diseases such as diabetes, cardiovascular diseases, and hypertension. Many modern people have a tendency to overeat owing to stress and loosening of self-control. Moreover, energy expenditure varies greatly among individuals. Scientific reduction of obesity is important under these circumstances. Furthermore, recent research on molecular levels has clarified the differentiation of adipocytes, the level of subsequent fat accumulation, and the secretion of the biologically active adipokines by adipocytes. Adipose tissues and obesity have become the most important target for the prevention and treatment of many chronic diseases. We have identified various food-derived compounds modulating nuclear receptors, especially peroxisome proliferators-activated receptor(PPAR), in the regulation of energy metabolism and obesity. In this review, we discuss the PPARs that are most important in obesity and energy metabolism.

Role of YAP activation in nuclear receptor CAR-mediated proliferation of mouse hepatocytes.

PubMed

Abe, Taiki; Amaike, Yuto; Shizu, Ryota; Takahashi, Miki; Kano, Makoto; Hosaka, Takuomi; Sasaki, Takamitsu; Kodama, Susumu; Matsuzawa, Atsushi; Yoshinari, Kouichi

2018-06-08

Constitutive androstane receptor (CAR) is a xenobiotic-responsive nuclear receptor that is highly expressed in the liver. CAR activation induces hepatocyte proliferation and hepatocarcinogenesis in rodents, but the mechanisms remain unclear. In this study, we investigated the association of CAR-dependent cell proliferation with Yes-associated protein (YAP), which is a transcriptional cofactor controlling organ size and cell growth through the interaction with various transcriptional factors including TEAD. In mouse livers, TCPOBOP (a mouse CAR activator) treatment increased the nuclear YAP accumulation and mRNA levels of YAP target genes as well as cell-cycle related genes along with liver hypertrophy and verteporfin (an inhibitor of YAP/TEAD interaction) cotreatment tended to attenuate them. Furthermore, in cell-based reporter gene assays, CAR activation enhanced the YAP/TEAD-dependent transcription. To investigate the role of YAP/TEAD activation in the CAR-dependent hepatocyte proliferation, we sought to establish an in vitro system completely reproducing CAR-dependent cell proliferation. Since CAR was only slightly expressed in cultured mouse primary hepatocytes compared to mouse livers and no proliferation was observed after treatment with TCPOBOP, we overexpressed CAR using mouse CAR expressing adenovirus (Ad-mCAR-V5) in mouse primary hepatocytes. Ad-mCAR-V5 infection and TCPOBOP treatment induced hepatocyte proliferation. Similar results were obtained with immortalized normal mouse hepatocytes as well. In the established in vitro system, CAR-dependent proliferation was strongly inhibited by Yap knockdown and completely abolished by verteporfin treatment. Our present results obtained in in vivo and in vitro experiments suggest that YAP/TEAD activation plays key roles in CAR-dependent proliferation of murine hepatocytes.

Function of multiple Lis-Homology domain/WD-40 repeat-containing proteins in feed-forward transcriptional repression by silencing mediator for retinoic and thyroid receptor/nuclear receptor corepressor complexes.

PubMed

Choi, Hyo-Kyoung; Choi, Kyung-Chul; Kang, Hee-Bum; Kim, Han-Cheon; Lee, Yoo-Hyun; Haam, Seungjoo; Park, Hyoung-Gi; Yoon, Ho-Geun

2008-05-01

Lis-homology (LisH) motifs are involved in protein dimerization, and the discovery of the conserved N-terminal LisH domain in transducin beta-like protein 1 and its receptor (TBL1 and TBLR1) led us to examine the role of this domain in transcriptional repression. Here we show that multiple beta-transducin (WD-40) repeat-containing proteins interact to form oligomers in solution and that oligomerization depends on the presence of the LisH domain in each protein. Repression of transcription, as assayed using Gal4 fusion proteins, also depended on the presence of the LisH domain, suggesting that oligomerization is a prerequisite for efficient transcriptional repression. Furthermore, we show that the LisH domain is responsible for the binding to the hypoacetylated histone H4 tail and for stable chromatin targeting by the nuclear receptor corepressor complex. Mutations in conserved residues in the LisH motif of TBL1 and TBLR1 block histone binding, oligomerization, and transcriptional repression, supporting the functional importance of the LisH motif in transcriptional repression. Our results indicate that another WD-40 protein, TBL3, also preferentially binds to the N-terminal domain of TBL1 and TBLR1, and forms oligomers with other WD-40 proteins. Finally, we observed that the WD-40 proteins RbAp46 and RbAp48 of the sin3A corepressor complex failed to dimerize. We also found the specific interaction UbcH/E2 with TBL1, but not RbAp46/48. Altogether, our results thus indicate that the presence of multiple LisH/WD-40 repeat containing proteins is exclusive to nuclear receptor corepressor/ silencing mediator for retinoic and thyroid receptor complexes compared with other class 1 histone deacetylase-containing corepessor complexes.

Androgen Receptor Content of the Normal and Hyperplastic Canine Prostate

PubMed Central

Shain, Sydney A.; Boesel, Robert W.

1978-01-01

A procedure was developed for measurement of androgen receptors in cytoplasmic extracts of prostates from intact dogs. The protocol utilized exchange saturation analysis at 15°C employing the synthetic androgen R1881 (17β-hydroxy-17α-methylestra-4,9,11-trien-3-one) as the ligand probe and quantitatively detected total cytoplasmic androgen receptor (Rc, androgen-free receptor, and RcA, androgen-occupied receptor) present at the initiation of the assay. This protocol was employed in conjunction with a tissue mince saturation analysis procedure (for quantitation of nuclear androgen receptor) to quantitate total androgen receptor content of normal and hyperplastic prostates obtained from young (2.5- or 4.6-yr old) and aged (12.5-yr old) purebred dogs of known birth date. The total cytoplasmic androgen receptor content (picomoles per prostate) of hyperplastic prostates was 4.6-fold greater than that of normal prostates. The total nuclear androgen receptor content of hyperplastic prostates (picomoles per prostate measured in crude nuclear preparations) was either 5.0- (4.6-yr-old dogs) or 7.8-fold (2.5-yr-old dogs) greater than that of normal prostates. However, androgen receptor content per cell was identical for hyperplastic and normal canine prostates, with the exception that nuclear androgen receptor was diminished in prostates from 2.5-yr-old dogs. The cell content per gram dry weight was identical for hyperplastic and normal canine prostates. We conclude that canine prostate hyperplasia is characterized by coordinate proliferation of androgen receptor-positive and androgen receptor-negative cells and is not a consequence of increased accumulation of 5α-dihydrotestosterone due to proliferation of androgen receptors per prostate cell. PMID:76635

Nuclear hormone retinoid X receptor (RXR) negatively regulates the glucose-stimulated insulin secretion of pancreatic ß-cells.

PubMed

Miyazaki, Satsuki; Taniguchi, Hidenori; Moritoh, Yusuke; Tashiro, Fumi; Yamamoto, Tsunehiko; Yamato, Eiji; Ikegami, Hiroshi; Ozato, Keiko; Miyazaki, Jun-ichi

2010-11-01

Retinoid X receptors (RXRs) are members of the nuclear hormone receptor superfamily and are thought to be key regulators in differentiation, cellular growth, and gene expression. Although several experiments using pancreatic β-cell lines have shown that the ligands of nuclear hormone receptors modulate insulin secretion, it is not clear whether RXRs have any role in insulin secretion. To elucidate the function of RXRs in pancreatic β-cells, we generated a double-transgenic mouse in which a dominant-negative form of RXRβ was inducibly expressed in pancreatic β-cells using the Tet-On system. We also established a pancreatic β-cell line from an insulinoma caused by the β-cell-specific expression of simian virus 40 T antigen in the above transgenic mouse. In the transgenic mouse, expression of the dominant-negative RXR enhanced the insulin secretion with high glucose stimulation. In the pancreatic β-cell line, the suppression of RXRs also enhanced glucose-stimulated insulin secretion at a high glucose concentration, while 9-cis-retinoic acid, an RXR agonist, repressed it. High-density oligonucleotide microarray analysis showed that expression of the dominant-negative RXR affected the expression levels of a number of genes, some of which have been implicated in the function and/or differentiation of β-cells. These results suggest that endogenous RXR negatively regulates the glucose-stimulated insulin secretion. Given these findings, we propose that the modulation of endogenous RXR in β-cells may be a new therapeutic approach for improving impaired insulin secretion in type 2 diabetes.

Transcriptional activation of NAD+-dependent protein deacetylase SIRT1 by nuclear receptor TLX.

PubMed

Iwahara, Naotoshi; Hisahara, Shin; Hayashi, Takashi; Horio, Yoshiyuki

2009-09-04

An orphan nuclear receptor TLX is a transcriptional repressor that promotes the proliferation and self-renewal of neural precursor cells (NPCs). SIRT1, an NAD(+)-dependent protein deacetylase, is highly expressed in the NPCs and participates in neurogenesis. Here, we found that TLX colocalized with SIRT1 and knockdown of TLX by small interfering RNAs decreased SIRT1 levels in NPCs. TLX increased the SIRT1 expression by binding to the newly identified TLX-activating element in the SIRT1 gene promoter in HEK293 cells. Thus, TLX is an inducer of SIRT1 and may contribute to neurogenesis both as a transactivator and as a repressor.

Expression of nuclear receptor interacting proteins TIF-1, SUG-1, receptor interacting protein 140, and corepressor SMRT in tamoxifen-resistant breast cancer.

PubMed

Chan, C M; Lykkesfeldt, A E; Parker, M G; Dowsett, M

1999-11-01

Regulation of gene transcription as a consequence of steroid receptor-DNA interaction is mediated via nuclear receptor interacting proteins (RIPs), including coactivator or corepressor proteins, which interact with both the receptor and components of the basic transcriptional unit and vary between cell types. The aim of this study was to test the hypothesis that resistance of some breast carcinomas to tamoxifen was associated with inappropriate expression of some of these RIPs. Using Northern analysis, we observed no significant difference between the amount of either TIF-1 or SUG-1 mRNA expressed in parental MCF-7 and MCF-7 tamoxifen-resistant cell lines. However, the expression of RIP140 mRNA was lower in the resistant cell line and in the presence of estradiol, the level of RIP140 mRNA was higher in the resistant cells but not in the parental cells. In a cohort of 19 tamoxifen-resistant breast tumor samples, there was no significant difference in the level of the RIP140 and TIF-1 and corepressor SMRT mRNA compared with tamoxifen-treated tumors (n = 6) or untreated tumors (n = 21). However, SUG-1 mRNA was lower in resistant breast tumors. These data provide no support for increased expression of these RIPs or decreased expression of corepressor SMRT for being a mechanism for resistance of breast tumors to tamoxifen.

SUMO proteases as potential targets for cancer therapy.

PubMed

Bialik, Piotr; Woźniak, Katarzyna

2017-12-08

Sumoylation is one of the post-translational modifications of proteins, responsible for the regulation of many cellular processes, such as DNA replication and repair, transcription, signal transduction and nuclear transport. During sumoylation, SUMO proteins are covalently attached to the ε-amino group of lysine in target proteins via an enzymatic cascade that requires the sequential action of E1, E2 and E3 enzymes. An important aspect of sumoylation is its reversibility, which involves SUMO-specific proteases called SENPs. SENPs (sentrin/SUMO-specific proteases) catalyze the deconjugation of SUMO proteins using their isopeptidase activity. These enzymes participate through hydrolase activity in the reaction of SUMO protein maturation, which involves the removal of a short fragment on the C-terminus of SUMO inactive form and exposure two glycine residues. SENPs are important for maintaining the balance between sumoylated and desumoylated proteins required for normal cellular physiology. Six SENP isoforms (SENP1, SENP2, SENP3, SENP5, SENP6 and SENP7) have been identified in mammals. These SENPs can be divided into three subfamilies based on their sequence homology, substrate specificity and subcellular localization. Results of studies indicate the role of SUMO proteases in the development of human diseases including cancer, suggesting that these proteins may be attractive targets for new drugs.

Increasing human Th17 differentiation through activation of orphan nuclear receptor retinoid acid-related orphan receptor γ (RORγ) by a class of aryl amide compounds.

PubMed

Zhang, Wei; Zhang, Jing; Fang, Leiping; Zhou, Ling; Wang, Shuai; Xiang, Zhijun; Li, Yuan; Wisely, Bruce; Zhang, Guifeng; An, Gang; Wang, Yonghui; Leung, Stewart; Zhong, Zhong

2012-10-01

In a screen for small-molecule inhibitors of retinoid acid-related orphan receptor γ (RORγ), we fortuitously discovered that a class of aryl amide compounds behaved as functional activators of the interleukin 17 (IL-17) reporter in Jurkat cells. Three of these compounds were selected for further analysis and found to activate the IL-17 reporter with potencies of ∼0.1 μM measured by EC₅₀. These compounds were shown to directly bind to RORγ by circular dichroism-based thermal stability experiments. Furthermore, they can enhance an in vitro Th17 differentiation process in human primary T cells. As RORγ remains an orphan nuclear receptor, discovery of these aryl amide compounds as functional agonists will now provide pharmacological tools for us to dissect functions of RORγ and facilitate drug discovery efforts for immune-modulating therapies.

Drosophila motor neuron retraction during metamorphosis is mediated by inputs from TGF-β/BMP signaling and orphan nuclear receptors.

PubMed

Boulanger, Ana; Farge, Morgane; Ramanoudjame, Christophe; Wharton, Kristi; Dura, Jean-Maurice

2012-01-01

Larval motor neurons remodel during Drosophila neuro-muscular junction dismantling at metamorphosis. In this study, we describe the motor neuron retraction as opposed to degeneration based on the early disappearance of β-Spectrin and the continuing presence of Tubulin. By blocking cell dynamics with a dominant-negative form of Dynamin, we show that phagocytes have a key role in this process. Importantly, we show the presence of peripheral glial cells close to the neuro-muscular junction that retracts before the motor neuron. We show also that in muscle, expression of EcR-B1 encoding the steroid hormone receptor required for postsynaptic dismantling, is under the control of the ftz-f1/Hr39 orphan nuclear receptor pathway but not the TGF-β signaling pathway. In the motor neuron, activation of EcR-B1 expression by the two parallel pathways (TGF-β signaling and nuclear receptor) triggers axon retraction. We propose that a signal from a TGF-β family ligand is produced by the dismantling muscle (postsynapse compartment) and received by the motor neuron (presynaptic compartment) resulting in motor neuron retraction. The requirement of the two pathways in the motor neuron provides a molecular explanation for the instructive role of the postsynapse degradation on motor neuron retraction. This mechanism insures the temporality of the two processes and prevents motor neuron pruning before postsynaptic degradation.

Nuclear and Membrane Actions of Estrogen Receptor Alpha: Contribution to the Regulation of Energy and Glucose Homeostasis.

PubMed

Guillaume, Maeva; Montagner, Alexandra; Fontaine, Coralie; Lenfant, Françoise; Arnal, Jean-François; Gourdy, Pierre

2017-01-01

Estrogen receptor alpha (ERα) has been demonstrated to play a key role in reproduction but also to exert numerous functions in nonreproductive tissues. Accordingly, ERα is now recognized as a key regulator of energy homeostasis and glucose metabolism and mediates the protective effects of estrogens against obesity and type 2 diabetes. This chapter attempts to summarize our current understanding of the mechanisms of ERα activation and their involvement in the modulation of energy balance and glucose metabolism. We first focus on the experimental studies that constitute the basis of the understanding of ERα as a nuclear receptor and more specifically on the key roles played by its two activation functions (AFs). We depict the consequences of the selective inactivation of these AFs in mouse models, which further underline the prominent role of nuclear ERα in the prevention of obesity and diabetes, as on the reproductive tract and the vascular system. Besides these nuclear actions, a fraction of ERα is associated with the plasma membrane and activates nonnuclear signaling from this site. Such rapid effects, called membrane-initiated steroid signals (MISS), have been characterized in a variety of cell lines and in particular in endothelial cells. The development of selective pharmacological tools that specifically activate MISS as well as the generation of mice expressing an ERα protein impeded for membrane localization has just begun to unravel the physiological role of MISS in vivo and their contribution to ERα-mediated metabolic protection. Finally, we discuss novel perspectives for the design of tissue-selective ER modulators.

Isolation, characterization, and expression analyses of ecdysone receptor 1, ecdysone receptor 2 and ultraspiracle genes in varroa destructor mite

USDA-ARS?s Scientific Manuscript database

The varroa mite, Varroa destructor, is a honeybee ectoparasite considered the most important pest in apiaries throughout the US. Ecdysone receptor is a hormone secreted by the prothoracic gland of insects that controls ecdysis and stimulates metamorphosis. The ecdysone receptor is a nuclear receptor...

The Drosophila FTZ-F1 Nuclear Receptor Mediates Juvenile Hormone Activation of E75A Gene Expression through an Intracellular Pathway*

PubMed Central

Dubrovsky, Edward B.; Dubrovskaya, Veronica A.; Bernardo, Travis; Otte, Valerie; DiFilippo, Robert; Bryan, Heather

2011-01-01

Juvenile hormone (JH) regulates a wide variety of biological activities in holometabolous insects, ranging from vitellogenesis and caste determination in adults to the timing of metamorphosis in larvae. The mechanism of JH signaling in such a diverse array of processes remains either unknown or contentious. We previously found that the nuclear receptor gene E75A is activated in S2 cells as a primary response to JH. Here, by expressing an intracellular form of JH esterase, we demonstrate that JH must enter the cell in order to activate E75A. To find intracellular receptors involved in the JH response, we performed an RNAi screen against nuclear receptor genes expressed in this cell line and identified the orphan receptor FTZ-F1. Removal of FTZ-F1 prevents JH activation of E75A, whereas overexpression enhances activation, implicating FTZ-F1 as a critical component of the JH response. FTZ-F1 is bound in vivo to multiple enhancers upstream of E75A, suggesting that it participates in direct JH-mediated gene activation. To better define the role of FTZ-F1 in JH signaling, we investigated interactions with candidate JH receptors and found that the bHLH-PAS proteins MET and GCE both interact with FTZ-F1 and can activate transcription through the FTZ-F1 response element. Removal of endogenous GCE, but not MET, prevents JH activation of E75A. We propose that FTZ-F1 functions as a competence factor by loading JH signaling components to the promoter, thus facilitating the direct regulation of E75A gene expression by JH. PMID:21832074

Spatial distribution of the messenger ribonucleic acid and protein of the nuclear receptor coactivator, amplified in breast cancer-3, in mice.

PubMed

Zhang, Hao; Liao, Lan; Kuang, Shao-Qing; Xu, Jianming

2003-04-01

Transcriptional activities of nuclear receptors are modulated by coactivators and corepressors. The amplified in breast cancer-3 protein (AIB3, also known as ASC-2, RAP250, PRIP, TRBP, and NCR) is a newly identified nuclear receptor coactivator that is amplified and overexpressed in breast cancers. This study aims to investigate the spatial expression of AIB3 mRNA and protein in various murine tissues. Quantitative measurements revealed that the concentrations of AIB3 mRNA differ substantially in different tissues in a descending order from the following: testis, brain, thymus, white fat, pituitary, ovary, adrenal gland, lung, uterus, kidney, heart, skeletal muscle, liver, and virgin mammary gland. The AIB3 mRNA level in the testis is 165-fold higher than that in the virgin mammary gland. Specific antiserum was generated and used to map the distribution of AIB3 protein by immunohistochemistry. Although AIB3 protein was detected in many tissues, the AIB3 immunoreactivities varied significantly from cell type to cell type. High levels of AIB3 immunoreactivity were observed in hormone target cells including the testicular Sertoli cells, follicular granulosa cells, and epithelial cells of the prostate, uterus, mammary gland, and kidney tubules. Medium and low levels of AIB3 immunoreactivities were also detected in a variety of other cell types. These results demonstrate that AIB3 mRNA and protein are preferentially expressed in specific cell types, suggesting that AIB3 may support the function of nuclear receptors in a cell type-specific manner.

Functional Analysis of Nuclear Estrogen Receptors in Zebrafish Reproduction by Genome Editing Approach.

PubMed

Lu, Huijie; Cui, Yong; Jiang, Liwen; Ge, Wei

2017-07-01

Estrogens signal through both nuclear and membrane receptors with most reported effects being mediated via the nuclear estrogen receptors (nERs). Although much work has been reported on nERs in the zebrafish, there is a lack of direct genetic evidence for their functional roles and importance in reproduction. To address this issue, we undertook this study to disrupt all three nERs in the zebrafish, namely esr1 (ERα), esr2a (ERβII), and esr2b (ERβI), by the genome-editing technology clustered regularly interspaced short palindromic repeats and its associated nuclease (CRISPR/Cas9). Using this loss-of-function genetic approach, we successfully created three mutant zebrafish lines with each nER knocked out. In addition, we also generated all possible double and triple knockouts of the three nERs. The phenotypes of these mutants in reproduction were analyzed in all single, double, and triple nER knockouts in both females and males. Surprisingly, all three single nER mutant fish lines display normal reproductive development and function in both females and males, suggesting functional redundancy among these nERs. Further analysis of double and triple knockouts showed that nERs, especially Esr2a and Esr2b, were essential for female reproduction, and loss of these two nERs led to an arrest of folliculogenesis at previtellogenic stage II followed by sex reversal from female to male. In addition, the current study also revealed a unique role for Esr2a in follicle cell proliferation and transdifferentiation, follicle growth, and chorion formation. Taken together, this study provides the most comprehensive genetic analysis for differential functions of esr1, esr2a, and esr2b in fish reproduction. Copyright © 2017 Endocrine Society.

Direct modification and regulation of a nuclear receptor-PIP2 complex by the nuclear inositol-lipid kinase IPMK

PubMed Central

Blind, Raymond D.; Suzawa, Miyuki; Ingraham, Holly A.

2012-01-01

Phosphatidylinositol (4,5)-bisphosphate (PIP2) is best known as a plasma membrane-bound regulatory lipid. While PIP2 and phosphoinositide-modifying enzymes coexist in the nucleus, their roles in the nucleus remain unclear. Here we show that the nuclear inositol polyphosphate multikinase (IPMK), which functions both as an inositol- and a PI3-kinase, interacts with the nuclear receptor SF-1 (NR5A1) and phosphorylates its bound ligand, PIP2. IPMK failed to recognize SF-1/PIP2 after blocking or displacing PIP2 from SF-1’s large hydrophobic pocket. In contrast to IPMK, p110 catalytic subunits of type 1 PI3-kinases were inactive on SF-1/PIP2. These and other in vitro analyses demonstrated specificity of IPMK for the SF-1/PIP2 protein/lipid complex. Once generated, SF-1/PIP3 is readily dephosphorylated by the lipid phosphatase PTEN. Importantly, decreasing IPMK or increasing PTEN expression greatly reduced SF-1 transcriptional activity. This ability of lipid kinases and phosphatases to alter the activity and directly remodel a non-membrane protein/lipid complex such SF-1/PIP2, establishes a new pathway for promoting lipid-mediated signaling in the nucleus. PMID:22715467

Cow's milk increases the activities of human nuclear receptors peroxisome proliferator-activated receptors alpha and delta and retinoid X receptor alpha involved in the regulation of energy homeostasis, obesity, and inflammation.

PubMed

Suhara, W; Koide, H; Okuzawa, T; Hayashi, D; Hashimoto, T; Kojo, H

2009-09-01

The nuclear peroxisome proliferator-activated receptors (PPAR) have been shown to play crucial roles in regulating energy homeostasis including lipid and carbohydrate metabolism, inflammatory responses, and cell proliferation, differentiation, and survival. Because PPAR agonists have the potential to prevent or ameliorate diseases such as hyperlipidemia, diabetes, atherosclerosis, and obesity, we have explored new natural agonists for PPAR. For this purpose, cow's milk was tested for agonistic activity toward human PPAR subtypes using a reporter gene assay. Milk increased human PPARalpha activity in a dose-dependent manner with a 3.2-fold increase at 0.5% (vol/vol). It also enhanced human PPARdelta activity in a dose-dependent manner with an 11.5-fold increase at 0.5%. However, it only slightly affected human PPARgamma activity. Ice cream, butter, and yogurt also increased the activities of PPARalpha and PPARdelta, whereas vegetable cream affected activity of PPARdelta but not PPARalpha. Skim milk enhanced the activity of PPAR to a lesser degree than regular milk. Milk and fresh cream increased the activity of human retinoid X receptor (RXR)alpha as well as PPARalpha and PPARdelta, whereas neither affected vitamin D3 receptor, estrogen receptors alpha and beta, or thyroid receptors alpha and beta. Both milk and fresh cream were shown by quantitative real-time PCR to increase the quantity of mRNA for uncoupling protein 2 (UCP2), an energy expenditure gene, in a dose-dependent manner. The increase in UCP2 mRNA was found to be reduced by treatment with PPARdelta-short interfering (si)RNA. This study unambiguously clarified at the cellular level that cow's milk increased the activities of human PPARalpha, PPARdelta, and RXRalpha. The possible role in enhancing the activities of PPARalpha, PPARdelta, and RXRalpha, and the health benefits of cow's milk were discussed.

A physical model describing the interaction of nuclear transport receptors with FG nucleoporin domain assemblies.

PubMed

Zahn, Raphael; Osmanović, Dino; Ehret, Severin; Araya Callis, Carolina; Frey, Steffen; Stewart, Murray; You, Changjiang; Görlich, Dirk; Hoogenboom, Bart W; Richter, Ralf P

2016-04-08

The permeability barrier of nuclear pore complexes (NPCs) controls bulk nucleocytoplasmic exchange. It consists of nucleoporin domains rich in phenylalanine-glycine motifs (FG domains). As a bottom-up nanoscale model for the permeability barrier, we have used planar films produced with three different end-grafted FG domains, and quantitatively analyzed the binding of two different nuclear transport receptors (NTRs), NTF2 and Importin β, together with the concomitant film thickness changes. NTR binding caused only moderate changes in film thickness; the binding isotherms showed negative cooperativity and could all be mapped onto a single master curve. This universal NTR binding behavior - a key element for the transport selectivity of the NPC - was quantitatively reproduced by a physical model that treats FG domains as regular, flexible polymers, and NTRs as spherical colloids with a homogeneous surface, ignoring the detailed arrangement of interaction sites along FG domains and on the NTR surface.

The Hsp90 Inhibitor, 17-AAG, Prevents the Ligand-Independent Nuclear Localization of Androgen Receptor in Refractory Prostate Cancer Cells

PubMed Central

Saporita, Anthony J.; Ai, Junkui; Wang, Zhou

2010-01-01

BACKGROUND Androgen receptor (AR) is the key molecule in androgen-refractory prostate cancer. Despite androgen ablative conditions, AR remains active and is necessary for the growth of androgen-refractory prostate cancer cells. Nuclear localization of AR is a prerequisite for its transcriptional activation. We examined AR localization in androgen-dependent and androgen-refractory prostate cancer cells. METHODS AND RESULTS We demonstrate increased nuclear localization of a GFP-tagged AR in the absence of hormone in androgen-refractory C4-2 cells compared to parental androgen-sensitive human prostate cancer LNCaP cells. Analysis of AR mutants impaired in ligand-binding indicates that the nuclear localization of AR in C4-2 cells is truly androgen-independent. The hsp90 inhibitor, 17-allylamino-17-demethoxygeldanamycin (17-AAG), inhibits basal PSA expression and disrupts the ligand-independent nuclear localization of AR at doses much lower than required to inhibit androgen-induced nuclear import. CONCLUSIONS Hsp90 is a key regulator of ligand-independent nuclear localization and activation of AR in androgen-refractory prostate cancer cells. PMID:17221841

Rational Quantitative Structure-Activity Relationship (RQSAR) Screen for PXR and CAR Isoform-Specific Nuclear Receptor Ligands

PubMed Central

Dring, Ann M.; Anderson, Linnea E.; Qamar, Saima; Stoner, Matthew A.

2010-01-01

Constitutive androstane receptor (CAR) and pregnane X receptor (PXR) are closely related orphan nuclear receptor proteins that share several ligands and target overlapping sets of genes involved in homeostasis and all phases of drug metabolism. CAR and PXR are involved in the development of certain diseases, including diabetes, metabolic syndrome and obesity. Ligand screens for these receptors so far have typically focused on steroid hormone analogs with pharmacophore-based approaches, only to find relatively few new hits. Multiple CAR isoforms have been detected in human liver, with the most abundant being the constitutively active reference, CAR1, and the ligand-dependent isoform CAR3. It has been assumed that any compound that binds CAR1 should also activate CAR3, and so CAR3 can be used as a ligand-activated surrogate for CAR1 studies. The possibility of CAR3-specific ligands has not, so far, been addressed. To investigate the differences between CAR1, CAR3 and PXR, and to look for more CAR ligands that may be of use in quantitative structure-activity relationship (QSAR) studies, we performed a luciferase transactivation assay screen of 60 mostly non-steroid compounds. Known active compounds with different core chemistries were chosen as starting points and structural variants were rationally selected for screening. Distinct differences in agonist versus inverse agonist/antagonist effects were seen in 49 compounds that had some ligand effect on at least one receptor and 18 that had effects on all three receptors; eight were CAR1 ligands only, three were CAR3 only ligands and four affected PXR only. This work provides evidence for new CAR ligands, some of which have CAR3-specific effects, and provides observational data on CAR and PXR ligands with which to inform in silico strategies. Compounds that demonstrated unique activity on any one receptor are potentially valuable diagnostic tools for the investigation of in vivo molecular targets. PMID:20869355

The dynamics of nuclear receptors and nuclear receptor coregulators in the pathogenesis of endometriosis

PubMed Central

Han, Sang Jun; O'Malley, Bert W.

2014-01-01

BACKGROUND Endometriosis is defined as the colonization and growth of endometrial tissue at anatomic sites outside the uterine cavity. Up to 15% of reproductive-aged women in the USA suffer from painful symptoms of endometriosis, such as infertility, pelvic pain, menstrual cycle abnormalities and increased risk of certain cancers. However, many of the current clinical treatments for endometriosis are not sufficiently effective and yield unacceptable side effects. There is clearly an urgent need to identify new molecular mechanisms that critically underpin the initiation and progression of endometriosis in order to develop more specific and effective therapeutics which lack the side effects of current therapies. The aim of this review is to discuss how nuclear receptors (NRs) and their coregulators promote the progression of endometriosis. Understanding the pathogenic molecular mechanisms for the genesis and maintenance of endometriosis as modulated by NRs and coregulators can reveal new therapeutic targets for alternative endometriosis treatments. METHODS This review was prepared using published gene expression microarray data sets obtained from patients with endometriosis and published literature on NRs and their coregulators that deal with endometriosis progression. Using the above observations, our current understanding of how NRs and NR coregulators are involved in the progression of endometriosis is summarized. RESULTS Aberrant levels of NRs and NR coregulators in ectopic endometriosis lesions are associated with the progression of endometriosis. As an example, endometriotic cell-specific alterations in gene expression are correlated with a differential methylation status of the genome compared with the normal endometrium. These differential epigenetic regulations can generate favorable cell-specific NR and coregulator milieus for endometriosis progression. Genetic alterations, such as single nucleotide polymorphisms and insertion/deletion polymorphisms of NR

Endogenous fatty acid ethanolamides suppress nicotine-induced activation of mesolimbic dopamine neurons through nuclear receptors.

PubMed

Melis, Miriam; Pillolla, Giuliano; Luchicchi, Antonio; Muntoni, Anna Lisa; Yasar, Sevil; Goldberg, Steven R; Pistis, Marco

2008-12-17

Nicotine stimulates the activity of mesolimbic dopamine neurons, which is believed to mediate the rewarding and addictive properties of tobacco use. Accumulating evidence suggests that the endocannabinoid system might play a major role in neuronal mechanisms underlying the rewarding properties of drugs of abuse, including nicotine. Here, we investigated the modulation of nicotine effects by the endocannabinoid system on dopamine neurons in the ventral tegmental area with electrophysiological techniques in vivo and in vitro. We discovered that pharmacological inhibition of fatty acid amide hydrolase (FAAH), the enzyme that catabolizes fatty acid ethanolamides, among which the endocannabinoid anandamide (AEA) is the best known, suppressed nicotine-induced excitation of dopamine cells. Importantly, this effect was mimicked by the administration of the FAAH substrates oleoylethanolamide (OEA) and palmitoylethanolamide (PEA), but not methanandamide, the hydrolysis resistant analog of AEA. OEA and PEA are naturally occurring lipid signaling molecules structurally related to AEA, but devoid of affinity for cannabinoid receptors. They blocked the effects of nicotine by activation of the peroxisome proliferator-activated receptor-alpha (PPAR-alpha), a nuclear receptor transcription factor involved in several aspects of lipid metabolism and energy balance. Activation of PPAR-alpha triggered a nongenomic stimulation of tyrosine kinases, which might lead to phosphorylation and negative regulation of neuronal nicotinic acetylcholine receptors. These data indicate for the first time that the anorexic lipids OEA and PEA possess neuromodulatory properties as endogenous ligands of PPAR-alpha in the brain and provide a potential new target for the treatment of nicotine addiction.

Uterine Development and Fertility Are Dependent on Gene Dosage of the Nuclear Receptor Coregulator REA

PubMed Central

Park, Sunghee; Yoon, Sangyeon; Zhao, Yuechao; Park, Seong-Eun; Liao, Lan; Xu, Jianming; Lydon, John P.; DeMayo, Francesco J.; O'Malley, Bert W.; Bagchi, Milan K.

2012-01-01

Although the effectiveness of nuclear hormone-receptor complexes is known to depend on coregulator partner proteins, relatively little is known about the roles of coregulators in uterine development and early stages of pregnancy and implantation. Because conventional genetic deletion of the coregulator, repressor of estrogen receptor activity (REA), was embryonic lethal, we here study REA conditional knockout mice generated by cre-loxP recombination, in which REA function was abrogated only in progesterone receptor-expressing tissues, to define the roles of REA in postembryonic stages and in a tissue-specific manner. We find that REA has gene dose-dependent activity impacting uterine development and fertility. Conditional homozygous mutant (REAd/d) mice developed to adulthood and showed normal ovarian function, but females were infertile with severely compromised uterine development and function characterized by cell cycle arrest, apoptosis, and altered adenogenesis (endometrial gland morphogenesis), resulting in failure of implantation and decidualization. By contrast, mice heterozygous for REA (REAf/d) had a very different phenotype, with estradiol treatment resulting in hyperstimulated, large uteri showing increased proliferation of luminal epithelial cells, and enhanced fluid imbibition associated with altered regulation of aquaporins. These REAf/d female mice showed a subfertility phenotype with reduced numbers and sizes of litters. These findings highlight that uterine development and regulation of estrogen receptor activities show a bimodal dependence on the gene dosage of REA. Optimal uterine development and functional activities require the normal gene dosage of REA, with partial or complete deletion resulting in hyperresponsiveness or underresponsiveness to hormone and subfertility or infertility, respectively. PMID:22585830

Profiling of 3696 Nuclear Receptor-Coregulator Interactions: A Resource for Biological and Clinical Discovery.

PubMed

Broekema, Marjoleine F; Hollman, Danielle A A; Koppen, Arjen; van den Ham, Henk-Jan; Melchers, Diana; Pijnenburg, Dirk; Ruijtenbeek, Rob; van Mil, Saskia W C; Houtman, René; Kalkhoven, Eric

2018-06-01

Nuclear receptors (NRs) are ligand-inducible transcription factors that play critical roles in metazoan development, reproduction, and physiology and therefore are implicated in a broad range of pathologies. The transcriptional activity of NRs critically depends on their interaction(s) with transcriptional coregulator proteins, including coactivators and corepressors. Short leucine-rich peptide motifs in these proteins (LxxLL in coactivators and LxxxIxxxL in corepressors) are essential and sufficient for NR binding. With 350 different coregulator proteins identified to date and with many coregulators containing multiple interaction motifs, an enormous combinatorial potential is present for selective NR-mediated gene regulation. However, NR-coregulator interactions have often been determined experimentally on a one-to-one basis across diverse experimental conditions. In addition, NR-coregulator interactions are difficult to predict because the molecular determinants that govern specificity are not well established. Therefore, many biologically and clinically relevant NR-coregulator interactions may remain to be discovered. Here, we present a comprehensive overview of 3696 NR-coregulator interactions by systematically characterizing the binding of 24 nuclear receptors with 154 coregulator peptides. We identified unique ligand-dependent NR-coregulator interaction profiles for each NR, confirming many well-established NR-coregulator interactions. Hierarchical clustering based on the NR-coregulator interaction profiles largely recapitulates the classification of NR subfamilies based on the primary amino acid sequences of the ligand-binding domains, indicating that amino acid sequence is an important, although not the only, molecular determinant in directing and fine-tuning NR-coregulator interactions. This NR-coregulator peptide interactome provides an open data resource for future biological and clinical discovery as well as NR-based drug design.

Importance of the pharmacological profile of the bound ligand in enrichment on nuclear receptors: toward the use of experimentally validated decoy ligands.

PubMed

Lagarde, Nathalie; Zagury, Jean-François; Montes, Matthieu

2014-10-27

The evaluation of virtual ligand screening methods is of major importance to ensure their reliability. Taking into account the agonist/antagonist pharmacological profile should improve the quality of the benchmarking data sets since ligand binding can induce conformational changes in the nuclear receptor structure and such changes may vary according to the agonist/antagonist ligand profile. We indeed found that splitting the agonist and antagonist ligands into two separate data sets for a given nuclear receptor target significantly enhances the quality of the evaluation. The pharmacological profile of the ligand bound in the binding site of the target structure was also found to be an additional critical parameter. We also illustrate that active compound data sets for a given pharmacological activity can be used as a set of experimentally validated decoy ligands for another pharmacological activity to ensure a reliable and challenging evaluation of virtual screening methods.