BCOR modulates transcriptional activity of a subset of glucocorticoid receptor target genes involved in cell growth and mobility

Abstract Several posttranslational modifications including phosphorylation have been detected on the glucocorticoid receptor (GR). However, the interdependence and combinatorial regulation of these modifications and their role in GR functions are poorly understood. We studied the effects of c-Jun N-terminal kinase (JNK)-dependent phosphorylation of GR on its sumoylation status and the impact that these modifications have on GR transcriptional activity. GR is targeted for phosphorylation at serine 246 (S246) by the JNK protein family in a rapid and transient manner. The levels of S246 phosphorylation of endogenous GR increased significantly in cells treated with UV radiation that activates JNK. S246 GR phosphorylation by JNK facilitated subsequent GR sumoylation at lysines 297 and 313. GR sumoylation increased with JNK activation and was inhibited in cells treated with JNK inhibitor. GR sumoylation in cells with activated JNK was mediated preferentially by small ubiquitin-like modifier (SUMO)2 rather than SUMO1. An increase in GR transcriptional activity was observed after inhibition of JNK or SUMO pathways and suppression of GR transcriptional activity after activation of both pathways in cells transfected with GR-responsive reporter genes. Endogenous GR transcriptional activity was inhibited on endogenous target genes IGF binding protein (IGFBP) and glucocorticoid-induced leucine zipper (GILZ) when JNK and SUMO pathways were induced individually or simultaneously. Activation of both of these signals inhibited GR-mediated regulation of human inhibitor of apoptosis gene (hIAP), whereas simultaneous activation had no effect. We conclude that phosphorylation aids GR sumoylation and that cross talk of JNK and SUMO pathways fine tune GR transcriptional activity in a target gene-specific manner, thereby modulating the hormonal response of cells exposed to stress.

Download Full-text

Lysine acetyltransferase Tip60 acetylates the APP adaptor Fe65 to increase its transcriptional activity

Biological Chemistry ◽

10.1515/hsz-2020-0279 ◽

2020 ◽

Vol 0 (0) ◽

Author(s):

Sabine Probst ◽

Florian Riese ◽

Larissa Kägi ◽

Maik Krüger ◽

Natalie Russi ◽

...

Keyword(s):

Transcriptional Activity ◽

Target Genes ◽

Adaptor Protein ◽

Proteolytic Processing ◽

Class Iii ◽

Lysine Deacetylase ◽

Lysine Residues ◽

Lysine Acetyltransferase ◽

Complex Forms ◽

Spot Formation

AbstractProteolytic processing of the amyloid precursor protein (APP) releases the APP intracellular domain (AICD) from the membrane. Bound to the APP adaptor protein Fe65 and the lysine acetyltransferase (KAT) Tip60, AICD translocates to the nucleus. Here, the complex forms spherical condensates at sites of endogenous target genes, termed AFT spots (AICD-Fe65-Tip60). We show that loss of Tip60 KAT activity prevents autoacetylation, reduces binding of Fe65 and abolishes Fe65-mediated stabilization of Tip60. Autoacetylation is a prerequisite for AFT spot formation, with KAT-deficient Tip60 retained together with Fe65 in speckles. We identify lysine residues 204 and 701 of Fe65 as acetylation targets of Tip60. We do not detect acetylation of AICD. Mutation of Fe65 K204 and K701 to glutamine, mimicking acetylation-induced charge neutralization, increases the transcriptional activity of Fe65 whereas Tip60 inhibition reduces it. The lysine deacetylase (KDAC) class III Sirt1 deacetylates Fe65 and pharmacological modulation of Sirt1 activity regulates Fe65 transcriptional activity. A second acetylation/deacetylation cycle, conducted by CBP and class I/II KDACs at different lysine residues, regulates stability of Fe65. This is the first report describing a role for acetylation in the regulation of Fe65 transcriptional activity, with Tip60 being the only KAT tested that supports AFT spot formation.

Download Full-text

Glucocorticoid Receptor Phosphorylation Differentially Affects Target Gene Expression

Molecular Endocrinology ◽

10.1210/me.2007-0219 ◽

2008 ◽

Vol 22 (8) ◽

pp. 1754-1766 ◽

Cited By ~ 157

Author(s):

Weiwei Chen ◽

Thoa Dang ◽

Raymond D. Blind ◽

Zhen Wang ◽

Claudio N. Cavasotto ◽

...

Keyword(s):

Glucocorticoid Receptor ◽

Transcriptional Activation ◽

Leucine Zipper ◽

Target Genes ◽

Divalent Cations ◽

Phosphorylation Site ◽

Transcriptional Response ◽

Receptor Occupancy ◽

Wild Type ◽

Interacting Protein

Abstract The glucocorticoid receptor (GR) is phosphorylated at multiple sites within its N terminus (S203, S211, S226), yet the role of phosphorylation in receptor function is not understood. Using a range of agonists and GR phosphorylation site-specific antibodies, we demonstrated that GR transcriptional activation is greatest when the relative phosphorylation of S211 exceeds that of S226. Consistent with this finding, a replacement of S226 with an alanine enhances GR transcriptional response. Using a battery of compounds that perturb different signaling pathways, we found that BAPTA-AM, a chelator of intracellular divalent cations, and curcumin, a natural product with antiinflammatory properties, reduced hormone-dependent phosphorylation at S211. This change in GR phosphorylation was associated with its decreased nuclear retention and transcriptional activation. Molecular modeling suggests that GR S211 phosphorylation promotes a conformational change, which exposes a novel surface potentially facilitating cofactor interaction. Indeed, S211 phosphorylation enhances GR interaction with MED14 (vitamin D receptor interacting protein 150). Interestingly, in U2OS cells expressing a nonphosphorylated GR mutant S211A, the expression of IGF-binding protein 1 and interferon regulatory factor 8, both MED14-dependent GR target genes, was reduced relative to cells expressing wild-type receptor across a broad range of hormone concentrations. In contrast, the induction of glucocorticoid-induced leucine zipper, a MED14-independent GR target, was similar in S211A- and wild-type GR-expressing cells at high hormone levels, but was reduced in S211A cells at low hormone concentrations, suggesting a link between GR phosphorylation, MED14 involvement, and receptor occupancy. Phosphorylation also affected the magnitude of repression by GR in a gene-selective manner. Thus, GR phosphorylation at S211 and S226 determines GR transcriptional response by modifying cofactor interaction. Furthermore, the effect of GR S211 phosphorylation is gene specific and, in some cases, dependent upon the amount of activated receptor.

Download Full-text

RSUME Enhances Glucocorticoid Receptor SUMOylation and Transcriptional Activity

Molecular and Cellular Biology ◽

10.1128/mcb.01470-12 ◽

2013 ◽

Vol 33 (11) ◽

pp. 2116-2127 ◽

Cited By ~ 30

Author(s):

J. Druker ◽

A. C. Liberman ◽

M. Antunica-Noguerol ◽

J. Gerez ◽

M. Paez-Pereda ◽

...

Keyword(s):

Glucocorticoid Receptor ◽

Transcriptional Activity

Download Full-text

Transcriptional transactivation functions localized to the glucocorticoid receptor N terminus are necessary for steroid induction of lymphocyte apoptosis

Molecular and Cellular Biology ◽

10.1128/mcb.12.2.589-597.1992 ◽

1992 ◽

Vol 12 (2) ◽

pp. 589-597

Author(s):

E S Dieken ◽

R L Miesfeld

Keyword(s):

Transcriptional Regulation ◽

Glucocorticoid Receptor ◽

Target Genes ◽

Dominant Negative ◽

Receptor Expression ◽

Transactivation Domain ◽

Lymphocyte Apoptosis ◽

Murine Cell Line ◽

N Terminus ◽

Simplex Virus

Genetic studies have suggested that transcriptional regulation of specific target genes (by either induction or repression) is the molecular basis of glucocorticoid-mediated lymphocyte apoptosis. To examine the role of transcriptional regulation more directly, we developed a complementation assay utilizing stable transfection of wild-type (wt) and mutant (nti) glucocorticoid receptor (GR) cDNA constructs into a GR-deficient S49 murine cell line (7r). Our data confirm that the level of functional GR is rate limiting for S49 apoptosis and moreover that the GR amino terminus (N terminus), which as been deleted from the nti GR, is absolutely required for complementation in this system. Surprisingly, we found that at physiological levels of receptor, expression of the nti GR in cells containing wt GR results in enhanced dexamethasone sensitivity rather than a dominant negative phenotype. One interpretation of these data is that DNA binding by wt-nti heterodimers may be functionally similar to that of wt-wt homodimers, indicating that GRE occupancy by at least one transactivation domain may be sufficient to induce the hormonal response. To determine whether acidic activating sequences such as those localized to the GR N terminus are important in the induction of lymphocyte apoptosis, we tested the activity of a chimeric receptor in which we replaced the entire GR N terminus with sequences from the herpes simplex virus VP16 protein. Our results demonstrate that 7r cells expressing VP-GR fusions are indeed steroid sensitive, strongly supporting the idea that S49 apoptosis is dependent on transcriptional regulation of specific genes which respond to acidic activating domains, implying that induction, rather than repression, may be the critical initiating event.

Download Full-text

Negative Correlation between the Diffusion Coefficient and Transcriptional Activity of the Glucocorticoid Receptor

International Journal of Molecular Sciences ◽

10.3390/ijms18091855 ◽

2017 ◽

Vol 18 (9) ◽

pp. 1855 ◽

Cited By ~ 3

Author(s):

◽

Keyword(s):

Diffusion Coefficient ◽

Glucocorticoid Receptor ◽

Negative Correlation ◽

Transcriptional Activity

Download Full-text

Differential Regulation of Estrogen-Inducible Proteolysis and Transcription by the Estrogen Receptor α N Terminus

Molecular and Cellular Biology ◽

10.1128/mcb.25.13.5417-5428.2005 ◽

2005 ◽

Vol 25 (13) ◽

pp. 5417-5428 ◽

Cited By ~ 67

Author(s):

Christopher C. Valley ◽

Raphaël Métivier ◽

Natalia M. Solodin ◽

Amy M. Fowler ◽

Mara T. Mashek ◽

...

Keyword(s):

Estrogen Receptor ◽

Transcriptional Activity ◽

Target Genes ◽

Estrogen Receptor Α ◽

Transactivation Domain ◽

Related Factors ◽

Additional Element ◽

Transactivation Domains ◽

Ubiquitin Proteasome ◽

Transcriptional Complex

ABSTRACT The ubiquitin-proteasome pathway has emerged as an important regulatory mechanism governing the activity of several transcription factors. While estrogen receptor α (ERα) is also subjected to rapid ubiquitin-proteasome degradation, the relationship between proteolysis and transcriptional regulation is incompletely understood. Based on studies primarily focusing on the C-terminal ligand-binding and AF-2 transactivation domains, an assembly of an active transcriptional complex has been proposed to signal ERα proteolysis that is in turn necessary for its transcriptional activity. Here, we investigated the role of other regions of ERα and identified S118 within the N-terminal AF-1 transactivation domain as an additional element for regulating estrogen-induced ubiquitination and degradation of ERα. Significantly, different S118 mutants revealed that degradation and transcriptional activity of ERα are mechanistically separable functions of ERα. We find that proteolysis of ERα correlates with the ability of ERα mutants to recruit specific ubiquitin ligases regardless of the recruitment of other transcription-related factors to endogenous model target genes. Thus, our findings indicate that the AF-1 domain performs a previously unrecognized and important role in controlling ligand-induced receptor degradation which permits the uncoupling of estrogen-regulated ERα proteolysis and transcription.

Download Full-text

Phosphorylation by Cyclin C/Cyclin-Dependent Kinase 2 following Mitogenic Stimulation of Murine Fibroblasts Inhibits Transcriptional Activity of LSF during G1 Progression

Molecular and Cellular Biology ◽

10.1128/mcb.00687-08 ◽

2009 ◽

Vol 29 (9) ◽

pp. 2335-2345 ◽

Cited By ~ 12

Author(s):

Utsav H. Saxena ◽

Christina M. H. Powell ◽

Jill K. Fecko ◽

Roxanne Cacioppo ◽

Hubert S. Chou ◽

...

Keyword(s):

Transcriptional Activity ◽

Target Genes ◽

Cyclin Dependent Kinase ◽

Mitogenic Stimulation ◽

Mouse Fibroblasts ◽

Cyclin C ◽

Murine Fibroblasts ◽

Interfering Rna ◽

Stimulation Of

ABSTRACT Transcription factor LSF is required for progression from quiescence through the cell cycle, regulating thymidylate synthase (Tyms) expression at the G1/S boundary. Given the constant level of LSF protein from G0 through S, we investigated whether LSF is regulated by phosphorylation in G1. In vitro, LSF is phosphorylated by cyclin E/cyclin-dependent kinase 2 (CDK2), cyclin C/CDK2, and cyclin C/CDK3, predominantly on S309. Phosphorylation of LSF on S309 is maximal 1 to 2 h after mitogenic stimulation of quiescent mouse fibroblasts. This phosphorylation is mediated by cyclin C-dependent kinases, as shown by coimmunoprecipitation of LSF and cyclin C in early G1 and by abrogation of LSF S309 phosphorylation upon suppression of cyclin C with short interfering RNA. Although mouse fibroblasts lack functional CDK3 (the partner of cyclin C in early G1 in human cells), CDK2 compensates for this absence. By transient transfection assays, phosphorylation at S309, mediated by cyclin C overexpression, inhibits LSF transactivation. Moreover, overexpression of cyclin C and CDK3 inhibits induction of endogenous Tyms expression at the G1/S transition. These results identify LSF as only the second known target (in addition to pRb) of cyclin C/CDK activity during progression from quiescence to early G1. Unexpectedly, this phosphorylation prevents induction of LSF target genes until late G1.

Download Full-text

RanBPM, a Nuclear Protein That Interacts with and Regulates Transcriptional Activity of Androgen Receptor and Glucocorticoid Receptor

Journal of Biological Chemistry ◽

10.1074/jbc.m209741200 ◽

2002 ◽

Vol 277 (50) ◽

pp. 48020-48027 ◽

Cited By ~ 64

Author(s):

Mira A. Rao ◽

Helen Cheng ◽

Alandra N. Quayle ◽

Hideo Nishitani ◽

Colleen C. Nelson ◽

...

Keyword(s):

Androgen Receptor ◽

Glucocorticoid Receptor ◽

Transcriptional Activity ◽

Nuclear Protein

Download Full-text

7-hydroxyfrullanolide, isolated from Sphaeranthus indicus, inhibits colorectal cancer cell growth by p53-dependent and -independent mechanism

Carcinogenesis ◽

10.1093/carcin/bgy176 ◽

2018 ◽

Vol 40 (6) ◽

pp. 791-804

Author(s):

Praveen Pandey ◽

Deepika Singh ◽

Mohammad Hasanain ◽

Raghib Ashraf ◽

Mayank Maheshwari ◽

...

Keyword(s):

Cell Growth ◽

Anticancer Activity ◽

Cancer Cell ◽

Molecular Mechanisms ◽

Target Genes ◽

Cancer Cell Growth ◽

Apoptotic Pathway ◽

Syngeneic Mouse Model ◽

Sphaeranthus Indicus

Abstract Sphaeranthus indicus Linn. is commonly used in Indian traditional medicine for management of multiple pathological conditions. However, there are limited studies on anticancer activity of this plant and its underlying molecular mechanisms. Here, we isolated an active constituent, 7-hydroxyfrullanolide (7-HF), from the flowers of this plant, which showed promising chemotherapeutic potential. The compound was more effective in inhibiting in vitro proliferation of colon cancers cells through G2/M phase arrest than other cancer cell lines that were used in this study. Consistent with in vitro data, 7-HF caused substantial regression of tumour volume in a syngeneic mouse model of colon cancer. The molecule triggered extrinsic apoptotic pathway, which was evident as upregulation of DR4 and DR5 expression as well as induction of their downstream effector molecules (FADD, Caspase-8). Concurrent activation of intrinsic pathway was demonstrated with loss of ΔΨm to release pro-apoptotic cytochrome c from mitochondria and activation of downstream caspase cascades (Caspase -9, -3). Loss of p53 resulted in decreased sensitivity of cells towards pro-apoptotic effect of 7-HF with increased number of viable cells indicating p53-dependent arrest of cancer cell growth. This notion was further supported with 7-HF-mediated elevation of endogenous p53 level, decreased expression of MDM2 and transcriptional upregulation of p53 target genes in apoptotic pathway. However, 7-HF was equally effective in preventing progression of HCT116 p53+/+ and p53−/− cell derived xenografts in nude mice, which suggests that differences in p53 status may not influence its in vivo efficacy. Taken together, our results support 7-HF as a potential chemotherapeutic agent and provided a new mechanistic insight into its anticancer activity.

Download Full-text