scholarly journals MDM2 Mediates p300/CREB-binding Protein-associated Factor Ubiquitination and Degradation

2004 ◽  
Vol 279 (19) ◽  
pp. 20035-20043 ◽  
Author(s):  
Yetao Jin ◽  
Shelya X. Zeng ◽  
Hunjoo Lee ◽  
Hua Lu
Keyword(s):  
Signal Sequence ◽  
Binding Protein ◽  
Mrna Level ◽  
Creb Binding Protein ◽  
Leptomycin B ◽  
Associated Factor ◽  
Sequence Deletion ◽  
Localization Signal ◽  
The Stability

We recently reported that MDM2, a negative feedback regulator of the tumor suppressor p53, inhibits p300/CREB-binding protein-associated factor (PCAF)-mediated p53 acetylation. Our further study showed that MDM2 also regulates the stability of PCAF. MDM2 ubiquitinated PCAFin vitroand in cells. PCAF ubiquitination occurred at the N terminus and in the nucleus, as the nuclear localization signal sequence-deletion mutant of MDM2, which localized in the cytoplasm and degraded p53, was unable to degrade nuclear PCAF. Restriction of PCAF in the nucleus by leptomycin B did not affect MDM2-mediated PCAF degradation. Consistently, overexpression of MDM2 in p53 null cells caused the reduction of the protein level of PCAF, but not the mRNA level. Conversely, PCAF levels were higher in MDM2-deficient mousep53-/-/mdm2-/-embryonic fibroblast (MEF) cells than that in MDM2-containing MEF cells. Furthermore, MDM2 reduced the half-life of PCAF by 50%. These results demonstrate that MDM2 regulates the stability of PCAF by ubiquitinating and degrading this protein.

scholarly journals Cubitus interruptus Requires DrosophilaCREB-Binding Protein To Activate wingless Expression in theDrosophila Embryo

2000 ◽  
Vol 20 (5) ◽  
pp. 1616-1625 ◽  
Author(s):  
Yang Chen ◽  
R. H. Goodman ◽  
Sarah M. Smolik
Keyword(s):  
Transcriptional Activation ◽  
Target Genes ◽  
Binding Protein ◽  
Point Mutations ◽  
Creb Binding Protein ◽  
Wild Type ◽  
Interaction Domain ◽  
Cubitus Interruptus

ABSTRACT CREB-binding protein (CBP) serves as a transcriptional coactivator in multiple signal transduction pathways. The Drosophilahomologue of CBP, dCBP, interacts with the transcription factors Cubitus interruptus (CI), MAD, and Dorsal (DL) and functions as a coactivator in several signaling pathways during Drosophiladevelopment, including the hedgehog (hh),decapentaplegic (dpp), and Tollpathways. Although dCBP is required for the expression of thehh target genes, wingless (wg) andpatched (ptc) in vivo, and potentiatesci-mediated transcriptional activation in vitro, it is not known that ci absolutely requires dCBP for its activity. We used a yeast genetic screen to identify several ci point mutations that disrupt CI-dCBP interactions. These mutant proteins are unable to transactivate a reporter gene regulated by cibinding sites and have a lower dCBP-stimulated activity than wild-type CI. When expressed exogenously in embryos, the CI point mutants cannot activate endogenous wg expression. Furthermore, a CI mutant protein that lacks the entire dCBP interaction domain functions as a negative competitor for wild-type CI activity, and the expression of dCBP antisense RNAs can suppress CI transactivation in Kc cells. Taken together, our data suggest that dCBP function is necessary forci-mediated transactivation of wg duringDrosophila embryogenesis.

scholarly journals The CREB-binding protein inhibitor ICG-001: a promising therapeutic strategy in sporadic meningioma with NF2 mutations

2020 ◽  
Vol 2 (1) ◽  
Author(s):  
Jiaojiao Deng ◽  
Lingyang Hua ◽  
Tao Han ◽  
Mi Tian ◽  
Daijun Wang ◽  
...  
Keyword(s):  
Binding Protein ◽  
Gene Mutations ◽  
Therapeutic Effects ◽  
Creb Binding Protein ◽  
Cycle Arrest ◽  
Large Patient ◽  
Wide Range ◽  
Patient Series ◽  
Responsive Element

Abstract Background Meningiomas with Neurofibromin 2 gene mutations (NF2-mutant meningiomas) account for ~40% of the sporadic meningiomas. However, there is still no effective drug treatment for the disease. Methods Expression profile of Merlin protein was explored through immunohistochemistry in a meningioma patient cohort (n = 346). A 20-agent library covering a wide range of meningioma relevant targets was tested using meningioma cell lines IOMM-Lee (NF2 wildtype) and CH157-MN (NF2 deficient). Therapeutic effects and biological mechanisms of the identified compound, ICG-001, in NF2-mutant meningiomas were further characterized in vitro and in patient-derived xenograft (PDX) models. Results Low Merlin expression was associated with meningioma proliferation and poor clinical outcomes in a large patient series. ICG-001, a cAMP-responsive element binding (CREB)-binding protein (CBP) inhibitor, selectively suppressed tumor growth of cells with low Merlin expression. Besides, ICG-001 mediated CH157-MN and IOMM-Lee growth inhibition primarily through robust induction of the G1 cell-cycle arrest. Treatment with ICG-001 alone significantly reduced the growth of NF2-mutant xenografts in mice, as well. We also provide further evidence that ICG-001 inhibits proliferation of NF2-mutant meningioma cells at least partly through attenuating the FOXM1-mediated Wnt/β-catenin signaling. Conclusions This study highlights the importance of ligand-mediated Wnt/β-catenin signaling as well as its drugable potency in NF2-mutant meningioma.

scholarly journals The N-Terminal Domain of p73 Interacts with the CH1 Domain of p300/CREB Binding Protein and Mediates Transcriptional Activation and Apoptosis

2000 ◽  
Vol 20 (4) ◽  
pp. 1299-1310 ◽  
Author(s):  
Xiaoya Zeng ◽  
Xiaorong Li ◽  
Ashley Miller ◽  
Zhimin Yuan ◽  
Wuchao Yuan ◽  
...  
Keyword(s):  
Transcriptional Activation ◽  
Binding Protein ◽  
Small Cell Lung Carcinoma ◽  
Creb Binding Protein ◽  
Cell Lung Carcinoma ◽  
Cell Extracts ◽  
N Terminus ◽  
Mcf 7

ABSTRACT The newly identified p53 homolog p73 mimics the transcriptional function of p53. We have investigated the regulation of p73's transcriptional activity by p300/CREB binding protein (CBP). p73-p300 complexes were identified in HeLa cell extracts by cofractionation and coimmunoprecipitation assays. The p73-p300 interaction was confirmed in vitro by glutathione S-transferase–protein association assays and in vivo by coimmunoprecipitating the overexpressed p300 and p73 in human p53-free small-cell lung carcinoma H1299 or osteosarcoma Saos-2 cells. The N terminus but not the N-terminal truncation of p73 bound to the CH1 domain (amino acids [aa] 350 to 450) of p300/CBP. Accordingly, this p73 N-terminal deletion was unable to activate transcription or to induce apoptosis. Overexpression of either p300 or CBP stimulated transcription mediated by p73 but not its N-terminally deleted mutant in vivo. The N-terminal fragment from aa 19 to 597, but not the truncated fragment from aa 242 to 1700 of p300, reduced p73-mediated transcription markedly. p73-dependent transcription or apoptosis was partially impaired in either p300- or CBP-deficient human breast carcinoma MCF-7 or H1299 cells, suggesting that both coactivators mediate transcription by p73 in cells. These results demonstrate that the N terminus of p73 directly interacts with the N-terminal CH1 domain of p300/CBP to activate transcription.

scholarly journals An mRNA Stability Complex Functions with Poly(A)-Binding Protein To Stabilize mRNA In Vitro

1999 ◽  
Vol 19 (7) ◽  
pp. 4552-4560 ◽  
Author(s):  
Zuoren Wang ◽  
Nancy Day ◽  
Panayiota Trifillis ◽  
Megerditch Kiledjian
Keyword(s):  
Mrna Stability ◽  
Binding Protein ◽  
Major Protein ◽  
Globin Mrna ◽  
Ideal System ◽  
Complex Functions ◽  
Cytosolic Extract ◽  
Protein Components ◽  
The Stability

ABSTRACT The stable globin mRNAs provide an ideal system for studying the mechanism governing mammalian mRNA turnover. α-Globin mRNA stability is dictated by sequences in the 3′ untranslated region (3′UTR) which form a specific ribonucleoprotein complex (α-complex) whose presence correlates with mRNA stability. One of the major protein components within this complex is a family of two polycytidylate-binding proteins, αCP1 and αCP2. Using an in vitro-transcribed and polyadenylated α-globin 3′UTR, we have devised an in vitro mRNA decay assay which reproduces the α-complex-dependent mRNA stability observed in cells. Incubation of the RNA with erythroleukemia K562 cytosolic extract results in deadenylation with distinct intermediates containing a periodicity of approximately 30 nucleotides, which is consistent with the binding of poly(A)-binding protein (PABP) monomers. Disruption of the α-complex by sequestration of αCP1 and αCP2 enhances deadenylation and decay of the mRNA, while reconstitution of the α-complex stabilizes the mRNA. Similarly, PABP is also essential for the stability of mRNA in vitro, since rapid deadenylation resulted upon its depletion. An RNA-dependent interaction between αCP1 and αCP2 with PABP suggests that the α-complex can directly interact with PABP. Therefore, the α-complex is an mRNA stability complex in vitro which could function at least in part by interacting with PABP.

scholarly journals Analysis of the Steroid Receptor Coactivator 1 (SRC1)-CREB Binding Protein Interaction Interface and Its Importance for the Function of SRC1

2001 ◽  
Vol 21 (1) ◽  
pp. 39-50 ◽  
Author(s):  
Hilary M. Sheppard ◽  
Janet C. Harries ◽  
Sagair Hussain ◽  
Charlotte Bevan ◽  
David M. Heery
Keyword(s):  
Binding Protein ◽  
Steroid Receptor ◽  
Creb Binding Protein ◽  
Interaction Domain ◽  
Steroid Receptor Coactivator ◽  
Helix Loop Helix ◽  
Rich Domain ◽  
Interaction Interface ◽  
Acetyltransferase Domain

ABSTRACT The transcriptional activity of nuclear receptors is mediated by coactivator proteins, including steroid receptor coactivator 1 (SRC1) and its homologues and the general coactivators CREB binding protein (CBP) and p300. SRC1 contains an activation domain (AD1) which functions via recruitment of CBP and and p300. In this study, we have used yeast two-hybrid and in vitro interaction-peptide inhibition experiments to map the AD1 domain of SRC1 to a 35-residue sequence potentially containing two α-helices. We also define a 72-amino-acid sequence in CBP necessary for SRC1 binding, designated the SRC1 interaction domain (SID). We show that in contrast to SRC1, direct binding of CBP to the estrogen receptor is weak, suggesting that SRC1 functions primarily as an adaptor to recruit CBP and p300. In support of this, we show that the ability of SRC1 to enhance ligand-dependent nuclear receptor activity in transiently transfected cells is dependent upon the integrity of the AD1 region. In contrast, the putative histone acetyltransferase domain, the Per-Arnt-Sim basic helix-loop-helix domain, the glutamine-rich domain, and AD2 can each be removed without loss of ligand-induced activity. Remarkably, a construct corresponding to residues 631 to 970, which contains only the LXXLL motifs and the AD1 region of SRC1, retained strong coactivator activity in our assays.

The DNA Binding Protein H-NS Binds to and Alters the Stability of RNA in vitro and in vivo

2004 ◽  
Vol 339 (3) ◽  
pp. 505-514 ◽  
Author(s):  
Cristin C Brescia ◽  
Meenakshi K Kaw ◽  
Darren D Sledjeski
Keyword(s):  
Dna Binding ◽  
Binding Protein ◽  
Dna Binding Protein ◽  
The Stability

Abstract 322: Incubation With Histone Deacetylases 3 and 4 or p300/CREB Binding Protein Associated Factor Does Not Significantly Alter Cardiac Sarcomere Function

2012 ◽  
Vol 111 (suppl_1) ◽  
Author(s):  
John S Walker ◽  
Xiaotao Li ◽  
Lori A Walker ◽  
Todd Horn ◽  
Timothy McKinsey ◽  
...  
Keyword(s):  
Histone Deacetylases ◽  
Binding Protein ◽  
Calcium Sensitivity ◽  
Creatine Phosphate ◽  
Creb Binding Protein ◽  
Histone Acetyl Transferase ◽  
Acetyl Coa ◽  
Associated Factor ◽  
Rigor Solution ◽  
Triton X

Recent studies have suggested that treatment of skinned cardiac trabeculae with a histone deacetylase (HDAC) inhibitor increases calcium sensitivity, measured as the calcium required for half maximal force development (pCa50). The proposed mechanism is the acetylation of sarcomeric proteins by an endogenous histone acetyl transferase, p300/CREB binding protein associated factor (pCAF). As a test of this proposed mechanism, we treated skinned cardiac myocytes from 8 rat left ventricles (LV) with supra-physiological concentrations of pCAF, with Acetyl COA as an acetyl group donor, and supra-physiological concentrations of constitutively active forms of HDACs 3 and 4, and examined the force-calcium relation. Myocytes were prepared from frozen rat LV samples by gentle homogenization in rigor solution (mM: 50 Tris, 1 EGTA, 100 KCl, 2 MgCl2, 2 DTT and a protease inhibitor cocktail) containing 0.1% Triton X-100. The myocyte homogenates were washed and resuspended in relaxing solution (mM: 5 MgATP, 10 Creatine phosphate, 40 K-Propionate and 10-6 Ca-EGTA) divided into four 300 µl aliquots, One was an untreated control and three were treated with either i) HDAC3/Ncor or HDAC4 , ii) pCAF + 1mM Acetyl CoA, or ii) 1mM Acetyl CoA. Force-calcium relationships were determined in mixtures of relaxing and activating solution (mM: 5 MgATP, 10 Creatine phosphate, 0.5 K-Propionate, 10-1.5 Ca-EGTA) to give 6 calcium concentrations between 10-9 and 10-3.5 mM. While HDAC 3 or 4 treatment tended to decrease calcium sensitivity, and pCAF+Acetyl COA, and Acetyl COA alone, tended to increase calcium sensitivity compared to controls, we found no statistically significant effects of HDAC3 or 4, or pCAF on the calcium sensitivity, maximum developed force or co-operativity of the myocytes (ANOVA, n=8, P<0.05).

scholarly journals Regulation of rRNA Synthesis by TATA-Binding Protein-Associated Factor Mot1

2007 ◽  
Vol 27 (8) ◽  
pp. 2886-2896 ◽  
Author(s):  
Arindam Dasgupta ◽  
Rebekka O. Sprouse ◽  
Sarah French ◽  
Pavel Aprikian ◽  
Robert Hontz ◽  
...  
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Binding Protein ◽  
Tata Binding Protein ◽  
Rrna Synthesis ◽  
Direct Role ◽  
Pol Ii ◽  
Associated Factor ◽  
Pol I

ABSTRACT Mot1 is an essential, conserved, TATA-binding protein (TBP)-associated factor in Saccharomyces cerevisiae with well-established roles in the global control of RNA polymerase II (Pol II) transcription. Previous results have suggested that Mot1 functions exclusively in Pol II transcription, but here we report a novel role for Mot1 in regulating transcription by RNA polymerase I (Pol I). In vivo, Mot1 is associated with the ribosomal DNA, and loss of Mot1 results in decreased rRNA synthesis. Consistent with a direct role for Mot1 in Pol I transcription, Mot1 also associates with the Pol I promoter in vitro in a reaction that depends on components of the Pol I general transcription machinery. Remarkably, in addition to Mot1's role in initiation, rRNA processing is delayed in mot1 cells. Taken together, these results support a model in which Mot1 affects the rate and efficiency of rRNA synthesis by both direct and indirect mechanisms, with resulting effects on transcription activation and the coupling of rRNA synthesis to processing.

scholarly journals The Neural RNA-Binding Protein Musashi1 Translationally Regulates Mammalian numb Gene Expression by Interacting with Its mRNA

2001 ◽  
Vol 21 (12) ◽  
pp. 3888-3900 ◽  
Author(s):  
Takao Imai ◽  
Akinori Tokunaga ◽  
Tetsu Yoshida ◽  
Mitsuhiro Hashimoto ◽  
Katsuhiko Mikoshiba ◽  
...  
Keyword(s):  
Notch Signaling ◽  
In Vitro Selection ◽  
Neural Progenitor Cells ◽  
Rna Binding ◽  
Binding Protein ◽  
Mrna Level ◽  
Rna Binding Protein ◽  
Down Regulation

ABSTRACT Musashi1 (Msi1) is an RNA-binding protein that is highly expressed in neural progenitor cells, including neural stem cells. In this study, the RNA-binding sequences for Msi1 were determined by in vitro selection using a pool of degenerate 50-mer sequences. All of the selected RNA species contained repeats of (G/A)U n AGU (n = 1 to 3) sequences which were essential for Msi1 binding. These consensus elements were identified in some neural mRNAs. One of these, mammaliannumb (m-numb), which encodes a membrane-associated antagonist of Notch signaling, is a likely target of Msi1. Msi1 protein binds in vitro-transcribed m-numb RNA in its 3′-untranslated region (UTR) and binds endogenousm-numb mRNA in vivo, as shown by affinity precipitation followed by reverse transcription-PCR. Furthermore, adenovirus-induced Msi1 expression resulted in the down-regulation of endogenous m-Numb protein expression. Reporter assays using a chimeric mRNA that combined luciferase and the 3′-UTR of m-numb demonstrated that Msi1 decreased the reporter activity without altering the reporter mRNA level. Thus, our results suggested that Msi1 could regulate the expression of its target gene at the translational level. Furthermore, we found that Notch signaling activity was increased by Msi1 expression in connection with the posttranscriptional down-regulation of them-numb gene.

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