p53 Targets Simian Virus 40 Large T Antigen for Acetylation by CBP

Poulin, Danielle L.; Kung, Andrew L.; DeCaprio, James A.

doi:10.1128/jvi.78.15.8245-8253.2004

Cited by 52 publications

(60 citation statements)

References 45 publications

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“…We tested our ability to reproduce these findings using the FLS platform technology. In CV-1 cells expressing p53/EGFP/ NS(471-721), consistent with the published results obtained by other methods (27), TAg(250 -627) was sufficient to associate (i.e. colocalize) with p53 in FLS (Fig.…”

Section: Bait/egfp/ns(471-721))supporting

confidence: 91%

“…The smallest contiguous region of TAg shown to retain strong interaction with p53 comprises 377 centrally located amino acids (TAg(250 -627)) (27). Furthermore deletion of 11 amino acids within this region of full-length TAg (TAg(⌬434 -444)) has been shown to ablate the interaction with p53 (28).…”

Section: Bait/egfp/ns(471-721))mentioning

confidence: 99%

“…pSG5-wtTAg, pSG5-TAg(250 -621)/HA, pSG5-TAg(⌬434 -444), and pCDNA3-wtp53 were generous gifts from Dr. J. A. DeCaprio (Harvard Medical School) and have been described previously (27,28). EGFP/NSP5 was a kind gift of Drs.…”

Section: Methodsmentioning

confidence: 99%

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Virus-derived Platforms for Visualizing Protein Associations inside Cells

Miller

Arnold

Broering³

et al. 2007

Molecular & Cellular Proteomics

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Protein-protein associations are vital to cellular functions. Here we describe a helpful new method to demonstrate protein-protein associations inside cells based on the capacity of orthoreovirus protein NS to form large cytoplasmic inclusions, easily visualized by light microscopy, and to recruit other proteins to these structures in a specific manner. We introduce this technology by the identification of a sixth orthoreovirus protein, RNA-dependent RNA polymerase 3, that was recruited to the structures through an association with NS. We then established the broader utility of this technology by using a truncated, fluorescently tagged form of NS as a fusion platform to present the mammalian tumor suppressor p53, which strongly recruited its known interactor simian virus 40 large T antigen to the NS-derived structures. In both examples, we further localized a region of the recruited protein that is key to its recruitment. Using either endogenous p53 or a second fluorescently tagged fusion of p53 with the rotavirus NSP5 protein, we demonstrated p53 oligomerization as well as p53 association with another of its cellular interaction partners, the CREB-binding proteins, within the inclusions. Furthermore using the p53-fused fluorescent NS platform in conjunction with threecolor microscopy, we identified a ternary complex comprising p53, simian virus 40 large T antigen, and retinoblastoma protein. The new method is technically simple, uses commonly available resources, and is adaptable to high throughput formats. Molecular & Cellular Proteomics 6:1027-1038, 2007.Protein-protein associations are vital to cellular functions. Numerous methods are available for addressing whether proteins associate either directly ("interaction") or indirectly (through bridging molecules), but many of these methods can be technically demanding, can be limited by the strength of the association, or cannot resolve whether the association indeed occurs inside cells. In addition, many current objectives and strategies demand methods that are simple and adaptable to high throughput formats. In this report, we describe a new such method that is based on an unusual property of the orthoreovirus-encoded NS protein.Double-stranded RNA viruses of the taxonomic family Reoviridae (e.g. orbiviruses, orthoreoviruses, and rotaviruses) form characteristically shaped, non-membrane-bound inclusions in the cytoplasm of infected cells. These structures are variously called viral inclusion bodies (orbiviruses) (1), viral factories (orthoreoviruses) (2, 3), or viroplasms (rotaviruses) (4) and are thought to be the sites of viral RNA replication and packaging into assembly intermediates that can later mature into infectious virions. The viral factories of some orthoreoviruses (such as certain isolates of strain Type 3 Dearing (T3D) 1 ), for example, have a globular morphology (5, 6) and can attain large diameters (10 m or more) that make them easily identifiable by light microscopy (see Fig. 1a, left panel).Previous studies have identified the medium-sized (-c...

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Section: Bait/egfp/ns(471-721))supporting

confidence: 91%

Section: Bait/egfp/ns(471-721))mentioning

confidence: 99%

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Virus-derived Platforms for Visualizing Protein Associations inside Cells

Miller

Arnold

Broering³

et al. 2007

Molecular & Cellular Proteomics

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“…As a result of such screening, we identified several acetylated proteins including the SV40 large T antigen (T-Ag). Most recently, it was demonstrated that T-Ag could be acetylated by CBP (Poulin et al, 2004). However, the regulation of the T-Ag acetylation and its biological function remained obscure.…”

Section: Introductionmentioning

confidence: 99%

Regulation of SV40 large T-antigen stability by reversible acetylation

et al. 2006

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Reversible acetylation on protein lysine residues has been shown to regulate the function of both nuclear proteins such as histones and p53 and cytoplasmic proteins such as a-tubulin. To identify novel acetylated proteins, we purified several proteins by the affinity to an antiacetylated-lysine antibody from cells treated with trichostatin A (TSA). Among the proteins identified, here we report acetylation of the SV40 large T antigen (T-Ag). The acetylation site was determined to be lysine-697, which is located adjacent to the C-terminal Cdc4 phosphodegron (CPD). Overexpression of the CBP acetyltransferase acetylated T-Ag, whereas HDAC1, HDAC3 and SIRT1 bound and deacetylated T-Ag. The acetylation and deacetylation occurred independently of p53, a binding partner of T-Ag, but the acetylation was enhanced in the presence of p53. T-Ag in the cells treated with TSA and NA or the acetylation mimic mutant (K697Q) became unstable in COS-7 cells, suggesting that acetylation regulates stability of T-Ag. Indeed, NIH3T3 cells stably expressing K697Q showed decreased anchorage-independent growth compared with those expressing wild type or the K697R mutant. These results demonstrate that acetylation destabilizes T-Ag and regulates the transforming activity of T-Ag in NIH3T3 cells.

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“…All studies used low passage C2C12 (passages 3-6). P300 Ϫ/Ϫ mouse embryonic fibroblasts (28) were cultured in DMEM supplemented with 10% FBS as described previously (29).…”

Section: Methodsmentioning

confidence: 99%

Kruppel-like Factor 15 Is a Critical Regulator of Cardiac Lipid Metabolism

Prosdocimo

Anand

Liao

et al. 2014

Journal of Biological Chemistry

107

113

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Background: Metabolic homeostasis is central to normal cardiac function. The molecular mechanisms underlying metabolic plasticity in the heart remain poorly understood. Results: Kruppel-like factor 15 (KLF15) is a direct and independent regulator of myocardial lipid flux. Conclusion: KLF15 is a core component of the transcriptional circuitry that governs cardiac metabolism. Significance: This work is the first to implicate the KLF transcription factor family in cardiac metabolism.

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p53 Targets Simian Virus 40 Large T Antigen for Acetylation by CBP

Cited by 52 publications

References 45 publications

Virus-derived Platforms for Visualizing Protein Associations inside Cells

Virus-derived Platforms for Visualizing Protein Associations inside Cells

Regulation of SV40 large T-antigen stability by reversible acetylation

Kruppel-like Factor 15 Is a Critical Regulator of Cardiac Lipid Metabolism

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