Evaluating Drosophila p53 as a Model System for Studying Cancer Mutations

Herzog, Gal; Joerger, A.C.; Shmueli, Merav D.; Fersht, Alan R.; Gazit, Ehud; Segal, Daniel

doi:10.1074/jbc.m112.417980

Cited by 13 publications

(18 citation statements)

References 45 publications

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“…Similar to mammalian p53, evidence suggests that the active Drosophila p53 transcription factor complex is a tetramer. 36,[51][52][53][54] We used the fluorescently tagged isoforms to examine whether some complexes contain both p53A and p53B. Larval brain and imaginal disc extracts were made from strains containing both the GFP-p53A and mCh-p53B BAC transgenes, GFP-p53A was immunoprecipitated (IP) with a highly efficient single chain nanobody against GFP, followed by western blotting with p53 antibodies.…”

Section: Resultsmentioning

confidence: 99%

The function of Drosophila p53 isoforms in apoptosis

et al. 2015

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The p53 protein is a major mediator of the cellular response to genotoxic stress and is a crucial suppressor of tumor formation. In a variety of organisms, p53 and its paralogs, p63 and p73, each encode multiple protein isoforms through alternative splicing, promoters, and translation start sites. The function of these isoforms in development and disease are still being defined. Here, we evaluate the apoptotic potential of multiple isoforms of the single p53 gene in the genetic model Drosophila melanogaster. Most previous studies have focused on the p53A isoform, but it has been recently shown that a larger p53B isoform can induce apoptosis when overexpressed. It has remained unclear, however, whether one or both isoforms are required for the apoptotic response to genotoxic stress. We show that p53B is a much more potent inducer of apoptosis than p53A when overexpressed. Overexpression of two newly identified short isoforms perturbed development and inhibited the apoptotic response to ionizing radiation. Analysis of physiological protein expression indicated that p53A is the most abundant isoform, and that both p53A and p53B can form a complex and co-localize to sub-nuclear compartments. In contrast to the overexpression results, new isoformspecific loss-of-function mutants indicated that it is the shorter p53A isoform, not full-length p53B, that is the primary mediator of pro-apoptotic gene transcription and apoptosis after ionizing radiation. Together, our data show that it is the shorter p53A isoform that mediates the apoptotic response to DNA damage, and further suggest that p53B and shorter isoforms have specialized functions.

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Section: Resultsmentioning

confidence: 99%

The function of Drosophila p53 isoforms in apoptosis

et al. 2015

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“…During the past decade, a wide range of real-time thermocyclers, as well as plate readers with temperature control, have been utilized to run DSF experiments, including the Stratagene M3005P [11,12], Opticon2 (MJ Research, now Biorad [13] [14]), iCycler/iQ by BioRad (an early example of use by Lo et al [15]), Rotorgene from Corbet Research [16], LightCycler 486 by Roche [17], a range of Applied Biosystems real-time PCR platforms, including the 7900 HT, 7500 Fast and StepOnePlus, and the recently launched ViiA 7 (http://www.invitrogen.com/site/us/en/home/Products-and-Services/Applications/PCR/real-time-pcr/real-time-pcr-applications/real-time-pcr-protein-analysis/protein-thermal-shift.html?icid=fr-proteinmelt), and the FluoDia T70 temperature-controlled plate reader by PTI (http://www.pti-nj.com/PlateReader/PlateReader-FluoDia.html). The main variations of the DSF technique are summarized within Table 1.…”

Section: Experimental Setup and Data Interpretationmentioning

confidence: 99%

“…The Drosophila Dmp53 has similar apoptotic functions as its human homolog and is therefore being considered as a convenient model system for studying human cancer mechanisms. The effect of point mutations in Dmp53, corresponding to cancer hot spot mutations in the human protein, on the stability and DNA binding affinity was tested through DSF [16]. The Hp53 and Dmp53 proteins had similar melting temperatures and showed similar energetic and functional responses to cancer-associated mutations, indicating general suitability of using Dmp53 as a model system for studying p53 function and discovery of p53-targeting drugs despite the low sequence similarity between the two orthologues.…”

Section: Examples Of Recent Dsf Usementioning

confidence: 99%

Recent developments in the use of differential scanning fluorometry in protein and small molecule discovery and characterization

Simeonov

2013

Expert Opinion on Drug Discovery

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Introduction Despite tremendous advances in the application of biophysical methods in drug discovery, the preponderance of instruments and techniques still require sophisticated analyses by dedicated personnel and/or large amounts of frequently hard-to-produce proteins. A technique which carries the promise of simplicity and relatively low protein consumption is the differential scanning fluorometry (DSF), wherein protein denaturation is monitored, through the use of environmentally sensitive fluorescent dye, in a temperature-ramp regime by observing the gradual exposure to the solvent of otherwise buried hydrophobic faces of protein domains. Areas covered This review describes recent developments in the field, with a special emphasis on advances published during the 2010–2013 period. Expert Opinion There has been a significant diversification of DSF applications beyond initial small molecule discovery into areas such as protein therapeutic development, formulation studies, and various mechanistic investigations, serving as a further indication of the broad penetration of the technique. In the small molecule arena, DSF has expanded towards sophisticated co-dependency MOA tests, demonstrating the wealth of information which the technique can provide. Importantly, the first public deposition of a large screening dataset may enable the use of thermal stabilization data in refining in silico models for small molecule binding.

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“…Hence, p53 was initially molded by selective pressures unrelated to cancer and only later became co‐opted for tumor suppression in long‐lived animals. If true, this inference suggests that attempts to identify ancient p53 effectors could open promising routes toward understanding adaptive functions of p53 that, when deranged, cause human diseases .…”

Section: Three Common Myths About P53mentioning

confidence: 99%

p53 in the game of transposons

Wylie

Jones

Abrams³

2016

BioEssays

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Throughout the animal kingdom, p53 genes function to restrain mobile elements and recent observations indicate that transposons become derepressed in human cancers. Together, these emerging lines of evidence suggest that cancers driven by p53 mutations could represent ‘transpospoathies’, i.e. disease states linked to eruptions of mobile elements. The transposopathy hypothesis predicts that p53 acts through conserved mechanisms to contain transposon movement and, in this way, prevents tumor formation. How transposon eruptions provoke neoplasias is not well understood but, from a broader perspective, this hypothesis also provides an attractive framework to explore unrestrained mobile elements as inciters of late-onset idiopathic disease.

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Evaluating Drosophila p53 as a Model System for Studying Cancer Mutations

Cited by 13 publications

References 45 publications

The function of Drosophila p53 isoforms in apoptosis

The function of Drosophila p53 isoforms in apoptosis

Recent developments in the use of differential scanning fluorometry in protein and small molecule discovery and characterization

p53 in the game of transposons

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