ORCID Profile
0000-0002-6271-3183
Current Organisations
Leuphana Universität Lüneburg
,
University of South Australia
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Publisher: Cold Spring Harbor Laboratory
Date: 06-2015
Abstract: Acridine orange is a cell-permeable fluorescent dye that binds to nucleic acids, resulting in an altered spectral emission. Acridine orange staining has been shown to be highly selective for apoptotic cells in Drosophila however, the precise mechanism underlying this effect is not known. Advantages of acridine orange staining include the speed and ease of the staining. But there are disadvantages: It should be performed on unfixed tissue that therefore must be examined immediately, and multiple labeling cannot be performed. Slightly different protocols for the uptake of acridine orange are required for different developmental stages. Here, we present protocols for use of acridine orange to detect apoptosis in Drosophila embryos and in larval tissue. Slight modifications might be required for other Drosophila tissues.
Publisher: Informa UK Limited
Date: 22-09-2020
Publisher: Springer Science and Business Media LLC
Date: 20-01-2021
Publisher: Springer Science and Business Media LLC
Date: 19-12-2019
Publisher: Royal Society of Chemistry (RSC)
Date: 2017
DOI: 10.1039/C7PY00345E
Abstract: A switchable polymeric 19 F magnetic resonance imaging (MRI) contrast agent was synthesised whereby the transverse ( T 2 ) relaxation times increased as a therapeutic was released from a hyperbranched polymer (HBP) scaffold.
Publisher: Rockefeller University Press
Date: 11-08-2008
Abstract: The sequential modifications of histones form the basis of the histone code that translates into either gene activation or repression. Nuclear receptors recruit a cohort of histone-modifying enzymes in response to ligand binding and regulate proliferation, differentiation, and cell death. In Drosophila melanogaster, the steroid hormone ecdysone binds its heterodimeric receptor ecdysone receptor/ultraspiracle to spatiotemporally regulate the transcription of several genes. In this study, we identify a novel cofactor, Drosophila lysine ketoglutarate reductase (dLKR)/saccharopine dehydrogenase (SDH), that is involved in ecdysone-mediated transcription. dLKR/SDH binds histones H3 and H4 and suppresses ecdysone-mediated transcription of cell death genes by inhibiting histone H3R17me2 mediated by the Drosophila arginine methyl transferase CARMER. Our data suggest that the dynamic recruitment of dLKR/SDH to ecdysone-regulated gene promoters controls the timing of hormone-induced gene expression. In the absence of dLKR/SDH, histone methylation occurs prematurely, resulting in enhanced gene activation. Consistent with these observations, the loss of dLKR/SDH in Drosophila enhances hormone-regulated gene expression, affecting the developmental timing of gene activation.
Publisher: Impact Journals, LLC
Date: 31-08-2016
Publisher: Informa UK Limited
Date: 28-10-2020
Publisher: Springer Science and Business Media LLC
Date: 04-05-2012
DOI: 10.1038/CDD.2012.43
Publisher: Springer Science and Business Media LLC
Date: 19-11-2020
Publisher: UPV/EHU Press
Date: 2015
Abstract: During Drosophila development, the steroid hormone ecdysone plays a key role in the transition from embryo into larva and then into pupa. It is during larval-pupal metamorphosis that extensive programmed cell death occurs to remove large obsolete larval tissues. During this transition, ecdysone pulses control the expression of specific transcription factors which drive the expression of key genes involved in cell death, thus spatially and temporally controlling programmed cell death. Ecdysone also controls cell death in specific larval and adult tissues. This review focuses on the current knowledge of ecdysone-mediated cell death in Drosophila.
Publisher: Elsevier BV
Date: 2002
Publisher: Springer Science and Business Media LLC
Date: 04-11-2012
DOI: 10.1038/CDD.2011.146
Publisher: Elsevier
Date: 2008
Publisher: Cold Spring Harbor Laboratory
Date: 07-2015
Abstract: A useful complement to animal studies is the use of Drosophila cell lines to analyze cell-death responses. There are numerous Drosophila cell lines available, such as S2 cells, which possess the advantages of being semi-adherent, fast growing, relatively robust, and useful for transfection and knockdown studies, whereas other lines, such as mbn2, are more suitable for analyzing hormone-induced cell death and gene expression. Drosophila cell lines are very amenable to knockdown studies as the cells take up double-stranded RNA (dsRNA) from the medium, initiating gene silencing and resulting in a high level of gene knockdown. This means that the cell lines are useful for investigating the response to death stimuli, following gene knockdown, by examining the expression of cell-death genes. This protocol describes the synthesis of dsRNA for treatment of Drosophila cells and the subsequent analysis of cell-death gene expression by quantitative real-time polymerase chain reaction (qPCR).
Publisher: Informa UK Limited
Date: 2010
Publisher: Informa UK Limited
Date: 23-06-2023
Publisher: Elsevier
Date: 2020
Publisher: Springer Science and Business Media LLC
Date: 17-04-2015
DOI: 10.1038/CDD.2015.28
Publisher: Springer Science and Business Media LLC
Date: 06-06-2018
Publisher: Elsevier BV
Date: 04-2019
DOI: 10.1016/J.BCP.2018.10.027
Abstract: Autophagy-dependent cell death is a distinct mode of regulated cell death required in a context specific manner. One of the best validated genetic models of autophagy-dependent cell death is the removal of the Drosophila larval midgut during larval-pupal transition. We have previously shown that down-regulation of growth signaling is essential for autophagy induction and larval midgut degradation. Sustained growth signaling through Ras and PI3K blocks autophagy and consequently inhibits midgut degradation. In addition, the morphogen Dpp plays an important role in regulating the correct timing of midgut degradation. Here we explore the potential roles of Hh and Wg signaling in autophagy-dependent midgut cell death. We demonstrate that Hh and Wg signaling are not involved in the regulation of autophagy-dependent cell death. However, surprisingly we found that one key component of these pathways, the Drosophila Glycogen Synthase Kinase 3, Shaggy (Sgg), may regulate midgut cell size independent of Hh and Wg signaling.
Publisher: Informa UK Limited
Date: 04-2012
DOI: 10.4161/AUTO.19496
Publisher: Elsevier BV
Date: 11-2009
Publisher: Elsevier
Date: 2017
DOI: 10.1016/BS.MIE.2016.09.089
Abstract: Drosophila is an excellent model system for studying autophagy during animal development due to the availability of genetic reagents and opportunity for in vivo cell biological analysis. The regulation and mechanism of autophagy are highly evolutionarily conserved and the role of autophagy has been characterized during various stages of Drosophila development as well as following starvation. Studies in Drosophila have revealed novel insights into the role of distinct components of the autophagy machinery. This chapter describes protocols for examining autophagy during Drosophila development. A crucial step in the induction of autophagy is the incorporation of Atg8a into the autophagosome. This can be measured as autophagic puncta using live fluorescent imaging, immunostaining, or immunoblot analysis of LC3/Atg8a processing. The level of autophagy can also be examined using other specific components of the autophagy pathway as markers detected by immunofluorescent imaging. Based on the distinct morphology of autophagy, it can also be examined by transmission electron microscopy. In addition, one of the advantages of using Drosophila as a model is the ability to undertake genetic analysis of in idual components of the autophagy machinery. Current approaches that can be used to monitor autophagy, including the overall flux and in idual steps in Drosophila melanogaster, will be discussed.
Publisher: Wiley
Date: 22-09-2015
DOI: 10.1002/GCC.22286
Abstract: Fragile site FRA16D exhibits DNA instability in cancer, resulting in diminished levels of protein from the WWOX gene that spans it. WWOX suppresses tumor growth by an undefined mechanism. WWOX participates in pathways involving aerobic metabolism and reactive oxygen species. WWOX comprises two WW domains as well as a short-chain dehydrogenase/reductase enzyme. Herein is described an in vivo genetic analysis in Drosophila melanogaster to identify functional interactions between WWOX and metabolic pathways. Altered WWOX levels modulate variable cellular outgrowths caused by genetic deficiencies of components of the mitochondrial respiratory complexes. This modulation requires the enzyme active site of WWOX, and the defective respiratory complex-induced cellular outgrowths are mediated by reactive oxygen species, dependent upon the Akt pathway and sensitive to levels of autophagy and hypoxia-inducible factor. WWOX is known to contribute to homeostasis by regulating the balance between oxidative phosphorylation and glycolysis. Reduction of WWOX levels results in diminished ability to respond to metabolic perturbation of normal cell growth. Thus, the ability of WWOX to facilitate escape from mitochondrial damage-induced glycolysis (Warburg effect) is, therefore, a plausible mechanism for its tumor suppressor activity.
Publisher: Springer Science and Business Media LLC
Date: 08-02-2019
DOI: 10.1038/S41419-019-1368-9
Abstract: The majority of developmentally programmed cell death (PCD) is mediated by caspase-dependent apoptosis however, additional modalities, including autophagy-dependent cell death, have important spatiotemporally restricted functions. Autophagy involves the engulfment of cytoplasmic components in a double membrane vesicle for delivery to the lysosome. An established model for autophagy-dependent PCD is Drosophila larval midgut removal during metamorphosis. Our previous work demonstrated that growth arrest is required to initiate autophagy-dependent midgut degradation and Target of rapamycin (Tor) limits autophagy induction. In further studies, we uncovered a role for Decapentaplegic (Dpp) in coordinating midgut degradation. Here, we provide new data to show that Dpp interacts with Tor during midgut degradation. Inhibiting Tor rescued the block in midgut degradation due to Dpp signaling. We propose that Dpp is upstream of Tor and down-regulation promotes growth arrest and autophagy-dependent midgut degradation. These findings underscore a relationship between Dpp and Tor signaling in the regulation of cell growth and tissue removal.
Publisher: Cold Spring Harbor Laboratory
Date: 07-2015
Abstract: The apoptotic machinery is highly conserved throughout evolution, and central to the regulation of apoptosis is the caspase family of cysteine proteases. Insights into the regulation and function of apoptosis in mammals have come from studies using model organisms. Drosophila provides an exceptional model system for identifying the function of conserved mechanisms regulating apoptosis, especially during development. The characteristic patterns of apoptosis during Drosophila development have been well described, as has the apoptotic response following DNA damage. The focus of this discussion is to introduce methodologies for monitoring apoptosis during Drosophila development and also in Drosophila cell lines.
Publisher: Elsevier BV
Date: 12-2013
DOI: 10.1016/J.BBAMCR.2013.06.014
Abstract: During the development of metazoans, programmed cell death (PCD) is essential for tissue patterning, removal of unwanted cells and maintaining homeostasis. In the past 20 years Drosophila melanogaster has been one of the systems of choice for studies involving developmental cell death, providing an ideal genetically tractable model of intermediary complexity between Caenorhabditis elegans and mammals. The lessons learned from studies using Drosophila indicate both the conserved nature of the many cell death pathways as well as novel and unexpected mechanisms. In this article we review the understanding of PCD during Drosophila development, highlighting the key mechanisms that are evolutionarily conserved as well as apparently unusual pathways, which indicate ergence, but provide evidence of complexity acquired during organismic evolution. This article is part of a Special Section entitled: Cell Death Pathways.
Publisher: Springer Science and Business Media LLC
Date: 22-10-2011
DOI: 10.1038/CDD.2010.130
Publisher: Cold Spring Harbor Laboratory
Date: 06-2015
Abstract: A characteristic feature of apoptosis is DNA fragmentation. This fragmentation can be detected by terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick-end labeling (TUNEL) of DNA in dying cells. Here, we present a protocol for TUNEL detection of apoptosis in Drosophila larval tissue, but these techniques can be adapted for other tissues and developmental stages.
Publisher: Cold Spring Harbor Laboratory
Date: 07-2015
Abstract: Central to the apoptotic pathway is the activation of caspases that are members of a highly conserved family of cysteine proteases. Caspases are synthesized as inactive zymogens and are generally activated by proteolytic cleavage to form the catalytically active enzyme. Caspase activity in apoptotic cells can be measured by assessing the cleavage of commercially available synthetic caspase substrates. The synthetic substrates contain a caspase cleavage site conjugated to a fluorochrome, such as 7-amino-4-methylcoumarin (AMC), or a chromophore, such as p -nitroaniline (pNA), for colorimetric detection. Here, we present a protocol for the measurement of caspase activity in Drosophila cell extracts by cleavage of the target peptide in the synthetic substrate that releases a fluorochrome or color-producing agent. The signal is measured by a spectrophotometer, with the intensity of the signal being proportional to the amount of substrate cleaved.
Publisher: Springer Science and Business Media LLC
Date: 13-12-2013
DOI: 10.1038/NCOMMS3916
Publisher: Hindawi Limited
Date: 2018
DOI: 10.1155/2018/5195416
Abstract: Autophagy is a conserved catabolic pathway that involves the engulfment of cytoplasmic components such as large protein aggregates and organelles that are delivered to the lysosome for degradation. This process is important in maintaining neuronal function and raises the possibility of a role for autophagy in neurodegenerative diseases. Alzheimer’s disease (AD) is the most prevalent form of these diseases and is characterized by the accumulation of amyloid plaques in the brain which arise due to the misfolding and aggregation of toxic peptides, including amyloid beta (A β ). There is substantial evidence from both AD patients and animal models that autophagy is dysregulated in this disease. However, it remains to be determined whether this is protective or pathogenic as there is evidence that autophagy can act to promote the degradation as well as function in the generation of toxic A β peptides. Understanding the molecular details of the extensive crosstalk that occurs between the autophagic and endolysosomal cellular pathways is essential for identifying the molecular details of amyloid toxicity. Drosophila models that express the toxic proteins that aggregate in AD have been generated and have been shown to recapitulate hallmarks of the disease. Here we focus on what is known about the role of autophagy in amyloid toxicity in AD from mammalian models and how Drosophila models can be used to further investigate AD pathogenesis.
Publisher: Springer Science and Business Media LLC
Date: 14-05-2020
Publisher: Wiley
Date: 21-10-2015
DOI: 10.1038/ICB.2014.85
Abstract: The evolutionarily conserved catabolic process of autophagy involves the degradation of cytoplasmic components through lysosomal enzymes. Basal levels of autophagy maintain cellular homeostasis and under stress conditions high levels of autophagy are induced. It is often under such stress conditions that high levels of autophagy and cell death have been observed, leading to the idea that autophagy may act as an executioner of cell death. However the notion of autophagy as a cell death mechanism has been controversial and remains mechanistically undefined. There is now growing evidence that in specific contexts autophagy can indeed facilitate cell death. The pro-death role of autophagy is however complicated due to the extensive cross-talk between different signalling pathways. This review summarises the ex les of where autophagy acts as a means of cell death and discusses the association of autophagy with the different cell death pathways.
Publisher: Cold Spring Harbor Laboratory
Date: 06-2015
Abstract: The activation of mammalian caspase-3 after proteolytic cleavage adjacent to residue Asp175 produces a large (17/19-kDa), active subunit. A commercially available antibody recognizes the large, active subunit of caspase-3 but not the full-length inactive caspase-3. This antibody has also been shown to detect active Drosophila effector caspases. Here, we present a protocol showing how this antibody can be used to detect apoptotic cells in various Drosophila tissues and developmental stages and discuss the specificity of the antibody.
Publisher: Springer Science and Business Media LLC
Date: 29-06-2019
No related grants have been discovered for Donna Denton.