Coenzyme A levels influence protein acetylation, CoAlation and 4'-phosphopantetheinylation:Expanding the impact of a metabolic nexus molecule

Coenzyme A (CoA) is a key molecule in cellular metabolism including the tricarboxylic acid cycle, fatty acid synthesis, amino acid synthesis and lipid metabolism. Moreover, CoA is required for biological processes like protein post-translational modifications (PTMs) including acylation. CoA levels affect the amount of histone acetylation and thereby modulate gene expression. A direct influence of CoA levels on other PTMs, like CoAlation and 4'-phosphopantetheinylation has been relatively less addressed and will be discussed here. Increased CoA levels are associated with increased CoAlation, whereas decreased 4'-phosphopantetheinylation is observed under circumstances of decreased CoA levels. We discuss how these two PTMs can positively or negatively influence target proteins depending on CoA levels. This review highlights the impact of CoA levels on post-translational modifications, their counteractive interplay and the far-reaching consequences thereof

Lgl regulates the hippo pathway independently of Fat/Dachs, Kibra/Expanded/Merlin and dRASSF/dSTRIPAK

In both Drosophila and mammalian systems, the Hippo (Hpo) signalling pathway controls tissue growth by inhibiting cell proliferation and promoting apoptosis. The core pathway consists of a protein kinase Hpo (MST1/2 in mammals) that is regulated by a number of upstream inputs including Drosophila Ras Association Factor, dRASSF. We have previously shown in the developing Drosophila eye epithelium that loss of the apico-basal cell polarity regulator lethal-(2)-giant-larvae (lgl), and the concomitant increase in aPKC activity, results in ectopic proliferation and suppression of developmental cell death by blocking Hpo pathway signalling. Here, we further explore how Lgl/aPKC interacts with the Hpo pathway. Deregulation of the Hpo pathway by Lgl depletion is associated with the mislocalization of Hpo and dRASSF. We demonstrate that Lgl/aPKC regulate the Hpo pathway independently of upstream inputs from Fat/Dachs and the Kibra/Expanded/Merlin complex. We show depletion of Lgl also results in accumulation and mislocalization of components of the dSTRIPAK complex, a major phosphatase complex that directly binds to dRASSF and represses Hpo activity. However, depleting dSTRIPAK components, or removal of dRASSF did not rescue the lgl−/− or aPKC overexpression phenotypes. Thus, Lgl/aPKC regulate Hpo activity by a novel mechanism, independently of dRASSF and dSTRIPAK. Surprisingly, removal of dRASSF in tissue with increased aPKC activity results in mild tissue overgrowth, indicating that in this context dRASSF acts as a tumor suppressor. This effect was independent of the Hpo and Ras Mitogen Activated Protein Kinase (MAPK) pathways, suggesting that dRASSF regulates a novel pathway to control tissue growth

The lethal giant larvae tumour suppressor mutation requires dMyc oncoprotein to promote clonal malignancy

<p>Abstract</p> <p>Background</p> <p>Neoplastic overgrowth depends on the cooperation of several mutations ultimately leading to major rearrangements in cellular behaviour. Precancerous cells are often removed by cell death from normal tissues in the early steps of the tumourigenic process, but the molecules responsible for such a fundamental safeguard process remain in part elusive. With the aim to investigate the molecular crosstalk occurring between precancerous and normal cells <it>in vivo</it>, we took advantage of the clonal analysis methods that are available in <it>Drosophila </it>for studying the phenotypes due to <it>lethal giant larvae </it>(<it>lgl</it>) neoplastic mutation induced in different backgrounds and tissues.</p> <p>Results</p> <p>We observed that <it>lgl </it>mutant cells growing in wild-type imaginal wing discs show poor viability and are eliminated by Jun N-terminal Kinase (JNK)-dependent cell death. Furthermore, they express very low levels of dMyc oncoprotein compared with those found in the surrounding normal tissue. Evidence that this is a cause of <it>lgl </it>mutant cells elimination was obtained by increasing dMyc levels in <it>lgl </it>mutant clones: their overgrowth potential was indeed re-established, with mutant cells overwhelming the neighbouring tissue and forming tumourous masses displaying several cancer hallmarks. Moreover, when <it>lgl </it>mutant clones were induced in backgrounds of slow-dividing cells, they upregulated dMyc, lost apical-basal cell polarity and were able to overgrow. Those phenotypes were abolished by reducing dMyc levels in the mutant clones, thereby confirming its key role in <it>lgl</it>-induced tumourigenesis. Furthermore, we show that the <it>eiger</it>-dependent Intrinsic Tumour Suppressor pathway plays only a minor role in eliminating <it>lgl </it>mutant cells in the wing pouch; <it>lgl</it>^-/-clonal death in this region is instead driven mainly by dMyc-induced Cell Competition.</p> <p>Conclusions</p> <p>Our results provide the first evidence that dMyc oncoprotein is required in <it>lgl </it>tumour suppressor mutant tissue to promote invasive overgrowth in larval and adult epithelial tissues. Moreover, we show that dMyc abundance inside <it>versus </it>outside the mutant clones plays a key role in driving neoplastic overgrowth.</p

North Sea Progressive Myoclonus Epilepsy is Exacerbated by Heat, A Phenotype Primarily Associated with Affected Glia

Progressive myoclonic epilepsies (PMEs) comprise a group of rare disorders of different genetic aetiologies, leading to childhood-onset myoclonus, myoclonic seizures and subsequent neurological decline. One of the genetic causes for PME, a mutation in the gene coding for Golgi SNAP receptor 2 (GOSR2), gives rise to a PME-subtype prevalent in Northern Europe and hence referred to as North Sea Progressive Myoclonic Epilepsy (NS-PME). Treatment for NS-PME, as for all PME subtypes, is symptomatic; the pathophysiology of NS-PME is currently unknown, precluding targeted therapy. Here, we investigated the pathophysiology of NS-PME. By means of chart review in combination with interviews with patients (n = 14), we found heat to be an exacerbating factor for a majority of NS-PME patients (86%). To substantiate these findings, we designed a NS-PME Drosophila melanogaster model. Downregulation of the Drosophila GOSR2-orthologue Membrin leads to heat-induced seizure-like behaviour. Specific downregulation of GOSR2/Membrin in glia but not in neuronal cells resulted in a similar phenotype, which was progressive as the flies aged and was partially responsive to treatment with sodium barbital. Our data suggest a role for GOSR2 in glia in the pathophysiology of NS-PME

CoA-dependent activation of mitochondrial acyl carrier protein links four neurodegenerative diseases

PKAN, CoPAN, MePAN, and PDH-E2 deficiency share key phenotypic features but harbor defects in distinct metabolic processes. Selective damage to the globus pallidus occurs in these genetic neurodegenerative diseases, which arise from defects in CoA biosynthesis (PKAN, CoPAN), protein lipoylation (MePAN), and pyruvate dehydrogenase activity (PDH-E2 deficiency). Overlap of their clinical features suggests a common molecular etiology, the identification of which is required to understand their pathophysiology and design treatment strategies. We provide evidence that CoA-dependent activation of mitochondrial acyl carrier protein (mtACP) is a possible process linking these diseases through its effect on PDH activity. CoA is the source for the 4 '-phosphopantetheine moiety required for the posttranslational 4 '-phosphopantetheinylation needed to activate specific proteins. We show that impaired CoA homeostasis leads to decreased 4 '-phosphopantetheinylation of mtACP. This results in a decrease of the active form of mtACP, and in turn a decrease in lipoylation with reduced activity of lipoylated proteins, including PDH. Defects in the steps of a linked CoA-mtACP-PDH pathway cause similar phenotypic abnormalities. By chemically and genetically re-activating PDH, these phenotypes can be rescued, suggesting possible treatment strategies for these diseases

Vps13 is required for timely removal of nurse cell corpses

Programmed cell death and consecutive removal of cellular remnants is essential for development. During late stages of Drosophila melanogaster oogenesis, the small somatic follicle cells that surround the large nurse cells, promote non-apoptotic nurse cell death, subsequently engulf them, and contribute to the timely removal of nurse cell corpses. Here we identify a role for Vps13 in the timely removal of nurse cell corpses downstream of developmental programmed cell death. Vps13 is an evolutionary conserved peripheral membrane protein associated with membrane contact sites and lipid transfer. Vps13 is expressed in late nurse cells and persistent nurse cell remnants are observed when Vps13 is depleted from nurse cells but not from follicle cells. Microscopic analysis revealed enrichment of Vps13 in close proximity to the plasma membrane and the endoplasmic reticulum in nurse cells undergoing degradation. Ultrastructural analysis uncovered the presence of an underlying Vps13-dependent membranous structure in close association with the plasma membrane. The newly identified structure and function suggests the presence of a Vps13-dependent process required for complete degradation of bulky remnants of dying cells

助成研究報告

textabstractIncreasing amounts of data support a role for guanine quadruplex (G4) DNA and RNA structures in various cellular processes. We stained different organisms with monoclonal antibody 1H6 specific for G4 DNA. Strikingly, immuno-electron microscopy showed exquisite specificity for heterochromatin. Polytene chromosomes from Drosophila salivary glands showed bands that co-localized with heterochromatin proteins HP1 and the SNF2 domain-containing protein SUUR. Staining was retained in SUUR knock-out mutants but lost upon overexpression of SUUR. Somatic cells in Macrostomum lignano were strongly labeled, but pluripotent stem cells labeled weakly. Similarly, germline stem cells in Drosophila ovaries were weakly labeled compared to most other cells. The unexpected presence of G4 structures in heterochromatin and the difference in G4 staining between somatic cells and stem cells with germline DNA in ciliates, flatworms, flies and mammals point to a conserved role for G4 structures in nuclear organization and cellular differentiation

Coenzyme A precursors flow from mother to zygote and from microbiome to host

Coenzyme A (CoA) is essential for metabolism and protein acetylation. Current knowledge holds that each cell obtains CoA exclusively through biosynthesis via the canonical five-step pathway, starting with pantothenate uptake. However, recent studies have suggested the presence of additional CoA-generating mechanisms, indicating a more complex system for CoA homeostasis. Here, we uncovered pathways for CoA generation through inter-organismal flows of CoA precursors. Using traceable compounds and fruit flies with a genetic block in CoA biosynthesis, we demonstrate that progeny survive embryonal and early larval development by obtaining CoA precursors from maternal sources. Later in life, the microbiome can provide the essential CoA building blocks to the host, enabling continuation of normal development. A flow of stable, long-lasting CoA precursors between living organisms is revealed. This indicates the presence of complex strategies to maintain CoA homeostasis

Supplementary data for article: Snijder, P. M.; Baratashvili, M.; Grzeschik, N. A.; Leuvenink, H. G. D.; Kuijpers, L.; Huitema, S.; Schaap, O.; Giepmans, B. N. G.; Kuipers, J.; Miljkovic, J. L.; et al. Overexpression of Cystathionine γ-Lyase Suppresses Detrimental Effects of Spinocerebellar Ataxia Type 3. Molecular Medicine 2015, 21, 758–768. https://doi.org/10.2119/molmed.2015.00221

Supplementary material for: [https://doi.org/10.2119/molmed.2015.00221]Related to published version: [http://cherry.chem.bg.ac.rs/handle/123456789/2255

Drosophila Vps13 Is Required for Protein Homeostasis in the Brain

Chorea-Acanthocytosis is a rare, neurodegenerative disorder characterized by progressive loss of locomotor and cognitive function. It is caused by loss of function mutations in the Vacuolar Protein Sorting 13A (VPS13A) gene, which is conserved from yeast to human. The consequences of VPS13A dysfunction in the nervous system are still largely unspecified. In order to study the consequences of VPS13A protein dysfunction in the ageing central nervous system we characterized a Drosophila melanogaster Vps13 mutant line. The Drosophila Vps13 gene encoded a protein of similar size as human VPS13A. Our data suggest that Vps13 is a peripheral membrane protein located to endosomal membranes and enriched in the fly head. Vps13 mutant flies showed a shortened life span and age associated neurodegeneration. Vps13 mutant flies were sensitive to proteotoxic stress and accumulated ubiquitylated proteins. Levels of Ref(2)P, the Drosophila orthologue of p62, were increased and protein aggregates accumulated in the central nervous system. Overexpression of the human Vps13A protein in the mutant flies partly rescued apparent phenotypes. This suggests a functional conservation of human VPS13A and Drosophila Vps13. Our results demonstrate that Vps13 is essential to maintain protein homeostasis in the larval and adult Drosophila brain. Drosophila Vps13 mutants are suitable to investigate the function of Vps13 in the brain, to identify genetic enhancers and suppressors and to screen for potential therapeutic targets for Chorea-Acanthocytosis