A gain-of-function screening reveals dAnkmy2 as a potential mediator of lifespan extension and oxidative stress resistance in Drosophila melanogaster.

Membrane Deformation Ability of the Ankyrin Repeat and KH Domain-Containing Protein 1 (ANKHD1) and Its Involvement in the Early Endosome Enlargement

Uncovering New Functions for MicroRNAs in Caenorhabditis elegans

BackgroundAerobic organisms are susceptible to damage by reactive oxygen species. Oxidative stress resistance is a quantitative trait with population variation attributable to the interplay between genetic and environmental factors. Drosophila melanogaster provides an ideal system to study the genetics of variation for resistance to oxidative stress.Methods and FindingsWe used 167 wild-derived inbred lines of the Drosophila Genetic Reference Panel for a genome-wide association study of acute oxidative stress resistance to two oxidizing agents, paraquat and menadione sodium bisulfite. We found significant genetic variation for both stressors. Single nucleotide polymorphisms (SNPs) associated with variation in oxidative stress resistance were often sex-specific and agent-dependent, with a small subset common for both sexes or treatments. Associated SNPs had moderately large effects, with an inverse relationship between effect size and allele frequency. Linear models with up to 12 SNPs explained 67–79% and 56–66% of the phenotypic variance for resistance to paraquat and menadione sodium bisulfite, respectively. Many genes implicated were novel with no known role in oxidative stress resistance. Bioinformatics analyses revealed a cellular network comprising DNA metabolism and neuronal development, consistent with targets of oxidative stress-inducing agents. We confirmed associations of seven candidate genes associated with natural variation in oxidative stress resistance through mutational analysis.ConclusionsWe identified novel candidate genes associated with variation in resistance to oxidative stress that have context-dependent effects. These results form the basis for future translational studies to identify oxidative stress susceptibility/resistance genes that are evolutionary conserved and might play a role in human disease.

Gender differences in lifespan and aging are known across species. Sex differences in longevity within a species can be useful to understand sex-specific aging. Drosophila melanogaster is a good model to study the problem of sex differences in longevity since females are longer lived than males. There is evidence that stress resistance influences longevity. The objective of this study was to investigate if there is a relationship between sex differences in longevity and oxidative stress resistance in D. melanogaster. We observed a progressive age-dependent decrease in the activity of SOD and catalase, major antioxidant enzymes involved in defense mechanisms against oxidative stress in parallel to the increased ROS levels over time. Longer-lived females showed lower ROS levels and higher antioxidant enzymes than males as a function of age. Using ethanol as a stressor, we have shown differential susceptibility of the sexes to ethanol wherein females exhibited higher resistance to ethanol-induced mortality and locomotor behavior compared to males. Our results show strong correlation between sex differences in oxidative stress resistance, antioxidant defenses and longevity. The study suggests that higher antioxidant defenses in females may confer resistance to oxidative stress, which could be a factor that influences sex-specific aging in D. melanogaster.

The PTEN tumour suppressor encodes a phosphatase, and its daf-18 orthologue in Caenorhabditis elegans negatively regulates the insulin/IGF-1 DAF-2 receptor pathway that influences lifespan in worms and other species. In order to identify new DAF-18 regulated pathways involved in aging, we initiated a candidate RNAi feeding screen for clones that lengthen lifespan. Here, we report that smg-1 inactivation increases average lifespan in a daf-18 dependent manner. Genetic analysis is consistent with SMG-1 acting at least in part in parallel to the canonical DAF-2 receptor pathway, but converging on the transcription factor DAF-16/FOXO. SMG-1 is a serine-threonine kinase which plays a conserved role in nonsense-mediated mRNA decay (NMD) in worms and mammals. In addition, human SMG-1 has also been implicated in the p53-mediated response to genotoxic stress. The effect of smg-1 inactivation on lifespan appears to be unrelated to its NMD function, but requires the p53 tumour suppressor orthologue cep-1. Furthermore, smg-1 inactivation confers a resistance to oxidative stress in a daf-18-, daf-16- and cep-1-dependent manner. We propose that the role of SMG-1 in lifespan regulation is at least partly dependent on its function in oxidative stress resistance. Taken together, our results unveil a novel role for SMG-1 in lifespan regulation.

Glucose is the preferred carbon and energy source in prokaryotes, unicellular eukaryotes, and metazoans. However, excess of glucose has been associated with several diseases, including diabetes and the less understood process of aging. On the contrary, limiting glucose (i.e., calorie restriction) slows aging and age-related diseases in most species. Understanding the mechanism by which glucose limits life span is therefore important for any attempt to control aging and age-related diseases. Here, we use the yeast Schizosaccharomyces pombe as a model to study the regulation of chronological life span by glucose. Growth of S. pombe at a reduced concentration of glucose increased life span and oxidative stress resistance as reported before for many other organisms. Surprisingly, loss of the Git3 glucose receptor, a G protein-coupled receptor, also increased life span in conditions where glucose consumption was not affected. These results suggest a role for glucose-signaling pathways in life span regulation. In agreement, constitutive activation of the Gα subunit acting downstream of Git3 accelerated aging in S. pombe and inhibited the effects of calorie restriction. A similar pro-aging effect of glucose was documented in mutants of hexokinase, which cannot metabolize glucose and, therefore, are exposed to constitutive glucose signaling. The pro-aging effect of glucose signaling on life span correlated with an increase in reactive oxygen species and a decrease in oxidative stress resistance and respiration rate. Likewise, the anti-aging effect of both calorie restriction and the Δgit3 mutation was accompanied by increased respiration and lower reactive oxygen species production. Altogether, our data suggest an important role for glucose signaling through the Git3/PKA pathway to regulate S. pombe life span.

Stress resistance is associated with longevity in Drosophila melanogaster and other model organisms used for genetic research. The present study tests for oxidative stress resistance in one set of lines selected for late-life reproduction and extended longevity. Both females and males from the selected lines were appreciably more resistant to oxidative stress than were flies from the control lines. A relative increase in oxidative stress resistance is a correlated response to selection in this laboratory selection experiment. Increased oxidative stress resistance appears to be a relatively robust correlated response to laboratory selection for late-life reproduction and extended longevity.

MAPK are activated by and orchestrate responses to multiple, diverse stimuli. Although these responses involve the increased phosphorylation of substrate effector proteins, e.g. transcription factors, the mechanisms by which responses are tailored to particular stimuli are unclear. In the fission yeast Schizosaccharomyces pombe, the Sty1 MAPK is crucial for changes in gene expression that allow adaptation to many forms of environmental stress. Here, we have identified two cysteine residues in Sty1, Cys-153 and Cys-158, that are important for hydrogen peroxide-induced gene expression and oxidative stress resistance but not for other functions of Sty1. Many Sty1-dependent changes in gene expression are mediated by the Atf1 transcription factor. In response to stress, Sty1 increases Atf1 levels by (i) promoting increases in atf1 mRNA and by (ii) directly phosphorylating and stabilizing Atf1 protein. Although dispensable for phosphorylation and stabilization of Atf1 protein, we find that both Cys-153 and Cys-158 are required for increases in atf1 mRNA levels and Atf1-dependent gene expression in response to hydrogen peroxide but not osmotic stress. Indeed, our data indicate that oxidation of Sty1, by formation of a disulfide bond between Cys-153 and Cys-158, is important for maintaining atf1 mRNA stability at high concentrations of hydrogen peroxide. Together, these data reveal that redox regulation of cysteine thiols in Sty1 is involved in a stress-specific mechanism regulating transcriptional responses to oxidative stress. Intriguingly, the conservation of these cysteine residues in other MAPK raises the possibility that similar mechanisms may ensure appropriate responses to hydrogen peroxide in other eukaryotes.

Role of ANKHD1/LINC00346/ZNF655 Feedback Loop in Regulating the Glioma Angiogenesis via Staufen1-Mediated mRNA Decay

BioTechniquesVol. 27, No. 2 BenchmarksOpen AccessIdentifying Mutations in Drosophila Genes by Direct Sequencing of PCR ProductsErin M. Schlag & David A. WassarmanErin M. Schlag*Address correspondence to Erin M. Schlag, Cell Biology and Metabolism Br., National Institute of Child Health and Human Dev., Bldg. 18T, Rm. 101, National Institutes of Health, Bethesda, MD 20892, USA. Internet: E-mail Address: eschlag@helix.nih.govNational Institute of Child Health and Human Development, NIH, Bethesda, MD, USASearch for more papers by this author & David A. WassarmanNational Institute of Child Health and Human Development, NIH, Bethesda, MD, USASearch for more papers by this authorPublished Online:22 Aug 2018https://doi.org/10.2144/99272bm09AboutSectionsPDF/EPUB ToolsAdd to favoritesDownload CitationsTrack Citations ShareShare onFacebookTwitterLinkedInRedditEmail "Identifying Mutations in Drosophila Genes by Direct Sequencing of PCR Products." , 27(2), pp. 262–264FiguresReferencesRelatedDetailsCited ByInter-individual and intragenomic variations in the ITS region of Clonorchis sinensis (Trematoda: Opisthorchiidae) from Russia and VietnamInfection, Genetics and Evolution, Vol. 55Characterising functionally important and ecologically meaningful genetic diversity using a candidate gene approach21 September 2008 | Genetica, Vol. 138, No. 4echinoid mutants exhibit neurogenic phenotypes and show synergistic interactions with the Notch signaling pathwayDevelopment, Vol. 130, No. 25MASK, a large ankyrin repeat and KH domain-containing protein involved in Drosophila receptor tyrosine kinase signalingDevelopment, Vol. 129, No. 1The RNA-binding protein Tsunagi interacts with Mago Nashi to establish polarity and localize oskar mRNA during Drosophila oogenesis1 November 2001 | Genes & Development, Vol. 15, No. 21Developmental and Transcriptional Consequences of Mutations in Drosophila TAFII60Molecular and Cellular Biology, Vol. 21, No. 20Clinical Use of Capillary PCR To Diagnose Mycoplasma PneumoniaJournal of Clinical Microbiology, Vol. 38, No. 4TAF250 is required for multiple developmental events in Drosophila1 February 2000 | Proceedings of the National Academy of Sciences, Vol. 97, No. 3 Vol. 27, No. 2 Follow us on social media for the latest updates Metrics Downloaded 114 times History Published online 22 August 2018 Published in print August 1999 Information© 2018 Author(s)PDF download

A recent genome-wide pharmacogenomics study showed that the rs7968606 single-nucleotide polymorphism (SNP) of the ankyrin repeat and sterile alpha motif domain-containing protein 1B (ANKS1B) gene approached the threshold of statistical significance. The aim of this study was to determine the association between the rs7968606 SNP of ANKS1B and the treatment response to amisulpride in schizophrenia patients. In total, 154 participants were enrolled from six university hospitals in Korea. All the subjects were interviewed before and after 6weeks of amisulpride treatment with the aid of the positive and negative syndrome scale and the clinical global impression-severity scale. Genotyping for the rs7968606 SNP of ANKS1B was performed in 101 subjects. Both the decrease (t=-2.067, p=0.041) and improvement rate (t=-1.990, p=0.049) in the positive and negative syndrome scale general score differed significantly between T-allele carriers and noncarriers of this polymorphism after 6weeks of amisulpride treatment. To the best of our knowledge, this is the first genetic association study of the relationship between the rs7968606 SNP of ANKS1B and the response of schizophrenia patients to treatment with amisulpride. Future larger-scale studies involving more SNPs of ANKS1B will improve the understanding of the pharmacogenetics underlying the treatment responses to amisulpride.

HBV infection is challenging to cure due to the persistence of viral covalently closed circular viral DNA (cccDNA). The dedicator of cytokinesis 11 (DOCK11) is recognized as a guanine nucleotide exchange factor (GEF) for CDC42 that has been reported to be required for HBV persistence. DOCK11 is expressed in both the cytoplasm and nucleus of human hepatocytes and is functionally associated with retrograde trafficking proteins Arf-GAP with GTPase domain, ankyrin repeat, and pleckstrin homology domain-containing protein 2 (AGAP2), and ADP-ribosylation factor 1 (ARF1), together with the HBV capsid, in the trans-Golgi network (TGN). This opens an alternative retrograde trafficking route for HBV from early endosomes (EEs) to the TGN and then to the endoplasmic reticulum (ER), thereby avoiding lysosomal degradation. DOCK11 also facilitates the association of cccDNA with H3K4me3 and RNA Pol II for activating cccDNA transcription. In addition, DOCK11 plays a crucial role in the host DNA repair system, being essential for cccDNA synthesis. This function can be inhibited by 10M-D42AN, a novel DOCK11-binding peptide, leading to the suppression of HBV replication both in vitro and in vivo. Treatment with a combination of 10M-D42AN and entecavir may represent a promising therapeutic strategy for patients with chronic hepatitis B (CHB). Consequently, DOCK11 may be seen as a potential candidate molecule in the development of molecularly targeted drugs against CHB.

BTB/POZ domain-containing proteins are increasingly recognized for their context-dependent roles in cancer, acting as either tumor suppressors or oncogenic drivers depending on the cancer type. Among them, the function of ankyrin repeat and BTB domain-containing protein 2 (ABTB2) in pancreatic ductal adenocarcinoma (PDAC), a highly lethal malignancy, has remained unexplored. In this study, we employed comprehensive functional genomics approaches—siRNA/shRNA knockdown, CRISPR-Cas9 knockout, plasmid-based overexpression, and a Cre-LoxP transgenic mouse model—to systemically modulate ABTB2 expression in human and murine PDAC cell lines, as well as in the KPC mouse model of PDAC. Our gain- and loss-of-function studies revealed that ABTB2 plays a pivotal tumor-suppressive role, significantly impairing PDAC cell oncogenicity in vitro and tumorigenesis in vivo. Importantly, therapeutic targeting of ABTB2 using adeno-associated virus serotype 2 (AAV2) and lipid nanoparticles (LNPs) demonstrated marked anti-tumor efficacy and synergized with 5-fluorouracil (5-FU) to enhance treatment outcomes. Transcriptomic analysis, immunoprecipitation, and functional assays demonstrated that ABTB2 interacts with tumour necrosis factor receptor-associated protein 1 (TRAP1), promoting its ubiquitin-dependent degradation and thereby suppressing key oncogenic Wnt/β-catenin and PI3K/Akt signaling pathways. Notably, TRAP1 inhibitors are currently in phase I clinical trials as potential anticancer agents. Our findings provide mechanistic insight and underscore the ABTB2/TRAP1 axis as a promising therapeutic target for PDAC treatment.

Eph receptors and ephrins have been implicated in a variety of cellular processes, including morphology and motility, because of their ability to modulate intricate signaling networks. Here we show that the phosphotyrosine binding (PTB) domain-containing proteins AIDA-1b and Odin are tightly associated with the EphA8 receptor in response to ligand stimulation. Both AIDA-1b and Odin belong to the ankyrin repeat and sterile alpha motif domain-containing (Anks) protein family. The PTB domain of Anks family proteins is crucial for their association with the juxtamembrane domain of EphA8, whereas EphA8 tyrosine kinase activity is not required for this protein-protein interaction. In addition, we found that Odin is a more physiologically relevant partner of EphA8 in mammalian cells. Interestingly, overexpression of the Odin PTB domain alone attenuated EphA8-mediated inhibition of cell migration in HEK293 cells, suggesting that it acts as a dominant-negative mutant of the endogenous Odin protein. More importantly, small interfering RNA-mediated Odin silencing significantly diminished ephrinA5-induced EphA8 signaling effects, which inhibit cell migration in HEK293 cells and retract growing neurites of Neuro2a cells. Taken together, our findings support a possible function for Anks family proteins as scaffolding proteins of the EphA8 signaling pathway.

Ankyrin repeat and single KH domain-containing protein 1 (ANKHD1) is a large, scaffolding protein composed of two stretches of ankyrin repeat domains that mediate protein–protein interactions and a KH domain that mediates RNA or single-stranded DNA binding. ANKHD1 interacts with proteins in several crucial signalling pathways, including receptor tyrosine kinase, JAK/STAT, mechanosensitive Hippo (YAP/TAZ), and p21. Studies into the role of ANKHD1 in cancer cell lines demonstrate a crucial role in driving uncontrolled cellular proliferation and growth, enhanced tumorigenicity, cell cycle progression through the S phase, and increased epithelial-to-mesenchymal transition. Furthermore, at a clinical level, the increased expression of ANKHD1 has been associated with greater tumour infiltration, increased metastasis, and larger tumours. Elevated ANKHD1 resulted in poorer prognosis, more aggressive growth, and a decrease in patient survival in numerous cancer types. This review aims to gather the current knowledge about ANKHD1 and explore its molecular properties and functions, focusing on the protein’s role in cancer at both a cellular and clinical level.