RESUMO
Mutations in glycosylation pathways, such as N-linked glycosylation, O-linked glycosylation, and GPI anchor synthesis, lead to Congenital Disorders of Glycosylation (CDG). CDG typically present with seizures, hypotonia, and developmental delay but display large clinical variability with symptoms affecting every system in the body. This variability suggests modifier genes might influence the phenotypes. Because of the similar physiology and clinical symptoms, there are likely common genetic modifiers between CDG. Here, we use evolution as a tool to identify common modifiers between CDG and glycosylation genes. Protein glycosylation is evolutionarily conserved from yeast to mammals. Evolutionary rate covariation (ERC) identifies proteins with similar evolutionary rates that indicate shared biological functions and pathways. Using ERC, we identified strong evolutionary rate signatures between proteins in the same and different glycosylation pathways. Genome-wide analysis of proteins showing significant ERC with GPI anchor synthesis proteins revealed strong signatures with ncRNA modification proteins and DNA repair proteins. We also identified strong patterns of ERC based on cellular sub-localization of the GPI anchor synthesis enzymes. Functional testing of the highest scoring candidates validated genetic interactions and identified novel genetic modifiers of CDG genes. ERC analysis of disease genes and biological pathways allows for rapid prioritization of potential genetic modifiers, which can provide a better understanding of disease pathophysiology and novel therapeutic targets.
Assuntos
Defeitos Congênitos da Glicosilação , Evolução Molecular , Glicosilação , Humanos , Defeitos Congênitos da Glicosilação/genética , Defeitos Congênitos da Glicosilação/metabolismo , Animais , Mutação , Glicosilfosfatidilinositóis/metabolismo , Glicosilfosfatidilinositóis/genética , FenótipoRESUMO
Loss of function mutations in the X-linked PIGA gene lead to PIGA-CDG, an ultra-rare congenital disorder of glycosylation (CDG), typically presenting with seizures, hypotonia, and neurodevelopmental delay. We identified two brothers (probands) with PIGA-CDG, presenting with epilepsy and mild developmental delay. Both probands carry PIGA S132C , an ultra-rare variant predicted to be damaging. Strikingly, the maternal grandfather and a great-uncle also carry PIGA S132C , but neither presents with symptoms associated with PIGA-CDG. We hypothesized genetic modifiers may contribute to this reduced penetrance. Using whole genome sequencing and pedigree analysis, we identified possible susceptibility variants found in the probands and not in carriers and possible protective variants found in the carriers and not in the probands. Candidate variants included heterozygous, damaging variants in three genes also involved directly in GPI-anchor biosynthesis and a few genes involved in other glycosylation pathways or encoding GPI-anchored proteins. We functionally tested the predicted modifiers using a Drosophila eye-based model of PIGA-CDG. We found that loss of CNTN2 , a predicted protective modifier, rescues loss of PIGA in Drosophila eye-based model, like what we predict in the family. Further testing found that loss of CNTN2 also rescues patient-relevant phenotypes, including seizures and climbing defects in Drosophila neurological models of PIGA-CDG. By using pedigree information, genome sequencing, and in vivo testing, we identified CNTN2 as a strong candidate modifier that could explain the incomplete penetrance in this family. Identifying and studying rare disease modifier genes in human pedigrees may lead to pathways and targets that may be developed into therapies.
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Stress preconditioning occurs when transient, sublethal stress events impact an organism's ability to counter future stresses. Although preconditioning effects are often noted in the literature, very little is known about the underlying mechanisms. To model preconditioning, we exposed a panel of genetically diverse Drosophila melanogaster to a sublethal heat shock and measured how well the flies survived subsequent exposure to endoplasmic reticulum (ER) stress. The impact of preconditioning varied with genetic background, ranging from dying half as fast to 4 and a half times faster with preconditioning compared to no preconditioning. Subsequent association and transcriptional analyses revealed that histone methylation, and transcriptional regulation are both candidate preconditioning modifier pathways. Strikingly, almost all subunits (7/8) in the Set1/COMPASS complex were identified as candidate modifiers of preconditioning. Functional analysis of Set1 knockdown flies demonstrated that loss of Set1 led to the transcriptional dysregulation of canonical ER stress genes during preconditioning. Based on these analyses, we propose a preconditioning model in which Set1 helps to establish an interim transcriptional "memory" of previous stress events, resulting in a preconditioned response to subsequent stress.
Assuntos
Proteínas de Drosophila , Drosophila , Animais , Drosophila/metabolismo , Drosophila melanogaster/genética , Drosophila melanogaster/metabolismo , Histonas/metabolismo , Regulação da Expressão Gênica , Metilação , Histona-Lisina N-Metiltransferase/genética , Histona-Lisina N-Metiltransferase/metabolismo , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismoRESUMO
Mutations in the phosphatidylinositol glycan biosynthesis class A (PIGA) gene cause a rare, X-linked recessive congenital disorder of glycosylation. Phosphatidylinositol glycan biosynthesis class A congenital disorder of glycosylation (PIGA-CDG) is characterized by seizures, intellectual and developmental delay, and congenital malformations. The PIGA gene encodes an enzyme involved in the first step of glycosylphosphatidylinositol (GPI) anchor biosynthesis. There are over 100 GPI-anchored proteins that attach to the cell surface and are involved in cell signaling, immunity, and adhesion. Little is known about the pathophysiology of PIGA-CDG. Here, we describe the first Drosophila model of PIGA-CDG and demonstrate that loss of PIG-A function in Drosophila accurately models the human disease. As expected, complete loss of PIG-A function is larval lethal. Heterozygous null animals appear healthy but, when challenged, have a seizure phenotype similar to what is observed in patients. To identify the cell-type specific contributions to disease, we generated neuron- and glia-specific knockdown of PIG-A. Neuron-specific knockdown resulted in reduced lifespan and a number of neurological phenotypes but no seizure phenotype. Glia-knockdown also reduced lifespan and, notably, resulted in a very strong seizure phenotype. RNA sequencing analyses demonstrated that there are fundamentally different molecular processes that are disrupted when PIG-A function is eliminated in different cell types. In particular, loss of PIG-A in neurons resulted in upregulation of glycolysis, but loss of PIG-A in glia resulted in upregulation of protein translation machinery. Here, we demonstrate that Drosophila is a good model of PIGA-CDG and provide new data resources for future study of PIGA-CDG and other GPI anchor disorders.
Assuntos
Drosophila , Glicosilfosfatidilinositóis , Animais , Humanos , Glicosilação , Fosfatidilinositóis , Fenótipo , Convulsões/genética , MutaçãoRESUMO
Several pathways and mutations must develop or be in place for the onset of cancer. Therefore, therapies should ideally target as many of these pathways as possible to improve outcomes. Combining several agents has proven to be more effective than the use of monotherapy in the treatment of renal cell carcinoma, hepatocellular carcinoma, and other cancers. Combination therapy can also include locoregional therapies such as ablation and embolization with systemic agents for synergistic effects. This review article discusses the current literature and clinical trials covering these multifactorial combination therapies in primary and metastatic liver tumors.
Assuntos
Carcinoma Hepatocelular , Embolização Terapêutica , Neoplasias Renais , Neoplasias Hepáticas , Humanos , Neoplasias Hepáticas/terapia , Neoplasias Hepáticas/patologia , Carcinoma Hepatocelular/terapia , Terapia Combinada , Neoplasias Renais/terapiaRESUMO
Mutations in the phosphatidylinositol glycan biosynthesis class A (PIGA) gene cause a rare, X-linked recessive congenital disorder of glycosylation (CDG). PIGA-CDG is characterized by seizures, intellectual and developmental delay, and congenital malformations. The PIGA gene encodes an enzyme involved in the first step of GPI anchor biosynthesis. There are over 100 GPI anchored proteins that attach to the cell surface and are involved in cell signaling, immunity, and adhesion. Little is known about the pathophysiology of PIGA-CDG. Here we describe the first Drosophila model of PIGA-CDG and demonstrate that loss of PIG-A function in Drosophila accurately models the human disease. As expected, complete loss of PIG-A function is larval lethal. Heterozygous null animals appear healthy, but when challenged, have a seizure phenotype similar to what is observed in patients. To identify the cell-type specific contributions to disease, we generated neuron- and glia-specific knockdown of PIG-A. Neuron-specific knockdown resulted in reduced lifespan and a number of neurological phenotypes, but no seizure phenotype. Glia-knockdown also reduced lifespan and, notably, resulted in a very strong seizure phenotype. RNAseq analyses demonstrated that there are fundamentally different molecular processes that are disrupted when PIG-A function is eliminated in different cell types. In particular, loss of PIG-A in neurons resulted in upregulation of glycolysis, but loss of PIG-A in glia resulted in upregulation of protein translation machinery. Here we demonstrate that Drosophila is a good model of PIGA-CDG and provide new data resources for future study of PIGA-CDG and other GPI anchor disorders.
RESUMO
Misfolded proteins in the endoplasmic reticulum (ER) elicit the ER stress response, a large transcriptional response driven by 3 well-characterized transcription factors (TFs). This transcriptional response is variable across different genetic backgrounds. One mechanism in which genetic variation can lead to transcriptional variability in the ER stress response is through altered binding and activity of the 3 main TFs: XBP1, ATF6, and ATF4. This work attempts to better understand this mechanism by first creating a computational pipeline to identify potential binding sites throughout the human genome. We utilized GTEx data sets to identify cis-eQTLs that fall within predicted TF binding sites (TFBSs). We also utilized the ClinVar database to compare the number of pathogenic vs benign variants at different positions of the binding motifs. Finally, we performed a cis-eQTL analysis on human cell lines experiencing ER stress to identify cis-eQTLs that regulate the variable ER stress response. The majority of these cis-eQTLs are unique to a given condition: control or ER stress. Some of these stress-specific cis-eQTLs fall within putative binding sites of the 3 main ER stress response TFs, providing a potential mechanism by which these cis-eQTLs might be impacting gene expression under ER stress conditions through altered TF binding. This study represents the first cis-eQTL analysis on human samples experiencing ER stress and is a vital step toward identifying the genetic components responsible for the variable ER stress response.
Assuntos
Locos de Características Quantitativas , Fatores de Transcrição , Humanos , Fatores de Transcrição/genética , Sítios de Ligação , Ligação Proteica , Variação Genética , Polimorfismo de Nucleotídeo ÚnicoRESUMO
CONTEXT: A genetic etiology accounts for the majority of unexplained primary ovarian insufficiency (POI). OBJECTIVE: We hypothesized a genetic cause of POI for a sister pair with primary amenorrhea. DESIGN: The study was an observational study. Subjects were recruited at an academic institution. SUBJECTS: Subjects were sisters with primary amenorrhea caused by POI and their parents. Additional subjects included women with POI analyzed previously (n = 291). Controls were recruited for health in old age or were from the 1000 Genomes Project (total n = 233). INTERVENTION: We performed whole exome sequencing, and data were analyzed using the Pedigree Variant Annotation, Analysis and Search Tool, which identifies genes harboring pathogenic variants in families. We performed functional studies in a Drosophila melanogaster model. MAIN OUTCOME: Genes with rare pathogenic variants were identified. RESULTS: The sisters carried compound heterozygous variants in DIS3. The sisters did not carry additional rare variants that were absent in publicly available datasets. DIS3 knockdown in the ovary of D. melanogaster resulted in lack of oocyte production and severe infertility. CONCLUSIONS: Compound heterozygous variants in highly conserved amino acids in DIS3 and failure of oocyte production in a functional model suggest that mutations in DIS3 cause POI. DIS3 is a 3' to 5' exoribonuclease that is the catalytic subunit of the exosome involved in RNA degradation and metabolism in the nucleus. The findings provide further evidence that mutations in genes important for transcription and translation are associated with POI.
Assuntos
Insuficiência Ovariana Primária , Animais , Humanos , Feminino , Insuficiência Ovariana Primária/genética , Insuficiência Ovariana Primária/patologia , Drosophila melanogaster/genética , Amenorreia/genética , Oogênese/genética , Complexo Multienzimático de Ribonucleases do ExossomoRESUMO
Stress preconditioning occurs when transient, sublethal stress events impact an organism's ability to counter future stresses. Although preconditioning effects are often noted in the literature, very little is known about the underlying mechanisms. To model preconditioning, we exposed a panel of genetically diverse Drosophila melanogaster to a sublethal heat shock and measured how well the flies survived subsequent exposure to endoplasmic reticulum (ER) stress. The impact of preconditioning varied with genetic background, ranging from dying half as fast to four and a half times faster with preconditioning compared to no preconditioning. Subsequent association and transcriptional analyses revealed that histone methylation, transcriptional regulation, and immune status are all candidate preconditioning modifier pathways. Strikingly, almost all subunits (7/8) in the Set1/COMPASS complex were identified as candidate modifiers of preconditioning. Functional analysis of Set1 knockdown flies demonstrated that loss of Set1 led to the transcriptional dysregulation of canonical ER stress genes during preconditioning. Based on these analyses, we propose a model of preconditioning in which Set1 helps to establish an interim transcriptional 'memory' of previous stress events, resulting in a preconditioned response to subsequent stress.
RESUMO
Partial loss-of-function mutations in glycosylation pathways underlie a set of rare diseases called Congenital Disorders of Glycosylation (CDGs). In particular, DPAGT1-CDG is caused by mutations in the gene encoding the first step in N-glycosylation, DPAGT1, and this disorder currently lacks effective therapies. To identify potential therapeutic targets for DPAGT1-CDG, we performed CRISPR knockout screens in Drosophila cells for genes associated with better survival and glycoprotein levels under DPAGT1 inhibition. We identified hundreds of candidate genes that may be of therapeutic benefit. Intriguingly, inhibition of the mannosyltransferase Dpm1, or its downstream glycosylation pathways, could rescue two in vivo models of DPAGT1 inhibition and ER stress, even though impairment of these pathways alone usually causes CDGs. While both in vivo models ostensibly cause cellular stress (through DPAGT1 inhibition or a misfolded protein), we found a novel difference in fructose metabolism that may indicate glycolysis as a modulator of DPAGT1-CDG. Our results provide new therapeutic targets for DPAGT1-CDG, include the unique finding of Dpm1-related pathways rescuing DPAGT1 inhibition, and reveal a novel interaction between fructose metabolism and ER stress.
Assuntos
Defeitos Congênitos da Glicosilação , Manosiltransferases , N-Acetilglucosaminiltransferases/metabolismo , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas , Defeitos Congênitos da Glicosilação/genética , Frutose , Genoma , Glicoproteínas/genética , Humanos , Manosiltransferases/genéticaRESUMO
NGLY1 deficiency, a rare disease with no effective treatment, is caused by autosomal recessive, loss-of-function mutations in the N-glycanase 1 (NGLY1) gene and is characterized by global developmental delay, hypotonia, alacrima, and seizures. We used a Drosophila model of NGLY1 deficiency to conduct an in vivo, unbiased, small molecule, repurposing screen of FDA-approved drugs to identify therapeutic compounds. Seventeen molecules partially rescued lethality in a patient-specific NGLY1 deficiency model, including multiple serotonin and dopamine modulators. Exclusive dNGLY1 expression in serotonin and dopamine neurons, in an otherwise dNGLY1 deficient fly, was sufficient to partially rescue lethality. Further, genetic modifier and transcriptomic data supports the importance of serotonin signaling in NGLY1 deficiency. Connectivity Map analysis identified glycogen synthase kinase 3 (GSK3) inhibition as a potential therapeutic mechanism for NGLY1 deficiency, which we experimentally validated with TWS119, lithium, and GSK3 knockdown. Strikingly, GSK3 inhibitors and a serotonin modulator rescued size defects in dNGLY1 deficient larvae upon proteasome inhibition, suggesting that these compounds act through NRF1, a transcription factor that is regulated by NGLY1 and regulates proteasome expression. This study reveals the importance of the serotonin pathway in NGLY1 deficiency, and serotonin modulators or GSK3 inhibitors may be effective therapeutics for this rare disease.
Assuntos
Reposicionamento de Medicamentos , Quinase 3 da Glicogênio Sintase , Animais , Defeitos Congênitos da Glicosilação , Drosophila/genética , Drosophila/metabolismo , Humanos , Peptídeo-N4-(N-acetil-beta-glucosaminil) Asparagina Amidase/deficiência , Complexo de Endopeptidases do Proteassoma/metabolismo , Doenças Raras , Serotonina/genéticaRESUMO
The genetic regulation of gene expression varies greatly across tissue-type and individuals and can be strongly influenced by the environment. Many variants, under healthy control conditions, may be silent or even have the opposite effect under diseased stress conditions. This study uses an in vivo mouse model to investigate how the effect of genetic variation changes with cellular stress across different tissues. Endoplasmic reticulum stress occurs when misfolded proteins accumulate in the endoplasmic reticulum. This triggers the unfolded protein response, a large transcriptional response which attempts to restore homeostasis. This transcriptional response, despite being a conserved, basic cellular process, is highly variable across different genetic backgrounds, making it an ideal system to study the dynamic effects of genetic variation. In this study, we sought to better understand how genetic variation alters expression across tissues, in the presence and absence of endoplasmic reticulum stress. The use of different mouse strains and their F1s allow us to also identify context-specific cis- and trans- regulatory variation underlying variable transcriptional responses. We found hundreds of genes that respond to endoplasmic reticulum stress in a tissue- and/or genotype-dependent manner. The majority of the regulatory effects we identified were acting in cis-, which in turn, contribute to the variable endoplasmic reticulum stress- and tissue-specific transcriptional response. This study demonstrates the need for incorporating environmental stressors across multiple different tissues in future studies to better elucidate the effect of any particular genetic factor in basic biological pathways, like the endoplasmic reticulum stress response.
Assuntos
Estresse do Retículo Endoplasmático , Resposta a Proteínas não Dobradas , Animais , Retículo Endoplasmático/metabolismo , Estresse do Retículo Endoplasmático/genética , Regulação da Expressão Gênica , Variação Genética , Camundongos , Resposta a Proteínas não Dobradas/genéticaRESUMO
CONTEXT: A genetic etiology likely accounts for the majority of unexplained primary ovarian insufficiency (POI). OBJECTIVE: We hypothesized that heterozygous rare variants and variants in enhanced categories are associated with POI. DESIGN: The study was an observational study. SETTING: Subjects were recruited at academic institutions. PATIENTS: Subjects from Boston (n = 98), the National Institutes of Health and Washington University (n = 98), Pittsburgh (n = 20), Italy (n = 43), and France (n = 32) were diagnosed with POI (amenorrhea with an elevated follicle-stimulating hormone level). Controls were recruited for health in old age or were from the 1000 Genomes Project (total n = 233). INTERVENTION: We performed whole exome sequencing (WES), and data were analyzed using a rare variant scoring method and a Bayes factor-based framework for identifying genes harboring pathogenic variants. We performed functional studies on identified genes that were not previously implicated in POI in a D. melanogaster model. MAIN OUTCOME: Genes with rare pathogenic variants and gene sets with increased burden of deleterious variants were identified. RESULTS: Candidate heterozygous variants were identified in known genes and genes with functional evidence. Gene sets with increased burden of deleterious alleles included the categories transcription and translation, DNA damage and repair, meiosis and cell division. Variants were found in novel genes from the enhanced categories. Functional evidence supported 7 new risk genes for POI (USP36, VCP, WDR33, PIWIL3, NPM2, LLGL1, and BOD1L1). CONCLUSIONS: Candidate causative variants were identified through WES in women with POI. Aggregating clinical data and genetic risk with a categorical approach may expand the genetic architecture of heterozygous rare gene variants causing risk for POI.
Assuntos
Insuficiência Ovariana Primária/genética , Adolescente , Adulto , Estudos de Casos e Controles , Análise Mutacional de DNA , Feminino , Heterozigoto , Humanos , Mutação , Sequenciamento do Exoma , Adulto JovemRESUMO
BACKGROUND: Numerous studies on the prognostic significance of lysine-specific demethylase 1 (LSD1) up-regulation in tumors have different outcomes. The inconsistency originated from various studies looking into the association between LSD1 and tumor cells has prompted the decision of this quantitative systematic review to decipher how up-regulated LSD1 and overall survival (OS) or recurrence-free survival (RFS) or disease-free survival (DFS) are linked in tumor patients. METHODS: Articles were searched from online databases such as Embase, Web of Science Core, PubMed, Google Scholar, and Scopus. The extraction of the hazard ratios (HR) with their 95% confidence intervals (CIs) was attained and survival data of 3151 tumor patients from 17 pieces of related research were used for this meta-analysis. RESULTS: To shed light on the link between LSD1 up-regulation and the prognosis of diverse tumors, the pooled hazard ratios (HRs) with their 95% confidence intervals (CIs) were determined. In this meta-analysis, it was observed that LSD1 up-regulation is linked with poor OS (HR = 2.08, 95% CI: 1.66-2.61, P < .01) and RFS (HR = 3.09, 95% CI: 1.81-5.26, P < .01) in tumor patients. However, LSD1 up-regulation was not linked to DFS (HR = 1.49, 95% CI: .83-2.69, P = .18) in tumor patients. The subcategory examination grouped by tumor type and ethnicity showed that LSD1 up-regulation was linked with a poor outcome in the esophageal tumor and hepatocellular carcinoma and Asian patients, respectively. For clinical-pathological factors, up-regulated LSD1 was significantly linked with Lymph node status. CONCLUSION: Despite the shortfall of the present work, this meta-analysis proposes that LSD1 up-regulation may be a prognostic biomarker for patients with tumors including esophageal tumors and hepatocellular carcinoma. We propose that large-scale studies are vital to substantiate these outcomes.
Assuntos
Biomarcadores Tumorais/genética , Histona Desmetilases/metabolismo , Neoplasias/genética , Neoplasias/mortalidade , Adolescente , Adulto , Idoso , Povo Asiático/genética , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/mortalidade , Criança , Progressão da Doença , Intervalo Livre de Doença , Neoplasias Esofágicas/genética , Neoplasias Esofágicas/mortalidade , Feminino , Humanos , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/mortalidade , Masculino , Pessoa de Meia-Idade , Prognóstico , Intervalo Livre de Progressão , Modelos de Riscos Proporcionais , Regulação para Cima/genética , Adulto JovemRESUMO
N-Glycanase 1 (NGLY1) is a cytoplasmic deglycosylating enzyme. Loss-of-function mutations in the NGLY1 gene cause NGLY1 deficiency, which is characterized by developmental delay, seizures, and a lack of sweat and tears. To model the phenotypic variability observed among patients, we crossed a Drosophila model of NGLY1 deficiency onto a panel of genetically diverse strains. The resulting progeny showed a phenotypic spectrum from 0 to 100% lethality. Association analysis on the lethality phenotype, as well as an evolutionary rate covariation analysis, generated lists of modifying genes, providing insight into NGLY1 function and disease. The top association hit was Ncc69 (human NKCC1/2), a conserved ion transporter. Analyses in NGLY1-/- mouse cells demonstrated that NKCC1 has an altered average molecular weight and reduced function. The misregulation of this ion transporter may explain the observed defects in secretory epithelium function in NGLY1 deficiency patients.
Assuntos
Defeitos Congênitos da Glicosilação/metabolismo , Peptídeo-N4-(N-acetil-beta-glucosaminil) Asparagina Amidase/deficiência , Membro 2 da Família 12 de Carreador de Soluto/metabolismo , Animais , Modelos Animais de Doenças , Drosophila melanogaster , Camundongos , Camundongos Knockout , Peptídeo-N4-(N-acetil-beta-glucosaminil) Asparagina Amidase/metabolismo , FenótipoAssuntos
COVID-19/imunologia , Células Matadoras Naturais/imunologia , Subpopulações de Linfócitos/imunologia , Antígenos CD19/imunologia , Povo Asiático , Linfócitos B/imunologia , Contagem de Células Sanguíneas , Complexo CD3/imunologia , Linfócitos T CD4-Positivos/imunologia , Antígeno CD56/imunologia , Linfócitos T CD8-Positivos/imunologia , COVID-19/epidemiologia , COVID-19/fisiopatologia , Feminino , Citometria de Fluxo , Corantes Fluorescentes/química , Proteínas Ligadas por GPI/imunologia , Humanos , Masculino , Pessoa de Meia-Idade , Pandemias , Receptores de IgG/imunologiaRESUMO
Endoplasmic reticulum (ER) stress-induced apoptosis is a primary cause and modifier of degeneration in a number of genetic disorders. Understanding how genetic variation influences the ER stress response and subsequent activation of apoptosis could improve individualized therapies and predictions of outcomes for patients. In this study, we find that the uncharacterized, membrane-bound metallopeptidase CG14516 in Drosophila melanogaster, which we rename as SUPpressor of ER stress-induced DEATH (superdeath), plays a role in modifying ER stress-induced apoptosis. We demonstrate that loss of superdeath reduces apoptosis and degeneration in the Rh1G69D model of ER stress through the JNK signaling cascade. This effect on apoptosis occurs without altering the activation of the unfolded protein response (IRE1 and PERK), suggesting that the beneficial prosurvival effects of this response are intact. Furthermore, we show that superdeath functions epistatically upstream of CDK5-a known JNK-activated proapoptotic factor in this model of ER stress. We demonstrate that superdeath is not only a modifier of this particular model, but affects the general tolerance to ER stress, including ER stress-induced apoptosis. Finally, we present evidence of Superdeath localization to the ER membrane. While similar in sequence to a number of human metallopeptidases found in the plasma membrane and ER membrane, its localization suggests that superdeath is orthologous to ERAP1/2 in humans. Together, this study provides evidence that superdeath is a link between stress in the ER and activation of cytosolic apoptotic pathways.
Assuntos
Apoptose , Proteínas de Drosophila/metabolismo , Estresse do Retículo Endoplasmático , N-Acetilgalactosaminiltransferases/metabolismo , Animais , Quinase 5 Dependente de Ciclina/genética , Proteínas de Drosophila/genética , Drosophila melanogaster , Epistasia Genética , N-Acetilgalactosaminiltransferases/genética , Resposta a Proteínas não DobradasRESUMO
The World Health Organization estimates that more than half of the world's population is at risk for vector-borne diseases, including arboviruses. Because many arboviruses are mosquito borne, investigation of the insect immune response will help identify targets to reduce the spread of arboviruses. Here, we use a genetic screening approach to identify an insulin-like receptor as a component of the immune response to arboviral infection. We determine that vertebrate insulin reduces West Nile virus (WNV) replication in Drosophila melanogaster as well as WNV, Zika, and dengue virus titers in mosquito cells. Mechanistically, we show that insulin signaling activates the JAK/STAT, but not RNAi, pathway via ERK to control infection in Drosophila cells and Culex mosquitoes through an integrated immune response. Finally, we validate that insulin priming of adult female Culex mosquitoes through a blood meal reduces WNV infection, demonstrating an essential role for insulin signaling in insect antiviral responses to human pathogens.
Assuntos
Culex , Flavivirus/fisiologia , Proteínas de Insetos/imunologia , Insulina/imunologia , Janus Quinases/imunologia , Mosquitos Vetores , Fatores de Transcrição STAT/imunologia , Transdução de Sinais/imunologia , Replicação Viral/imunologia , Animais , Linhagem Celular , Culex/imunologia , Culex/virologia , Drosophila melanogaster , Mosquitos Vetores/citologia , Mosquitos Vetores/imunologia , Mosquitos Vetores/virologiaRESUMO
Apoptosis is the primary cause of degeneration in a number of neuronal, muscular, and metabolic disorders. These diseases are subject to a great deal of phenotypic heterogeneity in patient populations, primarily due to differences in genetic variation between individuals. This creates a barrier to effective diagnosis and treatment. Understanding how genetic variation influences apoptosis could lead to the development of new therapeutics and better personalized treatment approaches. In this study, we examine the impact of the natural genetic variation in the Drosophila Genetic Reference Panel (DGRP) on two models of apoptosis-induced retinal degeneration: overexpression of p53 or reaper (rpr). We identify a number of known apoptotic, neural, and developmental genes as candidate modifiers of degeneration. We also use Gene Set Enrichment Analysis (GSEA) to identify pathways that harbor genetic variation that impact these apoptosis models, including Wnt signaling, mitochondrial metabolism, and redox homeostasis. Finally, we demonstrate that many of these candidates have a functional effect on apoptosis and degeneration. These studies provide a number of avenues for modifying genes and pathways of apoptosis-related disease.
Assuntos
Apoptose/genética , Drosophila/genética , Variação Genética , Homeostase/genética , Mitocôndrias/metabolismo , Via de Sinalização Wnt/genética , Animais , Proteínas de Drosophila/genética , Olho/anatomia & histologia , Redes Reguladoras de Genes , Estudo de Associação Genômica Ampla , Tamanho do Órgão/genética , Oxirredução , Proteína Supressora de Tumor p53/genéticaRESUMO
Endoplasmic reticulum (ER) stress is an important modifier of human disease. Genetic variation in response genes is linked to inter-individual differences in the ER stress response. However, the mechanisms and pathways by which genetic modifiers are acting on the ER stress response remain unclear. In this study, we characterize the role of the long chain fatty acid elongase Baldspot (ELOVL6) in modifying the ER stress response and disease. We demonstrate that loss of Baldspot rescues degeneration and reduces IRE1 and PERK signaling and cell death in a Drosophila model of retinitis pigmentosa and ER stress (Rh1G69D). Dietary supplementation of stearate bypasses the need for Baldspot activity. Finally, we demonstrate that Baldspot regulates the ER stress response across different tissues and induction methods. Our findings suggest that ELOVL6 is a promising target in the treatment of not only retinitis pigmentosa, but a number of different ER stress-related disorders.