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Lysosomal storage diseases result in various developmental and physiological complications, including cachexia. To study the causes for the negative energy balance associated with cachexia, we assessed the impact of sulfamidase deficiency and heparan sulfate storage on energy homeostasis and metabolism in a mouse model of type IIIa mucopolysaccharidosis (MPS IIIa, Sanfilippo A syndrome). At 12-weeks of age, MPS IIIa mice exhibited fasting and postprandial hypertriglyceridemia compared with wildtype mice, with a reduction of white and brown adipose tissues. Partitioning of dietary [3H]triolein showed a marked increase in intestinal uptake and secretion, whereas hepatic production and clearance of triglyceride-rich lipoproteins did not differ from wildtype controls. Uptake of dietary triolein was also elevated in brown adipose tissue (BAT), and notable increases in beige adipose tissue occurred, resulting in hyperthermia, hyperphagia, hyperdipsia, and increased energy expenditure. Furthermore, fasted MPS IIIa mice remained hyperthermic when subjected to low temperature but became cachexic and profoundly hypothermic when treated with a lipolytic inhibitor. We demonstrated that the reliance on increased lipid fueling of BAT was driven by a reduced ability to generate energy from stored lipids within the depot. These alterations arose from impaired autophagosome-lysosome fusion, resulting in increased mitochondria content in beige and BAT. Finally, we show that increased mitochondria content in BAT and postprandial dyslipidemia was partially reversed upon 5-week treatment with recombinant sulfamidase. We hypothesize that increased BAT activity and persistent increases in energy demand in MPS IIIa mice contribute to the negative energy balance observed in patients with MPS IIIa.
Assuntos
Hipertrigliceridemia , Mucopolissacaridose III , Tecido Adiposo Marrom/metabolismo , Animais , Caquexia , Camundongos , Mitofagia , Mucopolissacaridose III/metabolismo , Mucopolissacaridose III/terapia , TrioleínaRESUMO
Children with autism have an elevated frequency of large, rare copy number variants (CNVs). However, the global load of deletions or duplications, per se, and their size, location and relationship to clinical manifestations of autism have not been documented. We examined CNV data from 516 individuals with autism or typical development from the population-based Childhood Autism Risks from Genetics and Environment (CHARGE) study. We interrogated 120 regions flanked by segmental duplications (genomic hotspots) for events >50 kbp and the entire genomic backbone for variants >300 kbp using a custom targeted DNA microarray. This analysis was complemented by a separate study of five highly dynamic hotspots associated with autism or developmental delay syndromes, using a finely tiled array platform (>1 kbp) in 142 children matched for gender and ethnicity. In both studies, a significant increase in the number of base pairs of duplication, but not deletion, was associated with autism. Significantly elevated levels of CNV load remained after the removal of rare and likely pathogenic events. Further, the entire CNV load detected with the finely tiled array was contributed by common variants. The impact of this variation was assessed by examining the correlation of clinical outcomes with CNV load. The level of personal and social skills, measured by Vineland Adaptive Behavior Scales, negatively correlated (Spearman's r = -0.13, P = 0.034) with the duplication CNV load for the affected children; the strongest association was found for communication (P = 0.048) and socialization (P = 0.022) scores. We propose that CNV load, predominantly increased genomic base pairs of duplication, predisposes to autism.
Assuntos
Transtorno Autístico/genética , Variações do Número de Cópias de DNA , Estudos de Casos e Controles , Criança , Pré-Escolar , Feminino , Humanos , Masculino , Análise de Sequência com Séries de Oligonucleotídeos , Duplicações Segmentares Genômicas , Deleção de SequênciaRESUMO
Mutations in presenilin-1 (PSEN1) are the most common cause of familial, early-onset Alzheimer's disease (AD), typically producing cognitive deficits in the fourth decade. A variant of APOE, APOE3 Christchurch (APOE3ch) , was found associated with protection from both cognitive decline and Tau accumulation in a 70-year-old bearing the disease-causing PSEN1-E280A mutation. The amino acid change in ApoE3ch is within the heparan sulfate (HS) binding domain of APOE, and purified APOEch showed dramatically reduced affinity for heparin, a highly sulfated form of HS. The physiological significance of ApoE3ch is supported by studies of a mouse bearing a knock-in of this human variant and its effects on microglia reactivity and Aß-induced Tau deposition. The studies reported here examine the function of heparan sulfate-modified proteoglycans (HSPGs) in cellular and molecular pathways affecting AD-related cell pathology in human cell lines and mouse astrocytes. The mechanisms of HSPG influences on presenilin- dependent cell loss and pathology were evaluated in Drosophila using knockdown of the presenilin homolog, Psn , together with partial loss of function of sulfateless (sfl) , a homolog of NDST1 , a gene specifically affecting HS sulfation. HSPG modulation of autophagy, mitochondrial function, and lipid metabolism were shown to be conserved in cultured human cell lines, Drosophila , and mouse astrocytes. RNAi of Ndst1 reduced intracellular lipid levels in wild-type mouse astrocytes or those expressing humanized variants of APOE, APOE3 , and APOE4 . RNA-sequence analysis of human cells deficient in HS synthesis demonstrated effects on the transcriptome governing lipid metabolism, autophagy, and mitochondrial biogenesis and showed significant enrichment in AD susceptibility genes identified by GWAS. Neuron-directed knockdown of Psn in Drosophila produced cell loss in the brain and behavioral phenotypes, both suppressed by simultaneous reductions in sfl mRNA levels. Abnormalities in mitochondria, liposome morphology, and autophagosome-derived structures in animals with Psn knockdown were also rescued by simultaneous reduction of sfl. sfl knockdown reversed Psn- dependent transcript changes in genes affecting lipid transport, metabolism, and monocarboxylate carriers. These findings support the direct involvement of HSPGs in AD pathogenesis.
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We examined the function of heparan-sulfate-modified proteoglycans (HSPGs) in pathways affecting Alzheimer disease (AD)-related cell pathology in human cell lines and mouse astrocytes. Mechanisms of HSPG influences on presenilin-dependent cell loss were evaluated in Drosophila using knockdown of the presenilin homolog, Psn, together with partial loss-of-function of sulfateless (sfl), a gene specifically affecting HS sulfation. HSPG modulation of autophagy, mitochondrial function, and lipid metabolism were shown to be conserved in human cell lines, Drosophila, and mouse astrocytes. RNA interference (RNAi) of Ndst1 reduced intracellular lipid levels in wild-type mouse astrocytes or those expressing humanized variants of APOE, APOE3, and APOE4. Neuron-directed knockdown of Psn in Drosophila produced apoptosis and cell loss in the brain, phenotypes suppressed by reductions in sfl expression. Abnormalities in mitochondria, liposomes, and autophagosome-derived structures in animals with Psn knockdown were also rescued by reduction of sfl. These findings support the direct involvement of HSPGs in AD pathogenesis.
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Heparan sulfate modified proteins or proteoglycans (HSPGs) are an abundant class of cell surface and extracellular matrix molecules. They serve important co-receptor functions in the regulation of signaling as well as membrane trafficking. Many of these activities directly affect processes associated with neurodegeneration including uptake and export of Tau protein, disposition of Amyloid Precursor Protein-derived peptides, and regulation of autophagy. In this review we focus on the impact of HSPGs on autophagy, membrane trafficking, mitochondrial quality control and biogenesis, and lipid metabolism. Disruption of these processes are a hallmark of Alzheimer's disease (AD) and there is evidence that altering heparan sulfate structure and function could counter AD-associated pathological processes. Compromising presenilin function in several systems has provided instructive models for understanding the molecular and cellular underpinnings of AD. Disrupting presenilin function produces a constellation of cellular deficits including accumulation of lipid, disruption of autophagosome to lysosome traffic and reduction in mitochondrial size and number. Inhibition of heparan sulfate biosynthesis has opposing effects on all these cellular phenotypes, increasing mitochondrial size, stimulating autophagy flux to lysosomes, and reducing the level of intracellular lipid. These findings suggest a potential mechanism for countering pathology found in AD and related disorders by altering heparan sulfate structure and influencing cellular processes disrupted broadly in neurodegenerative disease. Vertebrate and invertebrate model systems, where the cellular machinery of autophagy and lipid metabolism are conserved, continue to provide important translational guideposts for designing interventions that address the root cause of neurodegenerative pathology.
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Mutations in human Exostosin genes (EXTs) confer a disease called Hereditary Multiple Exostoses (HME) that affects 1 in 50,000 among the general population. Patients with HME have a short stature and develop osteochondromas during childhood. Here we show that two zebrafish mutants, dackel (dak) and pinscher (pic), have cartilage defects that strongly resemble those seen in HME patients. We have previously determined that dak encodes zebrafish Ext2. Positional cloning of pic reveals that it encodes a sulphate transporter required for sulphation of glycans (Papst1). We show that although both dak and pic are required during cartilage morphogenesis, they are dispensable for chondrocyte and perichondral cell differentiation. They are also required for hypertrophic chondrocyte differentiation and osteoblast differentiation. Transplantation analysis indicates that dak(-/-) cells are usually rescued by neighbouring wild-type chondrocytes. In contrast, pic(-/-) chondrocytes always act autonomously and can disrupt the morphology of neighbouring wild-type cells. These findings lead to the development of a new model to explain the aetiology of HME.
Assuntos
Proteínas de Transporte de Ânions/genética , Regulação da Expressão Gênica no Desenvolvimento , N-Acetilglucosaminiltransferases/genética , Osteogênese/genética , Proteínas de Peixe-Zebra/genética , Peixe-Zebra/crescimento & desenvolvimento , Animais , Proteínas de Transporte de Ânions/fisiologia , Clonagem Molecular , Embrião não Mamífero , Marcadores Genéticos , Homozigoto , Perda de Heterozigosidade , Repetições de Microssatélites , Modelos Animais , Mutação , N-Acetilglucosaminiltransferases/fisiologia , Osteogênese/fisiologia , Mapeamento Físico do Cromossomo , RNA Mensageiro/metabolismo , Peixe-Zebra/embriologia , Peixe-Zebra/genética , Proteínas de Peixe-Zebra/fisiologiaRESUMO
Autophagy is a fundamental cellular process discovered as a response to nutrient deprivation. It provides the cellular and molecular machinery for catabolism of cellular constituents, generating energy and providing building blocks for continued survival. However, autophagy does much more than provide an entry into catabolic pathways, it provides a mechanism for intracellular quality control, removing damaged organelles and misfolded proteins, processes critical for cellular health. Autophagy serves as a counterpoint to cell growth and anabolic events, activated when growth is not possible or is suppressed. Hence, there is an inherent antagonism between autophagy and growth. Heparan sulfate modified proteins are important co-receptors that generally promote growth factor activity and are therefore positioned within signaling networks that inhibit, or negatively regulate autophagy levels. This review summarizes evidence that heparan sulfate modified proteins provide an evolutionarily conserved inhibitory modulation of autophagy that can have profound effects on cell physiology and organismal responses to stress.
Assuntos
Autofagia , Heparitina Sulfato , Controle de Qualidade , Transdução de SinaisRESUMO
Heparan sulfate proteoglycans (HSPGs) are concentrated at neuromuscular synapses in many species, including Drosophila. We have established the physiological and patterning functions of HSPGs at the Drosophila neuromuscular junction by using mutations that block heparan sulfate synthesis or sulfation to compromise HSPG function. The mutant animals showed defects in synaptic physiology and morphology suggesting that HSPGs function both presynaptically and postsynaptically; these defects could be rescued by appropriate transgene expression. Of particular interest were selective disruptions of mitochondrial localization, abnormal distributions of Golgi and endoplasmic reticulum markers in the muscle, and a markedly increased level of stimulus-dependent endocytosis in the motoneuron. Our data support the emerging view that HSPG functions are not limited to the cell surface and matrix environments, but also affect a diverse set of cellular processes including membrane trafficking and organelle distributions.
Assuntos
Movimento Celular/fisiologia , Heparitina Sulfato/biossíntese , Mitocôndrias/metabolismo , Junção Neuromuscular/fisiologia , Neurônios/classificação , Neurônios/fisiologia , Animais , Animais Geneticamente Modificados , Comportamento Animal/fisiologia , Comunicação Celular/fisiologia , Movimento Celular/genética , Drosophila , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Endocitose/genética , Potenciais Pós-Sinápticos Excitadores/efeitos dos fármacos , Potenciais Pós-Sinápticos Excitadores/fisiologia , Regulação da Expressão Gênica/fisiologia , Proteínas de Fluorescência Verde/genética , Proteoglicanas de Heparan Sulfato/biossíntese , Proteoglicanas de Heparan Sulfato/genética , Proteoglicanas de Heparan Sulfato/metabolismo , Heparitina Sulfato/genética , Peroxidase do Rábano Silvestre/metabolismo , Larva , Locomoção/fisiologia , Microscopia Eletrônica de Transmissão/métodos , Mitocôndrias/ultraestrutura , Músculo Esquelético/metabolismo , Músculo Esquelético/ultraestrutura , Mutação , Junção Neuromuscular/citologia , Junção Neuromuscular/ultraestruturaRESUMO
Heparan sulfate proteoglycans are abundant molecules in the extracellular matrix and at the cell surface. Heparan sulfate chains are composed of groups of disaccharides whose side chains are modified through a series of enzymatic reactions. Deletion of these enzymes alters heparan sulfate fine structure and leads to changes in cell proliferation and tissue development. The role of heparan sulfate modification has not been explored in the vessel wall. The goal of this study was to test the hypothesis that altering heparan sulfate fine structure would impact vascular smooth muscle cell (VSMC) proliferation, vessel structure, and remodeling in response to injury. A heparan sulfate modifying enzyme, N-deacetylase N-sulfotransferase1 (Ndst1) was deleted in smooth muscle resulting in decreased N- and 2-O sulfation of the heparan sulfate chains. Smooth muscle specific deletion of Ndst1 led to a decrease in proliferating VSMCs and the circumference of the femoral artery in neonatal and adult mice. In response to vascular injury, mice lacking Ndst1 exhibited a significant reduction in lesion formation. Taken together, these data provide new evidence that modification of heparan sulfate fine structure through deletion of Ndst1 is sufficient to decrease VSMC proliferation and alter vascular remodeling.
Assuntos
Vasos Sanguíneos/patologia , Vasos Sanguíneos/fisiopatologia , Músculo Liso Vascular/patologia , Miócitos de Músculo Liso/enzimologia , Miócitos de Músculo Liso/patologia , Sulfotransferases/deficiência , Envelhecimento/patologia , Animais , Animais Recém-Nascidos , Vasos Sanguíneos/enzimologia , Proliferação de Células , Artéria Femoral/enzimologia , Artéria Femoral/patologia , Deleção de Genes , Testes de Função Cardíaca , Heparitina Sulfato/metabolismo , Camundongos , Tamanho do Órgão , Sulfotransferases/metabolismo , Túnica Íntima/enzimologia , Túnica Íntima/patologia , Túnica Íntima/fisiopatologia , Túnica Média/enzimologia , Túnica Média/patologia , Túnica Média/fisiopatologiaRESUMO
Autophagy is a catabolic process that provides cells with energy and molecular building blocks during nutritional stress. Autophagy also removes misfolded proteins and damaged organelles, a critical mechanism for cellular repair. Earlier work demonstrated that heparan sulfate proteoglycans, an abundant class of carbohydrate-modified proteins found on cell surfaces and in the extracellular matrix, suppress basal levels of autophagy in several cell types during development in Drosophila melanogaster In studies reported here, we examined the capacity of heparan sulfate synthesis to influence events affected by autophagy, including lifespan, resistance to reactive oxygen species (ROS) stress, and accumulation of ubiquitin-modified proteins in the brain. Compromising heparan sulfate synthesis increased autophagy-dependent processes, evident by extended lifespan, increased resistance to ROS, and reduced accumulation of ubiquitin-modified proteins in the brains of ROS exposed adults. The capacity of altering heparan sulfate biosynthesis to protect cells from injury was also evaluated in two different models of neurodegeneration, overexpression of Presenilin and parkin mutants. Presenilin overexpression in the retina produces cell loss, and compromising heparan sulfate biosynthesis rescued retinal patterning and size abnormalities in these animals. parkin is the fly homolog of human PARK2, one of the genes responsible for juvenile onset Parkinson's Disease. Parkin is involved in mitochondrial surveillance and compromising parkin function results in degeneration of both flight muscle and dopaminergic neurons in Drosophila Altering heparan sulfate biosynthesis suppressed flight muscle degeneration and mitochondrial dysmorphology, indicating that activation of autophagy-mediated removal of mitochondria (mitophagy) is potentiated in these animals. These findings provide in vivo evidence that altering the levels of heparan sulfate synthesis activates autophagy and can provide protection from a variety of cellular stressors.
Assuntos
Autofagia , Proteínas de Drosophila/genética , Heparitina Sulfato/biossíntese , Longevidade , Estresse Oxidativo , Ubiquitina-Proteína Ligases/genética , Animais , Encéfalo/metabolismo , Olho Composto de Artrópodes/metabolismo , Proteínas de Drosophila/metabolismo , Drosophila melanogaster , Heparitina Sulfato/genética , Músculos/metabolismo , Mutação , Presenilinas/genética , Presenilinas/metabolismo , Ubiquitina-Proteína Ligases/metabolismoRESUMO
Wingless (Wg) is a morphogen required for the patterning of many Drosophila tissues. Several lines of evidence implicate heparan sulfate-modified proteoglycans (HSPGs) such as Dally-like protein (Dlp) in the control of Wg distribution and signaling. We show that dlp is required to limit Wg levels in the matrix, contrary to the expectation from overexpression studies. dlp mutants show ectopic activation of Wg signaling at the presumptive wing margin and a local increase in extracellular Wg levels. dlp somatic cell clones disrupt the gradient of extracellular Wg, producing ectopic activation of high threshold Wg targets but reducing the expression of lower threshold Wg targets where Wg is limiting. Notum encodes a secreted protein that also limits Wg distribution, and genetic interaction studies show that dlp and Notum cooperate to restrict Wg signaling. These findings suggest that modification of an HSPG by a secreted hydrolase can control morphogen levels in the matrix.
Assuntos
Proteínas de Drosophila/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Proteínas de Insetos/metabolismo , Proteoglicanas/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Transdução de Sinais , Animais , Padronização Corporal/genética , Células Clonais , Drosophila/embriologia , Drosophila/crescimento & desenvolvimento , Proteínas de Drosophila/genética , Olho/ultraestrutura , Deleção de Genes , Genes de Insetos , Heterozigoto , Proteínas de Insetos/genética , Morfogênese , Proteoglicanas/genética , Proteínas Proto-Oncogênicas/genética , Distribuição Tecidual , Asas de Animais/embriologia , Asas de Animais/crescimento & desenvolvimento , Asas de Animais/ultraestrutura , Proteína Wnt1RESUMO
Retinal ganglion cell (RGC) axons are topographically ordered in the optic tract according to their retinal origin. In zebrafish dackel (dak) and boxer (box) mutants, some dorsal RGC axons missort in the optic tract but innervate the tectum topographically. Molecular cloning reveals that dak and box encode ext2 and extl3, glycosyltransferases implicated in heparan sulfate (HS) biosynthesis. Both genes are required for HS synthesis, as shown by biochemical and immunohistochemical analysis, and are expressed maternally and then ubiquitously, likely playing permissive roles. Missorting in box can be rescued by overexpression of extl3. dak;box double mutants show synthetic pathfinding phenotypes that phenocopy robo2 mutants, suggesting that Robo2 function requires HS in vivo; however, tract sorting does not require Robo function, since it is normal in robo2 null mutants. This genetic evidence that heparan sulfate proteoglycan function is required for optic tract sorting provides clues to begin understanding the underlying molecular mechanisms.
Assuntos
Axônios/metabolismo , Proteoglicanas de Heparan Sulfato/biossíntese , N-Acetilglucosaminiltransferases/fisiologia , Vias Visuais/metabolismo , Proteínas de Peixe-Zebra/fisiologia , Animais , Proteoglicanas de Heparan Sulfato/genética , Dados de Sequência Molecular , N-Acetilglucosaminiltransferases/biossíntese , N-Acetilglucosaminiltransferases/genética , Células Ganglionares da Retina/metabolismo , Vias Visuais/embriologia , Peixe-ZebraRESUMO
Recently, several evolutionary conserved signaling pathways that play prominent roles in regulating early neurodevelopment have been found to regulate synaptic remodeling in the adult. To test whether adult neuronal expression of bone morphogenic protein (BMP) signaling components also plays a postnatal role in regulating neuronal plasticity, we modulated BMP signaling in mice both in vivo and in vitro by genetic removal of the BMP inhibitor chordin or by perfusing recombinant BMP signaling pathway components onto acute hippocampal slices. Chordin null mice exhibited a significant increase in presynaptic transmitter release from hippocampal neurons, resulting in enhanced paired-pulse facilitation and long-term potentiation. These mice also showed a decreased acquisition time in a water maze test along with less exploratory activity during Y-maze and open-field tests. Perfusion of BMP ligands onto hippocampal slices replicated the presynaptic phenotype of chordin null slices, but bath application of Noggin, another antagonist of BMP signaling pathway, significantly decrease the frequency of miniature EPSCs. These results demonstrate that the BMP signaling pathway contributes to synaptic plasticity and learning likely through a presynaptic mechanism.
Assuntos
Glicoproteínas/metabolismo , Hipocampo/citologia , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Aprendizagem/fisiologia , Plasticidade Neuronal/fisiologia , Terminações Pré-Sinápticas/metabolismo , Comportamento Espacial/fisiologia , Animais , Animais Recém-Nascidos , Proteínas Morfogenéticas Ósseas/metabolismo , Células Cultivadas , Relação Dose-Resposta à Radiação , Estimulação Elétrica/métodos , Embrião de Mamíferos , Potenciais Pós-Sinápticos Excitadores/efeitos dos fármacos , Potenciais Pós-Sinápticos Excitadores/fisiologia , Potenciais Pós-Sinápticos Excitadores/efeitos da radiação , Comportamento Exploratório/fisiologia , Glicoproteínas/deficiência , Hipocampo/fisiologia , Técnicas In Vitro , Peptídeos e Proteínas de Sinalização Intercelular/deficiência , Masculino , Aprendizagem em Labirinto/fisiologia , Camundongos , Camundongos Knockout , Microscopia Eletrônica de Transmissão/métodos , Plasticidade Neuronal/genética , Neurônios/citologia , Neurônios/efeitos dos fármacos , Neurônios/efeitos da radiação , Técnicas de Patch-Clamp , Terminações Pré-Sinápticas/ultraestrutura , Fatores de TempoRESUMO
Heparan sulfate proteoglycans (HSPGs), a class of glycosaminoglycan-modified proteins, control diverse patterning events via their regulation of growth-factor signaling and morphogen distribution. In C. elegans, zebrafish, and the mouse, heparan sulfate (HS) biosynthesis is required for normal axon guidance, and mutations affecting Syndecan (Sdc), a transmembrane HSPG, disrupt axon guidance in Drosophila embryos. Glypicans, a family of glycosylphosphatidylinositol (GPI)-linked HSPGs, are expressed on axons and growth cones in vertebrates, but their role in axon guidance has not been determined. We demonstrate here that the Drosophila glypican Dally-like protein (Dlp) is required for proper axon guidance and visual-system function. Mosaic studies revealed that Dlp is necessary in both the retina and the brain for different aspects of visual-system assembly. Sdc mutants also showed axon guidance and visual-system defects, some that overlap with dlp and others that are unique. dlp+ transgenes were able to rescue some sdc visual-system phenotypes, but sdc+ transgenes were ineffective in rescuing dlp abnormalities. Together, these findings suggest that in some contexts HS chains provide the biologically critical component, whereas in others the structure of the protein core is also essential.
Assuntos
Axônios/metabolismo , Movimento Celular/fisiologia , Proteínas de Drosophila/metabolismo , Drosophila/embriologia , Glicoproteínas de Membrana/metabolismo , Morfogênese , Proteoglicanas/metabolismo , Vias Visuais/fisiologia , Animais , Encéfalo/metabolismo , Drosophila/metabolismo , Eletrofisiologia , Regulação da Expressão Gênica no Desenvolvimento , Heparitina Sulfato/metabolismo , Imuno-Histoquímica , Microscopia Confocal , Microscopia Eletrônica de Varredura , Células Fotorreceptoras de Invertebrados/metabolismo , Células Fotorreceptoras de Invertebrados/ultraestrutura , Retina/metabolismo , Sindecanas , Vias Visuais/metabolismoRESUMO
Heparan sulfate proteoglycans (HSPGs) are ubiquitous molecules that are critical for signaling mediated by many growth factors, including members of the Wnt, transforming growth factor-beta, Hedgehog, and fibroblast growth factor families, and are essential for cell specification, axon guidance, and the establishment of morphogen gradients. Although the heparan sulfate modifications of HSPGs are critical, there is much to learn about how the protein cores contribute to the specific signaling functions of these cell-surface and matrix molecules. Recent work has demonstrated that glypican-1 and syndecan-1 expressed by tumor cells have specific roles in FGF2 signaling, affecting their responses to this mitogenic stimulus.
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Fator 2 de Crescimento de Fibroblastos/fisiologia , Proteoglicanas de Heparan Sulfato/fisiologia , Glicoproteínas de Membrana/fisiologia , Neoplasias/metabolismo , Proteoglicanas/fisiologia , Animais , Proteoglicanas de Heparan Sulfato/metabolismo , Humanos , Glicoproteínas de Membrana/metabolismo , Proteoglicanas/metabolismo , Transdução de Sinais , Sindecana-1 , SindecanasRESUMO
Heparan sulfate-modified proteoglycans (HSPGs) are important regulators of signaling and molecular recognition at the cell surface and in the extracellular space. Disruption of HSPG core proteins, HS-synthesis, or HS-degradation can have profound effects on growth, patterning, and cell survival. The Drosophila neuromuscular junction provides a tractable model for understanding the activities of HSPGs at a synapse that displays developmental and activity-dependent plasticity. Muscle cell-specific knockdown of HS biosynthesis disrupted the organization of a specialized postsynaptic membrane, the subsynaptic reticulum (SSR), and affected the number and morphology of mitochondria. We provide evidence that these changes result from a dysregulation of macroautophagy (hereafter referred to as autophagy). Cellular and molecular markers of autophagy are all consistent with an increase in the levels of autophagy in the absence of normal HS-chain biosynthesis and modification. HS production is also required for normal levels of autophagy in the fat body, the central energy storage and nutritional sensing organ in Drosophila. Genetic mosaic analysis indicates that HS-dependent regulation of autophagy occurs non-cell autonomously, consistent with HSPGs influencing this cellular process via signaling in the extracellular space. These findings demonstrate that HS biosynthesis has important regulatory effects on autophagy and that autophagy is critical for normal assembly of postsynaptic membrane specializations.
Assuntos
Autofagia , Drosophila melanogaster/citologia , Drosophila melanogaster/metabolismo , Proteoglicanas de Heparan Sulfato/metabolismo , Animais , Autofagossomos/metabolismo , Autofagossomos/ultraestrutura , Proteínas Relacionadas à Autofagia/genética , Proteínas Relacionadas à Autofagia/metabolismo , Regulação para Baixo , Drosophila/genética , Drosophila/metabolismo , Drosophila melanogaster/genética , Drosophila melanogaster/ultraestrutura , Corpo Adiposo/metabolismo , Corpo Adiposo/ultraestrutura , Proteoglicanas de Heparan Sulfato/biossíntese , Homozigoto , Larva/metabolismo , Larva/ultraestrutura , Mitocôndrias/metabolismo , Mitocôndrias/ultraestrutura , Músculos/metabolismo , Músculos/ultraestrutura , Mutação/genética , Junção Neuromuscular/metabolismo , Fenótipo , Interferência de RNA , Sinapses/metabolismo , Sinapses/ultraestruturaRESUMO
Autism spectrum disorder is a complex trait with a high degree of heritability as well as documented susceptibility from environmental factors. In this study the contributions of copy number variation, exposure to air pollutants, and the interaction between the two on autism risk, were evaluated in the population-based case-control Childhood Autism Risks from Genetics and Environment (CHARGE) Study. For the current investigation, we included only those CHARGE children (a) who met criteria for autism or typical development and (b) for whom our team had conducted both genetic evaluation of copy number burden and determination of environmental air pollution exposures based on mapping addresses from the pregnancy and early childhood. This sample consisted of 158 cases of children with autism and 147 controls with typical development. Multiple logistic regression models were fit with and without environmental variable-copy number burden interactions. We found no correlation between average air pollution exposure from conception to age 2 years and the child's CNV burden. We found a significant interaction in which a 1SD increase in duplication burden combined with a 1SD increase in ozone exposure was associated with an elevated autism risk (OR 3.4, P < 0.005) much greater than the increased risks associated with either genomic duplication (OR 1.85, 95% CI 1.25-2.73) or ozone (OR 1.20, 95% CI 0.93-1.54) alone. Similar results were obtained when CNV and ozone were dichotomized to compare those in the top quartile relative to those having a smaller CNV burden and lower exposure to ozone, and when exposures were assessed separately for pregnancy, the first year of life, and the second year of life. No interactions were observed for other air pollutants, even those that demonstrated main effects; ozone tends to be negatively correlated with the other pollutants examined. While earlier work has demonstrated interactions between the presence of a pathogenic CNV and an environmental exposure [Webb et al., 2016], these findings appear to be the first indication that global copy number variation may increase susceptibility to certain environmental factors, and underscore the need to consider both genomics and environmental exposures as well as the mechanisms by which each may amplify the risks for autism associated with the other. Autism Res 2017, 10: 1470-1480. © 2017 International Society for Autism Research, Wiley Periodicals, Inc.
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Poluição do Ar/estatística & dados numéricos , Transtorno do Espectro Autista/epidemiologia , Transtorno do Espectro Autista/fisiopatologia , Variações do Número de Cópias de DNA/fisiologia , Exposição Ambiental/estatística & dados numéricos , Estudos de Casos e Controles , Pré-Escolar , Feminino , Humanos , Masculino , Material Particulado , GravidezRESUMO
A clear and predictive understanding of the etiology of autism spectrum disorders (ASD), a group of neurodevelopmental disorders characterized by varying deficits in social interaction and communication as well as repetitive behaviors, has not yet been achieved. There remains active debate about the origins of autism, and the degree to which genetic and environmental factors, and their interplay, produce the range and heterogeneity of cognitive, developmental, and behavioral features seen in children carrying a diagnosis of ASD. Unlocking the causes of these complex developmental disorders will require a collaboration of experts in many disciplines, including clinicians, environmental exposure experts, bioinformaticists, geneticists, and computer scientists. For this workshop we invited prominent researchers in the field of autism, covering a range of topics from genetic and environmental research to ethical considerations. The goal of this workshop: provide an introduction to the current state of autism research, highlighting the potential for multi-disciplinary collaborations that rigorously evaluate the many potential contributors to ASD. It is further anticipated that approaches that successfully advance the understanding of ASD can be applied to the study of other common, complex disorders. Herein we provide a short review of ASD and the work of the invited speakers.
Assuntos
Transtornos Globais do Desenvolvimento Infantil/etiologia , Transtornos Globais do Desenvolvimento Infantil/genética , Biologia Computacional , Coleta de Dados , Meio Ambiente , Genômica , HumanosRESUMO
The identification and validation of gene-gene interactions is a major challenge in human studies. Here, we explore an approach for studying epistasis in humans using a Drosophila melanogaster model of neonatal diabetes mellitus. Expression of the mutant preproinsulin (hINS(C96Y)) in the eye imaginal disc mimics the human disease: it activates conserved stress-response pathways and leads to cell death (reduction in eye area). Dominant-acting variants in wild-derived inbred lines from the Drosophila Genetics Reference Panel produce a continuous, highly heritable distribution of eye-degeneration phenotypes in a hINS(C96Y) background. A genome-wide association study (GWAS) in 154 sequenced lines identified a sharp peak on chromosome 3L, which mapped to a 400-bp linkage block within an intron of the gene sulfateless (sfl). RNAi knockdown of sfl enhanced the eye-degeneration phenotype in a mutant-hINS-dependent manner. RNAi against two additional genes in the heparan sulfate (HS) biosynthetic pathway (ttv and botv), in which sfl acts, also modified the eye phenotype in a hINS(C96Y)-dependent manner, strongly suggesting a novel link between HS-modified proteins and cellular responses to misfolded proteins. Finally, we evaluated allele-specific expression difference between the two major sfl-intronic haplotypes in heterozygtes. The results showed significant heterogeneity in marker-associated gene expression, thereby leaving the causal mutation(s) and its mechanism unidentified. In conclusion, the ability to create a model of human genetic disease, map a QTL by GWAS to a specific gene, and validate its contribution to disease with available genetic resources and the potential to experimentally link the variant to a molecular mechanism demonstrate the many advantages Drosophila holds in determining the genetic underpinnings of human disease.
Assuntos
Diabetes Mellitus/genética , Variação Genética , Proinsulina/genética , Alelos , Animais , Animais Geneticamente Modificados , Diabetes Mellitus/metabolismo , Modelos Animais de Doenças , Proteínas de Drosophila/química , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Drosophila melanogaster , Epistasia Genética , Olho/metabolismo , Olho/patologia , Feminino , Expressão Gênica , Técnicas de Silenciamento de Genes , Estudo de Associação Genômica Ampla , Heparitina Sulfato/biossíntese , Humanos , Íntrons , Masculino , Mutação , Fenótipo , Proinsulina/química , Dobramento de Proteína , Interferência de RNA , Sulfotransferases/química , Sulfotransferases/genética , Sulfotransferases/metabolismoRESUMO
In omic research, such as genome wide association studies, researchers seek to repeat their results in other datasets to reduce false positive findings and thus provide evidence for the existence of true associations. Unfortunately this standard validation approach cannot completely eliminate false positive conclusions, and it can also mask many true associations that might otherwise advance our understanding of pathology. These issues beg the question: How can we increase the amount of knowledge gained from high throughput genetic data? To address this challenge, we present an approach that complements standard statistical validation methods by drawing attention to both potential false negative and false positive conclusions, as well as providing broad information for directing future research. The Diverse Convergent Evidence approach (DiCE) we propose integrates information from multiple sources (omics, informatics, and laboratory experiments) to estimate the strength of the available corroborating evidence supporting a given association. This process is designed to yield an evidence metric that has utility when etiologic heterogeneity, variable risk factor frequencies, and a variety of observational data imperfections might lead to false conclusions. We provide proof of principle examples in which DiCE identified strong evidence for associations that have established biological importance, when standard validation methods alone did not provide support. If used as an adjunct to standard validation methods this approach can leverage multiple distinct data types to improve genetic risk factor discovery/validation, promote effective science communication, and guide future research directions.