RESUMO
Disruption of sphingolipid homeostasis is known to cause neurological disorders, but the mechanisms by which specific sphingolipid species modulate pathogenesis remain unclear. The last step of de novo sphingolipid synthesis is the conversion of dihydroceramide to ceramide by dihydroceramide desaturase (human DEGS1; Drosophila Ifc). Loss of ifc leads to dihydroceramide accumulation, oxidative stress, and photoreceptor degeneration, whereas human DEGS1 variants are associated with leukodystrophy and neuropathy. In this work, we demonstrate that DEGS1/ifc regulates Rac1 compartmentalization in neuronal cells and that dihydroceramide alters the association of active Rac1 with organelle-mimicking membranes. We further identify the Rac1-NADPH oxidase (NOX) complex as the major cause of reactive oxygen species (ROS) accumulation in ifc-knockout (ifc-KO) photoreceptors and in SH-SY5Y cells with the leukodystrophy-associated DEGS1H132R variant. Suppression of Rac1-NOX activity rescues degeneration of ifc-KO photoreceptors and ameliorates oxidative stress in DEGS1H132R-carrying cells. Therefore, we conclude that DEGS1/ifc deficiency causes dihydroceramide accumulation, resulting in Rac1 mislocalization and NOX-dependent neurodegeneration.
Assuntos
Proteínas de Drosophila/genética , Drosophila melanogaster/genética , Ácidos Graxos Dessaturases/genética , Proteínas de Membrana/genética , NADPH Oxidases/genética , Proteínas rac1 de Ligação ao GTP/genética , Animais , Linhagem Celular Tumoral , Ceramidas/metabolismo , Proteínas de Drosophila/deficiência , Drosophila melanogaster/metabolismo , Eletrorretinografia , Ácidos Graxos Dessaturases/antagonistas & inibidores , Ácidos Graxos Dessaturases/metabolismo , Regulação da Expressão Gênica , Humanos , Proteínas de Membrana/deficiência , NADPH Oxidases/metabolismo , Neurônios/metabolismo , Neurônios/patologia , Estresse Oxidativo , Células Fotorreceptoras de Invertebrados/metabolismo , Células Fotorreceptoras de Invertebrados/patologia , Mutação Puntual , Ligação Proteica , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Retina/metabolismo , Retina/patologia , Transdução de Sinais , Proteínas rac1 de Ligação ao GTP/metabolismoRESUMO
Impaired clearance of amyloid-ß peptide (Aß) leads to abnormal extracellular accumulation of this neurotoxic protein that drives neurodegeneration in sporadic Alzheimer's disease (AD). Connective tissue growth factor (CTGF/CCN2) expression is elevated in plaque-surrounding astrocytes in AD patients. However, the role of CTGF in AD pathogenesis remains unclear. Here we characterized the neuroprotective activity of CTGF. We found that CTGF facilitated Aß uptake and subsequent degradation within primary glia and neuroblastoma cells. CTGF enhanced extracellular Aß degradation via membrane-bound matrix metalloproteinase-14 (MMP14) in glia and extracellular MMP13 in neurons. In the brain of a Drosophila AD model, glial-expression of CTGF reduced Aß deposits, improved locomotor function, and rescued memory deficits. Neuroprotective potential of CTGF against Aß42-induced photoreceptor degeneration was disrupted through silencing MMPs. Therefore, CTGF may represent a node for potential AD therapeutics as it intervenes in glia-neuron communication via specific MMPs to alleviate Aß neurotoxicity in the central nervous system.
Assuntos
Peptídeos beta-Amiloides/toxicidade , Fator de Crescimento do Tecido Conjuntivo/metabolismo , Doença de Alzheimer/metabolismo , Peptídeos beta-Amiloides/metabolismo , Animais , Animais Geneticamente Modificados , Astrócitos/metabolismo , Encéfalo/metabolismo , Fator de Crescimento do Tecido Conjuntivo/fisiologia , Modelos Animais de Doenças , Drosophila , Humanos , Metaloproteinase 14 da Matriz/metabolismo , Camundongos , Neuroglia/metabolismo , Neuroglia/fisiologia , Neurônios/metabolismo , Neurônios/fisiologia , Fármacos Neuroprotetores/metabolismo , Fármacos Neuroprotetores/farmacologia , Síndromes Neurotóxicas/metabolismo , Fragmentos de Peptídeos/metabolismo , Placa Amiloide/metabolismo , RatosRESUMO
Dihydroceramide desaturases are evolutionarily conserved enzymes that convert dihydroceramide (dhCer) to ceramide (Cer). While elevated Cer levels cause neurodegenerative diseases, the neuronal activity of its direct precursor, dhCer, remains unclear. We show that knockout of the fly dhCer desaturase gene, infertile crescent (ifc), results in larval lethality with increased dhCer and decreased Cer levels. Light stimulation leads to ROS increase and apoptotic cell death in ifc-KO photoreceptors, resulting in activity-dependent neurodegeneration. Lipid-containing Atg8/LC3-positive puncta accumulate in ifc-KO photoreceptors, suggesting lipophagy activation. Further enhancing lipophagy reduces lipid droplet accumulation and rescues ifc-KO defects, indicating that lipophagy plays a protective role. Reducing dhCer synthesis prevents photoreceptor degeneration and rescues ifc-KO lethality, while supplementing downstream sphingolipids does not. These results pinpoint that dhCer accumulation is responsible for ifc-KO defects. Human dhCer desaturase rescues ifc-KO larval lethality, and rapamycin reverses defects caused by dhCer accumulation in human neuroblastoma cells, suggesting evolutionarily conserved functions. This study demonstrates a novel requirement for dhCer desaturase in neuronal maintenance in vivo and shows that lipophagy activation prevents activity-dependent degeneration caused by dhCer accumulation.