RESUMEN
Mitophagy is thought to play a key role in eliminating damaged mitochondria, with diseases such as cancer and neurodegeneration exhibiting defects in this process. Mitophagy is also involved in cell differentiation and maturation, potentially through modulating mitochondrial metabolic reprogramming. Here we examined mitophagy that is induced upon iron chelation and found that the transcriptional activity of HIF1α, in part through upregulation of BNIP3 and NIX, is an essential mediator of this pathway in SH-SY5Y cells. In contrast, HIF1α is dispensable for mitophagy occurring upon mitochondrial depolarisation. To examine the role of this pathway in a metabolic reprogramming and differentiation context, we utilised the H9c2 cell line model of cardiomyocyte maturation. During differentiation of these cardiomyoblasts, mitophagy increased and required HIF1α-dependent upregulation of NIX. Though HIF1α was essential for expression of key cardiomyocyte markers, mitophagy was not directly required. However, enhancing mitophagy through NIX overexpression, accelerated marker gene expression. Taken together, our findings provide a molecular link between mitophagy signalling and cardiomyocyte differentiation and suggest that although mitophagy may not be essential per se, it plays a critical role in maintaining mitochondrial integrity during this energy demanding process.
RESUMEN
Mutations in the gene encoding the microtubule severing ATPase spastin are the most frequent cause of hereditary spastic paraplegia, a genetic condition characterised by length-dependent axonal degeneration. Here, we show that HeLa cells lacking spastin and embryonic fibroblasts from a spastin knock-in mouse model become highly polarised and develop cellular protrusions. In HeLa cells, this phenotype was rescued by wild-type spastin, but not by forms unable to sever microtubules or interact with endosomal ESCRT-III proteins. Cells lacking the spastin-interacting ESCRT-III-associated proteins IST1 or CHMP1B also developed protrusions. The protrusion phenotype required protrudin, a RAB-interacting protein that interacts with spastin and localises to ER-endosome contact sites, where it promotes KIF5-dependent endosomal motility to protrusions. Consistent with this, the protrusion phenotype in cells lacking spastin also required KIF5. Lack or mutation of spastin resulted in functional consequences for receptor traffic of a pathway implicated in HSP, as Bone Morphogenetic Protein receptor distribution became polarised. Our results, therefore, identify a novel role for ESCRT-III proteins and spastin in regulating polarised membrane traffic.
Asunto(s)
Extensiones de la Superficie Celular/metabolismo , Complejos de Clasificación Endosomal Requeridos para el Transporte/metabolismo , Espastina/metabolismo , Animales , Receptores de Proteínas Morfogenéticas Óseas/metabolismo , Membrana Celular/metabolismo , Polaridad Celular , Extensiones de la Superficie Celular/ultraestructura , Células Cultivadas , Fibroblastos/citología , Fibroblastos/metabolismo , Técnicas de Sustitución del Gen , Células HeLa , Humanos , Cinesinas/fisiología , Ratones , Transporte de Proteínas , Paraplejía Espástica Hereditaria/genética , Espastina/genética , Proteínas de Transporte Vesicular/fisiologíaRESUMEN
Many genetic neurological disorders exhibit variable expression within affected families, often exemplified by variations in disease age at onset. Epistatic effects (i.e. effects of modifier genes on the disease gene) may underlie this variation, but the mechanistic basis for such epistatic interactions is rarely understood. Here we report a novel epistatic interaction between SPAST and the contiguous gene DPY30, which modifies age at onset in hereditary spastic paraplegia, a genetic axonopathy. We found that patients with hereditary spastic paraplegia caused by genomic deletions of SPAST that extended into DPY30 had a significantly younger age at onset. We show that, like spastin, the protein encoded by SPAST, the DPY30 protein controls endosomal tubule fission, traffic of mannose 6-phosphate receptors from endosomes to the Golgi, and lysosomal ultrastructural morphology. We propose that additive effects on this pathway explain the reduced age at onset of hereditary spastic paraplegia in patients who are haploinsufficient for both genes.
Asunto(s)
Epistasis Genética/genética , Mutación/genética , Proteínas Nucleares/genética , Paraplejía Espástica Hereditaria/genética , Espastina/genética , Adulto , Edad de Inicio , Antígenos CD8/genética , Antígenos CD8/metabolismo , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Femenino , Factores de Intercambio de Guanina Nucleótido/genética , Factores de Intercambio de Guanina Nucleótido/metabolismo , Células HeLa/metabolismo , Células HeLa/ultraestructura , Humanos , Proteína 1 de la Membrana Asociada a los Lisosomas/metabolismo , Proteína 1 de la Membrana Asociada a los Lisosomas/ultraestructura , Lisosomas/metabolismo , Lisosomas/ultraestructura , Masculino , Proteínas de la Membrana/genética , Proteínas de la Membrana/metabolismo , Persona de Mediana Edad , Proteínas Nucleares/metabolismo , Proteínas Nucleares/ultraestructura , Transporte de Proteínas/genética , Factores de Transcripción/genética , Factores de Transcripción/metabolismoRESUMEN
The autophagic turnover of mitochondria, termed mitophagy, is thought to play an essential role in not only maintaining the health of the mitochondrial network but also that of the cell and organism as a whole. We have come a long way in identifying the molecular components required for mitophagy through extensive in vitro work and cell line characterisation, yet the physiological significance and context of these pathways remain largely unexplored. This is highlighted by the recent development of new mouse models that have revealed a striking level of variation in mitophagy, even under normal conditions. Here, we focus on programmed mitophagy and summarise our current understanding of why, how and where this takes place in mammals.