RESUMEN
BACKGROUND: CMTM7 is a tumor suppressor that positively regulates EGFR degradation by promoting Rab5 activation, and plays a vital role in tumor progression. Rab5 forms complexes with Beclin1 and VPS34, and acts in the early stage of autophagy. However, the affects of CMTM7 on autophagy and its mechanism are still unclear. METHODS: The effect of CMTM7 on autophagy induction was confirmed by western blotting, confocal microscopy and transmission electron microscopy. Co-immunoprecipitation was used to analyse the interaction of CMTM7 with autophagy initiation complex and Rab5. The xenograft model in nude mice was used to elucidate the function of CMTM7 in tumorigenicity and autophagy in vivo. RESULTS: In this study, we first demonstrated that CMTM7 facilitated the initiation of autophagosome formation, which consequently promoted the subsequent multistage process of autophagic flux, i.e. from autophagosome assembly till autolysosome formation and degradation. Confocal and co-immunoprecipitation showed that CMTM7 interacted with Rab5, VPS34, Beclin1, and ATG14L, but not with ULK1, UVRAG and LC3B. CMTM7 also increased the activity of ATG14L-linked VPS34 complex and its association with Rab5. Both in vitro and in vivo experiments demonstrated that knockdown of CMTM7 enhanced tumor growth by impairing autophagy. CONCLUSION: These findings highlighted the role of CMTM7 in the regulation of autophagy and tumorigenicity, revealing it as a novel molecule that is associated with the interaction of Rab5 and ATG14L-Beclin1-VPS34 complex. Video Abstract.
Asunto(s)
Proteínas Adaptadoras del Transporte Vesicular/genética , Proteínas Relacionadas con la Autofagia/genética , Beclina-1/genética , Quimiocinas/genética , Fosfatidilinositol 3-Quinasas Clase III/genética , Proteínas con Dominio MARVEL/genética , Neoplasias/genética , Proteínas de Unión al GTP rab5/genética , Animales , Autofagia/genética , Carcinogénesis/genética , Línea Celular Tumoral , Regulación Neoplásica de la Expresión Génica/genética , Xenoinjertos , Humanos , Ratones , Microscopía Electrónica de Transmisión , Complejos Multiproteicos/genética , Complejos Multiproteicos/ultraestructura , Neoplasias/patología , Proteínas de Unión al GTP rab5/ultraestructuraRESUMEN
Sporadic motor neuron disease (MND) is characterized by progressive degeneration of motor neurons and intraneuronal cytoplasmic translocation and deposition of the nuclear protein TDP-43. There is a paucity of data on the subcellular mechanisms of the nuclear-cytoplasmic trafficking of TDP-43, particularly about the precise role of the endosomal-lysosomal system (ELS). In the present study, using a neuron-specific morphometric approach, we examined the expression of the early endosomal marker Rab5 and lysosomal cathepsins B, D, F, and L as well as PAS-stained structures in the anterior horn cells in 11 individuals affected by sporadic MND and 5 age-matched controls. This was compared with the expression of ubiquitin, p62 and TDP-43 and its phosphorylated form. The principal finding was the increased expression of the endosomal marker Rab5 and lysosomal cathepsin D, and of PAS-positive structures in motor neurons of MND cases. Furthermore, the area-portion of Rab5 immunoreactivity correlated well with the intracellular accumulation of ubiquitin, p62 and (phosphorylated) TDP-43. However, double immunolabelling and immunogold electron microscopy excluded colocalization of phosphorylated TDP-43 with the ELS. These data contrast with observations on neuronal cytopathology in Alzheimer's or prion diseases where the disease-specific proteins are processed within endosomes, and suggest a distinct role of the ELS in MND.