RESUMO
The HSP60/HSP10 chaperonin assists folding of proteins in the mitochondrial matrix space by enclosing them in its central cavity. The chaperonin forms part of the mitochondrial protein quality control system. It is essential for cellular survival and mutations in its subunits are associated with rare neurological disorders. Here we present the first survey of interactors of the human mitochondrial HSP60/HSP10 chaperonin. Using a protocol involving metabolic labeling of HEK293 cells, cross-linking, and immunoprecipitation of HSP60, we identified 323 interacting proteins. As expected, the vast majority of these proteins are localized to the mitochondrial matrix space. We find that approximately half of the proteins annotated as mitochondrial matrix proteins interact with the HSP60/HSP10 chaperonin. They cover a broad spectrum of functions and metabolic pathways including the mitochondrial protein synthesis apparatus, the respiratory chain, and mitochondrial protein quality control. Many of the genes encoding HSP60 interactors are annotated as disease genes. There is a correlation between relative cellular abundance and relative abundance in the HSP60 immunoprecipitates. Nineteen abundant matrix proteins occupy more than 60% of the HSP60/HSP10 chaperonin capacity. The reported inventory of interactors can form the basis for interrogating which proteins are especially dependent on the chaperonin.
Assuntos
Chaperonina 10/metabolismo , Chaperonina 60/metabolismo , Proteínas Mitocondriais/metabolismo , Células HEK293 , Humanos , Mitocôndrias/metabolismoRESUMO
Histopathological studies and animal models have suggested an inflammatory component in the pathomechanism of the CHMP2B associated frontotemporal dementia (FTD-3). In this cross-sectional study, serum and cerebrospinal fluid were analyzed for inflammatory markers in CHMP2B mutation carriers. Serum levels of CCL4 were increased throughout life. Serum levels of IL-15, CXCL10, CCL22 and TNF-α were significantly associated with cognitive decline, suggesting a peripheral inflammatory response to neurodegeneration. CSF levels of sTREM2 appeared to increase more rapidly with age in CHMP2B mutation carriers. The identification of a peripheral inflammatory response to disease progression supports the involvement of an inflammatory component in FTD-3.
Assuntos
Complexos Endossomais de Distribuição Requeridos para Transporte , Demência Frontotemporal/sangue , Demência Frontotemporal/líquido cefalorraquidiano , Mediadores da Inflamação/sangue , Mediadores da Inflamação/líquido cefalorraquidiano , Adulto , Idoso , Biomarcadores/sangue , Biomarcadores/líquido cefalorraquidiano , Estudos Transversais , Complexos Endossomais de Distribuição Requeridos para Transporte/genética , Feminino , Demência Frontotemporal/genética , Humanos , Masculino , Pessoa de Meia-Idade , Mutação/genéticaRESUMO
Spinocerebellar ataxia type 2 (SCA2) is a rare neurodegenerative disorder caused by a CAG repeat expansion in the ataxin-2 gene. We show increased oxidative stress, abnormalities in the antioxidant system, changes in complexes involved in oxidative phosphorylation and changes in mitochondrial morphology in SCA2 patient fibroblasts compared to controls, and we show that treatment with CoQ10 can partially reverse these changes. Together, our results suggest that oxidative stress and mitochondrial dysfunction may be contributory factors to the pathophysiology of SCA2 and that therapeutic strategies involving manipulation of the antioxidant system could prove to be of clinical benefit.
Assuntos
Fibroblastos/patologia , Mitocôndrias/patologia , Estresse Oxidativo , Ataxias Espinocerebelares/patologia , Ubiquinona/análogos & derivados , Vitaminas/metabolismo , Adolescente , Adulto , Idoso , Células Cultivadas , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Ubiquinona/metabolismo , Adulto JovemRESUMO
Chaperonins, a subgroup of molecular chaperones, form ring-shaped structures and assist folding of proteins by enclosing them in their inner cavity. The mitochondrial Hsp60/Hsp10 chaperonin system is essential for cell viability and only a very small number of mutations causing human disease have so far been found that appear to selectively affect neuronal tissues. We here review the knowledge on the mammalian Hsp60/Hsp10 system and discuss evidence and observations, which may explain why this is the case. The Hsp60 mutations shown to be associated with neurodegenerative diseases mildly affect the protein and leave residual function. We present arguments for the notion that the neuron/glia specificity may be due to an effect of Hsp60 deficiency on myelination, a neuron-specific property. The substrates of the Hsp60/Hsp10 system are only poorly defined, but the combination of deficiency of a number of mitochondrial enzymes and proteins that are highly dependent on this system for folding is the likely trigger for deficient myelination. However, a number of experimental observations indicate that Hsp60 may also have roles outside mitochondria and deficiency of Hsp60 due to mutation may also affect myelination via these signaling pathways. Taken together, it appears that mild Hsp60 deficiency primarily affects neuronal and/or glia cells whereas more severe deficiency of Hsp60 would affect all tissues and not be compatible with life. We discuss in the end what approaches may lead to a further understanding of the functions of the Hsp60/Hsp10 system in mammalian cells and thus its role in disease conditions.
Assuntos
Chaperonina 60/genética , Chaperonina 60/metabolismo , Proteínas Mitocondriais/genética , Proteínas Mitocondriais/metabolismo , Chaperonas Moleculares/metabolismo , Doenças Neurodegenerativas/metabolismo , Animais , Chaperonina 10/metabolismo , Regulação da Expressão Gênica , Humanos , Mamíferos , Camundongos , Camundongos Knockout , Mitocôndrias/metabolismo , Chaperonas Moleculares/genética , Mutação , Doenças Neurodegenerativas/genética , Doenças Neurodegenerativas/fisiopatologia , Neuroglia/metabolismo , Neuroglia/patologia , Dobramento de Proteína , Paraplegia Espástica Hereditária/genética , Paraplegia Espástica Hereditária/metabolismoRESUMO
Mitochondrial dysfunction is associated with neurodegenerative diseases and mutations in the HSPD1 gene, encoding the mitochondrial Hsp60 chaperone, are the causative factors of two neurodegenerative diseases, hereditary spastic paraplegia and MitChap60 disease. In cooperation with Hsp10, Hsp60 forms a barrel-shaped complex, which encloses unfolded polypeptides and provides an environment facilitating folding. We have generated an Hsp60 variant with a mutation (Asp423Ala) in the ATPase domain and established a stable human embryonic kidney (HEK293) cell line allowing tetracycline-controlled expression of this mutant variant. We monitored expression of the Hsp60-Asp423Ala variant protein following induction and examined its effects on cellular properties. We showed that the folding of mitochondrial-targeted green fluorescent protein, a well-known substrate protein of Hsp60, was consistently impaired in cells expressing Hsp60-Asp423Ala. The level of the Hsp60-Asp423Ala variant protein increased over time upon induction, cell proliferation stopped after 48-h induction and mitochondrial membrane potential decreased in a time-dependent manner. In summary, we have established a stable cell line with controllable expression of an Hsp60 variant, which allows detailed studies of different degrees of Hsp60 deficiency.