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C10ORF10/DEPP, a transcriptional target of FOXO3 regulates ROS-sensitivity by destabilizing peroxisomes in human neuroblastoma

ABSTRACT: FOXO transcription factors control cellular formation of reactive oxygen species (ROS), which critically contribute to cell survival and cell death in neuroblastoma. Here, we report that C10orf10, also named “Decidual Protein induced by Progesterone (DEPP)”, is a direct transcriptional target of FOXO3 in human neuroblastoma. As FOXO3-mediated apoptosis involves a biphasic ROS accumulation, we analyzed cellular ROS levels in DEPP-knockdown cells by live-cell imaging. Knockdown of DEPP prevented the primary and secondary ROS accumulation during FOXO3 activation and attenuates FOXO3-induced apoptosis, whereas its overexpression raises cellular ROS levels and sensitizes to cell death. In neuronal cells, cellular steady state ROS are mainly detoxified in peroxisomes by the enzyme CAT/catalase. As DEPP contains a peroxisomal-targeting-signal-type-2 (PTS2) sequence at its N-terminus that enables protein import into peroxisomes, we analyzed the effect of DEPP on peroxisomal function by measuring the catalase enzyme activity. Catalase activity was reduced by conditional DEPP overexpression and significantly increased in DEPP-knockdown cells. Using live cell imaging and fluorescent peroxisomal and mitochondrial probes we demonstrate that DEPP localizes to peroxisomes and mitochondria in neuroblastoma cells. The combined data indicate that DEPP reduces peroxisomal activity and thereby impairs the cellular ROS detoxification capacity and contributes to death sensitization. SH-EP, NB15 neuroblastoma cells and CCRF-CEM-C7H2 acute lymphoblastic leukemia cells were infected with the retrovirus plasmid pLIB-FOXO3(A3)-Ertm-iresNeo.

ORGANISM(S): Homo sapiens

PROVIDER: GSE53046 | GEO | 2014/10/06

SECONDARY ACCESSION(S): PRJNA230747

REPOSITORIES: GEO

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Project description:FOXO transcription factors control cellular formation of reactive oxygen species (ROS), which critically contribute to cell survival and cell death in neuroblastoma. Here, we report that C10orf10, also named M-bM-^@M-^\Decidual Protein induced by Progesterone (DEPP)M-bM-^@M-^], is a direct transcriptional target of FOXO3 in human neuroblastoma. As FOXO3-mediated apoptosis involves a biphasic ROS accumulation, we analyzed cellular ROS levels in DEPP-knockdown cells by live-cell imaging. Knockdown of DEPP prevented the primary and secondary ROS accumulation during FOXO3 activation and attenuates FOXO3-induced apoptosis, whereas its overexpression raises cellular ROS levels and sensitizes to cell death. In neuronal cells, cellular steady state ROS are mainly detoxified in peroxisomes by the enzyme CAT/catalase. As DEPP contains a peroxisomal-targeting-signal-type-2 (PTS2) sequence at its N-terminus that enables protein import into peroxisomes, we analyzed the effect of DEPP on peroxisomal function by measuring the catalase enzyme activity. Catalase activity was reduced by conditional DEPP overexpression and significantly increased in DEPP-knockdown cells. Using live cell imaging and fluorescent peroxisomal and mitochondrial probes we demonstrate that DEPP localizes to peroxisomes and mitochondria in neuroblastoma cells. The combined data indicate that DEPP reduces peroxisomal activity and thereby impairs the cellular ROS detoxification capacity and contributes to death sensitization. SH-EP, NB15 neuroblastoma cells and CCRF-CEM-C7H2 acute lymphoblastic leukemia cells were infected with the retrovirus plasmid pLIB-FOXO3(A3)-Ertm-iresNeo. Gene expression measures of samples with activated FOXO3 transcription factor (3h OHT treated) have been compared to untreated samples (0h time point). To rule out gene regulations by estrogen samples treated for 3 hours with tamoxifen have been compared to the untreated samples. Only genes that were more than two-fold regulated in the first, but not in the second comparison were defined to be FOXO3 regulated.

Project description:Fruit ripening is a very important physiological process which gains relevance in crop species due to their economical and nutritional repercussions. During fruit ripening enormous metabolic changes occur in a genetically-controlled scenario affecting the physiology of most cell compartments. Peroxisomes are single-membrane bounded organelles present in all eukaryotes which display a noteworthy nitro-oxidative metabolism, and harbor catalase as one of the major antioxidant enzymes in cells. Quantitative proteomics analysis by isobaric tags for relative and absolute quantification (iTRAQ) was used to investigate the ripening process in sweet pepper (Capsicum annuum L.) fruits. Out of 2,574 quantified proteins, 692 were found to be significantly more abundant in immature green fruits compared to red ones, but 497 showed a lower expression as the ripening proceeded. Overall, about 50% of the detectable proteins were estimated to modify their expression due to the ripening process. Data obtained from the Gene Ontology algorithms (AgriGO platform) showed that from all the identified proteins, 46 (2%) were only predicted to have a peroxisomal origin, with a high number of them displaying up-expression tendency during ripening. Catalase was framed within the group which showed lower expression in ripe fruits, but this enzyme was also susceptible to undergo posttranslational modifications (PTMs) promoted by reactive nitrogen and oxygen species (RNS and ROS) through nitration, S-nitrosylation and oxidation events which provoked its inhibition. The biochemical characterization of catalase indicates that it has atypical native molecular mass (125-135 kDa), since it behaves as a homodimer, and isoelectric point of 7.4, which is higher than that of the majority of plant catalases reported so far. Taking together, these data suggest that ROS and RNS could be essential to modulate the role of catalase for the maintenance of basic cellular peroxisomal functions during pepper fruit ripening where nitro-oxidative stress occur.

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C10ORF10/DEPP, a transcriptional target of FOXO3 regulates ROS-sensitivity by destabilizing peroxisomes in human neuroblastoma

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