Dataset Information

RAS-dependent ROS signaling in C. elegans.

ABSTRACT: Mitochondrial superoxide produced by low concentrations of the pro-oxidant paraquat increases C. elegans lifespan. We found that paraquat acts by intensifying RAS-dependent ROS signaling (RDRS) to alter the expression of >50% of the genome and control animal composition and physiology. We show that RDRS is regulated by negative feedback from SOD-1-dependent conversion of superoxide into cytoplasmic hydrogen peroxide, which in turn acts on a redox-sensitive cysteine (C118) of the RAS homologue LET-60ras. Preventing C118 oxidation by replacement with serine, or mimicking oxidation by replacement with aspartic acid, leads to opposite changes in the expression of thousands of genes. We show that paraquat acts through RDRS to exacerbate and accelerate gene expression changes that are normally observed at the end of post-embryonic development. The identities of the genes affected by paraquat suggest that RDRS stimulation extends lifespan by increasing turnover and repair while moderating damage from metabolic activity.

ORGANISM(S): Caenorhabditis elegans

PROVIDER: GSE200912 | GEO | 2022/12/07

REPOSITORIES: GEO

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Project description:Exposure to Paraquat and RNA interference knockdown of mitochondrial superoxide dismutase (Sod2) are known to result in significant lifespan reduction, locomotor dysfunction, and mitochondrial degeneration in Drosophila melanogaster. Both perturbations increase the flux of superoxide, a progenitor reactive oxygen species, but the molecular underpinnings of the resulting phenotypes are poorly understood. Improved understanding of such processes could lead to advances in the treatment of numerous age-related disorders. Superoxide toxicity can act through protein carbonylation. Analysis of carbonylated proteins is attractive since reactive carbonyl groups are not present in the twenty canonical amino acids and are amenable to labeling and enrichment strategies. Here, carbonylated proteins were labeled with biotin hydrazide and enriched on streptavidin-coated beads. On-bead digestion was used to release carbonylated protein peptides, with relative abundance ratios versus controls obtained using the iTRAQ mass spectrometry-based proteomics approach. While Paraquat exposure and Sod2 knockdown have similar phenotypes, differences in protein carbonylation were anticipated because Paraquat exposure was expected to increase the concentration of superoxide throughout the cell while Sod2 knockdown was only expected to raise the concentration of superoxide in the mitochondrial matrix. Paraquat exposure resulted in widespread increases in carbonylated protein relative abundance: the median Paraquat-exposed to control carbonylated protein relative abundance ratio was 1.53. For Sod2 knockdown, in contrast, the median carbonylated protein relative abundance ratios were 1.13 versus the RNA interference driver control and 1.05 versus the RNA interference transgene control. However, some proteins did show large increases in carbonylated protein relative abundance on Sod2 knockdown, most notably cytochrome c oxidase subunit Vb, possibly providing some indication of the molecular basis of the Sod2-knockdown phenotype.

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