Metabolic effects of skn-1 down-regulation in C.elegans

skn-1;phase II detoxification;C.elegans;metabolomics;615

약학대학 약학과 ;

University:서울대학교 대학원

关键词: skn-1;    phase II detoxification;    C.elegans;    metabolomics;    615;   

Others  :  http://s-space.snu.ac.kr/bitstream/10371/133562/1/000000025279.pdf

美国|英语

来源: Seoul National University Open Repository

【 摘 要 】

SKN-1/Nrf2, a transcription factor with a well-established role in regulating oxidative stress resistance and detoxification pathway, has also been proved to affect the lifespan of C.elegans. While many studies have characterized the SKN-1 function at genomic level, there has been lack of a systematic investigation at individual metabolites. As SKN-1 function relates to metabolism, we studied metabolite profiles of skn-1 knockdown C.elegans strain by NMR and LC-MS/MS metabolomics approach. First, we fed C.elegans with E.coli HT115 containing the double-strand RNA of skn-1 to knock down skn-1 gene in the worms. The effects of skn-1 knockdown were confirmed by the shorter lifespan and the lower level of skn-1 expression measured by qPCR in knockdown worms. We then extracted metabolites from the worms and obtained metabolite profiles through NMR and LC-MS/MS. Multivariate analysis showed a distinctive metabolic profile with the significant decrease in the oxidative stress defense system represented by the reduction in NADPH/NADP+ ratio, the decline in the transsulfuration pathway-main source of GSH synthesis, and the lower level of total GSH (GSHt) in skn-1 RNAi worms. We also tested the performance of phase II detoxification system since we previously observed the involvement of the skn-1 dependent phase II detoxification in the dietary restriction’s beneficial effects. It is showed that there is an impairment of this system in the knockdown worms as exemplified by the lower level of glutathione conjugate of paracetamol, a compound detoxified through this pathway. By interrogating existing microarray data, we also observed the consistency in the changes of metabolite data with the gene expression levels, including the decline of cbl-1 and gpx encoding gamma-cystathionase and glutathione peroxidase which are necessary for the synthesis and function of GSH respectively, the reduction in the T25B9.9 encoding 6-gluconate phosphate dehydrogenase, which is the main enzyme of NADPH production, and the decrease of ugt and gst encoding UDP-glucuronosyl transferase and Glutathione-S-transferase in the Phase II detoxification pathway. As the correlation between aging and oxidative damage has been hypothesized previously, our results demonstrate the debilitation of the cytoprotective pathway including oxidative stress defense system and xenobiotic detoxification system, underlies the lifespan reduction of skn-1 knockdown worms.