Toward the Understanding of the Metabolic Regulation of Sirtuins

The ability to sense and respond to fluctuations in environmental nutrient levels is a requisite for life. Therefore, multiple nutrient sensing mechanisms are developed though evolution. Silent information regulator 2 (Sir2) proteins, or sirtuins, are highly conserved nicotinamide adenine dinucleotide (NAD+) dependent protein deacetylases. Their dependence on NAD+ links their activity to cellular metabolic status to serve as critical intracellular nutrient sensors. There are seven sirtuins in mammals and these proteins may mediate some of the benefits of caloric restriction by modulating energy metabolism, genomic stability and stress resistance. However, the molecular mechanisms by which sirtuins exert their physiological or pathological influences are not fully understood. The aim of this dissertation work was to gain molecular insights into this knowledge by studying the two aspects of biology: chronic inflammation-induced insulin resistance, and mitochondrial unfolded protein response (mtUPR)./nChronic low-grade inflammation has been shown to contribute to the pathophysiology of many diseases, such as atherosclerosis, diabetes, Alzheimer’s disease, Parkinson’s disease, and cancer. In addition, SIRT2, a cytosol localized sirtuin, has been reported to suppress inflammation in multiple inflammation-inducing mouse models. Based on these findings, we investigated whether SIRT2 suppresses chronic inflammation through regulating inflammasome activity. We found that SIRT2 specifically inhibits the NOD-like receptor family, pyrin domain containing 3 (NLRP3) inflammasome in macrophages by directly deacetylating NLRP3. We confirmed that NLRP3 acetylation enhances its inflammasome function by mutating the deacetylated lysine residues to mimic the constitutively deacetylated state. Under conditions of aging and overnutrition, two prominent risk factors associated with insulin resistance, SIRT2 ablation in mice leads to increased systematic chronic inflammation and insulin resistance. These results establish a novel regulation of the NLRP3 inflammasome by SIRT2, and identify a nodal control point at the interface of nutrient sensing and innate immunity./nPerturbation of mitochondrial protein homeostasis (proteostasis), also known as mitochondrial protein folding stress (mtPFS), activates the mtUPR, a retrograde signaling pathway leading to transcriptional upregulation of mitochondrial chaperones, repression of translation, and stress relief. The mtUPR is a nascent cellular pathway: the molecular components and physiological relevance of which are not well understood. Our lab has previously shown that SIRT7, a nucleus localized sirtuin, alleviates the mtPFS. Here we set up the experimental system of mtUPR by treating the synthetic triterpenoid 2-cyano-3,12-dioxooleana-1,9-dien-28-oic acid (CDDO) in HEK293T and Hela cells. We found that CDDO treatment leads to mitochondrial aggregation to a focus nearby the nucleus, and widespread increased electron density in the mitochondrial matrix under electron microscopy. We are going to analyze whether genetic manipulation of SIRT7 will also change these two newly identified mtPFS markers. In addition, we demonstrated global transcriptional downregulation of genes encoded by mitochondrial DNA in response to mtPFS, which can potentially relieve the stress and serve as a novel branch of mtUPR./nThese results not only broaden the understanding of inflammasome regulations and inflammasome-related diseases, but also provides important insights into detailed molecular mechanisms of mtUPR. Further, from the therapeutic standpoints, these findings also open novel avenues to treat diseases characteristic of chronic inflammation or mitochondrial dysfunction.

Advisor: Chen, Danica