Sirtuins,, What Are Sirtuins and What Do They Do? Stephen Rose By Stephen Rose June 5, 2022

Lifespan Logo WHAT WE DO EDUCATION NEWS EVENTS RESEARCH GET INVOLVED SERVICES Donate What Are Sirtuins and What Do They Do? Stephen Rose By Stephen Rose June 5, 2022 Resveratrol What Are Sirtuins and What Do They Do? Publisher: Lifespan Extension Advocacy Foundation Date Published: 06/05/2022 Date Modified: 06/23/2022 Sirtuins are a family of seven SIRT enzymes found in the nucleus, cytoplasm, and mitochondria of human cells. These enzymes exist in nearly all species and perform many functions that are associated with increased longevity [1]. SIRT3, SIRT4, and SIRT5 are primarily found in mitochondria; SIRT1, SIRT6, and SIRT7 are mostly located in the nucleus; and SIRT2 is found in the cytoplasm, occasionally accompanied by SIRT1 [2,3]. What functions do sirtuins perform and how? Chromatin is made of two things: DNA and histones. Histones are molecular spools, and DNA is the thread that wraps around these spools. Sirtuins regulate how much DNA is wound around the histones and how much is open to the surrounding environment, as DNA that is wrapped around the spool can’t be expressed. First and foremost, sirtuins regulate the spooling and unspooling process in response to environmental changes, protecting our DNA from damage. Through this mechanism and others, sirtuins regulate the cell cycle, DNA repair, and mitochondrial energetics [1]. Sirtuins do this by manipulating histones, specifically their tails, which are made from lysine. Lysine is a protein building block that is attracted to water. As the inside of a cell is mostly water, lysine is found on the outside surface of proteins. This long tail projecting out from the protein’s surface makes it a reaction hot spot. Sirtuins are shaped in a way that enables them to change the design of lysine. However, these changes have ripple effects, which cause the histones to spool up surrounding DNA [4]. Thus, sirtuins are very good at dialing down gene expression in many ways. Mammalian SIRT1 alone uses this mechanism on at least 40 different molecules [5]. Sirtuins and gene expression In the nucleus, DNA is spooled around molecules called histones. Sirtuins modify histones, epigenetic enzymes, transcription factors, and transcription factor regulators to control gene expression [6-9]. Sirtuins control access to DNA through deacetylation of histones and epigenetic enzymes, such as histone methyltransferase. Sirtuins can remove an acetyl group from lysine on histone proteins in a process called deacetylation. This causes the histones to wind up DNA, limiting its accessibility to the machinery of gene expression [10]. Sirtuins can also increase gene expression. In this case, sirtuins deacetylate non-histone molecules called histone methyltransferases (HMTs). HMTs modify the lysine tails of histones by adding methyl groups. Adding methyl groups to lysine has the opposite effect on histones as that of deacetylation: it unwinds DNA from histones, making genes available for expression or repair. SIRT1 recruits and activates histone methyltransferase (HMT) by deacetylating a particular lysine on it [8]. Sirtuins also deacetylate transcription factors to control patterns of gene expression. Transcription factors are proteins that promote or repress gene expression by helping or hindering the attachment of RNA polymerase to DNA. The job of RNA polymerase is to copy genes encoded in DNA into a form that can be used for making proteins. Sirtuins can control this process by deacetylating the lysine residues present on transcription factors [6]. The transcription factors p53, FoxO1, FoxO3, FoxO4, FoxO6, and NF-κB, which can influence longevity, are among those influenced by sirtuins [11-13]. Sirtuins promote the expression of FoxO transcription factors, which, in turn, promote the activation of longevity genes. Activation of the FoxO family of transcription factors has long been associated with longevity. Sirtuins deacetylate lysine on FoxO transcription factors, which enables their entry into the nucleus. Upon entry, they bind to DNA, which promotes the expression of genes that improve stress resistance, metabolism, cell cycle arrest, and apoptosis [12]. Many of sirtuins’ effects on longevity appear to be mediated through FoxO gene expression. ADVERTISEMENT Longevity-associated gene expression patterns driven by FoxO transcription factors Sirtuins heavily influence FoxO transcription factor activity through complex mechanisms. Sirtuins enhance FoxO activity, and things that stimulate the insulin and IGF-1 receptors prevent FoxO activity. Insulin and IGF-1 both include PI3K-AKT in their signaling cascades, and PI3K-AKT activity prevents FoxOs from entering the nucleus. Therefore, insulin and IGF-1 can shut down FoxO activity; sirtuins can turn it back on. For example, glucose stimulates the release of insulin. Insulin attaches to the insulin receptor, which, in turn, prompts PI3K-AKT to disable FoxO transcription factors. Other changes to FoxO proteins, such as methylation and ubiquitination, also affect these processes. These processes can change the concentration of available FoxO transcription factors and when, where, and how FoxO proteins bind to DNA [14]. Sirtuins and FoxO affect how cells clear out junk FoxO activation increases autophagy and the activity of the ubiquitin-proteasomal system to clean up cells. One mechanism by which FoxOs may act as pro-longevity factors is through the maintenance of protein balance (proteostasis). Cells continually produce new proteins and get rid of old ones. This is done for housekeeping purposes and to adapt to a changing environment. FoxO genes promote clearance of unwanted cellular proteins in two ways: autophagy and the ubiquitin proteosome system. Autophagy, a cell’s consumption of its own organelles, is a programmed response to a lack of food that serves two main functions. It gets rid of cellular garbage and uses it to produce energy in the cell when times are lean [14,15]. Ubiquitin is a protein that is attached to lysine to mark a given protein for destruction. The proteasome, a cellular organelle that deconstructs proteins, relies on ubiquitin. This system helps cells keep the right balance of working proteins based on their needs [16]. Aging is associated with the accumulation of excess damaged proteins resulting from compromised proteasomal activity. This phenomenon is especially pronounced in muscle, liver, and heart tissue. Additionally, abnormalities in proteasomal activities exacerbate neurodegenerative diseases, such as Parkinson’s, Alzheimer’s, and Huntington’s disease. Therefore, sirtuin activation through FoxO-mediated effects on autophagy and the ubiquitin-proteasomal system could help slow the progress of these age-related diseases [12]. Inflammation and oxidative stress FoxO expression limits oxidative stress. FoxO transcription factor activity encourages the upregulation of gene sets that better enable organisms to cope with oxidative stress, increasing the production of antioxidants such as MnSOD, catalase, and GADD45. This shift in gene expression is thought to decrease the risk of certain cancers and other age-associated diseases [12]. Sirtuins depress NF-κB transcription through multiple mechanisms to reduce inflammation. SIRT1 inhibits NF-κB signaling directly by deacetylating the NF-κB complex. SIRT1 also stimulates oxidative energy production via the activation of AMPK, PPARα and PGC-1α, which also inhibit NF-κB signaling and suppress inflammation [13]. SIRT6 disables the promoter-transcription factor for the expression of the NF-kB gene [6]. Sirtuins regulate transcriptional regulators such as PGC-1α, which is a coactivator of many genes, including those that produce mitochondrial DNA [5]. PGC-1α is present in both the nucleus and the mitochondria, and it interacts with SIRT1 in both locations [17]. SIRT1 binds to and activates PGC-1α through deacetylation, affecting gene expression, energy production, and mitochondrial biogenesis. PGC-1α controls the number of mitochondria produced in cells (mitochondrial biogenesis). PGC-1α expression also induces the production of glucose from other energy sources, such as amino acids, fatty acids, glycerol (gluconeogenesis). It appears that these processes can occur independently or at the same time, depending on context [18]. This topic has been the subject of controversy and also involves questions about how exactly resveratrol works. The questions are “Does deacetylation of PGC-1α by sirtuins lead to increased production of mitochondria?” and “Does resveratrol increases mitochondrial biogenesis?” Although there is ongoing discussion about what the direct target of resveratrol is, there is clear consensus that resveratrol’s metabolic action converges on pathways involving AMPK, SIRT1, and PGC-1α [19]. It is not entirely clear whether PGC-1α deacetylation of sirtuins increases the production of mitochondria [19-21]. SIRT1, methylation, and cancer SIRT1 participates in DNA methylation but plays a controversial role in cancer development. SIRT1 is often found in the same place as the DNA methylation machinery of the cell, and it is associated with the intense methylation of tumor suppressor genes. Preventing such genes from being expressed should increase the risk of cancer, and SIRT1 is not found near these promoters when they are not hypermethylated. Therefore, sirtuins’ activation may encourage tumor growth under some circumstances. ADVERTISEMENT A paper from 2009 suggests that increasing sirtuin expression could cause cancer by inhibiting the tumor suppressor p53 [11]. A 2011 paper indicates that this effect on p53 is abolished by methyltransferase Set 7/9 [22]. The net effects of sirtuins on cancer development and prevention are still unclear [23]. When SIRT1 is inhibited in cancer cells, the tumor suppressor genes are expressed, at least in breast and colon cancer cells [24]. Therefore, SIRT1 inhibition, in some cases, may prevent or slow cancer progression. However, this is not the whole story. Sirtuins can also protect DNA from damage and oxidative stress, maintain genomic stability, and limit replicative lifespan. All of these factors tend to prevent the development of cancer [25]. SIRT1 is part of a large silencing complex that methylates DNA promoters specifically in areas where DNA damage has occurred. It can make the complex effective or ineffective in its role in the methylation process, depending upon what part of the complex it deacetylates. SIRT1 also exerts control over NoRC, which silences ribosomal RNA. If ribosomal RNA is silenced, the cell can not translate mRNA into protein. Protein synthesis in the cell is shut down. The primary function of sirtuins is to maintain genomic stability, which they do in three main ways. They regulate facultative and constitutive heterochromatin, they control cell-cycle progression, and finally, they are critical for DNA damage signaling. Sirtuins and telomeres As cells continue to divide, the end portions of the DNA (telomeres) gradually erode. An enzyme called telomerase slows this erosion down by assisting in the continual rebuilding of telomeres. When this process falls too far behind, genetic information is lost, and this can trigger the cell to self-destruct. If the cell doesn’t self-destruct, other DNA damage response programs can cause the cell to enter senescence. Telomeres and sirtuins are both linked to disease and aging, but their interaction is poorly understood. Recent evidence shows that telomere erosion depresses sirtuin activity. Conversely, increasing sirtuin activity stabilizes telomeres and improves telomerase-dependent disease states [26]. SIRT1 and SIRT6 play an important but poorly understood role in managing the energy state of the cell and the maintenance of heterochromatin. Interestingly, SIRT1 and SIRT6 are mobilized from telomeres to sites of DNA double strand breaks. Evidence indicates that SIRT1 inhibits telomerase activity, which would be expected to drive the cell to senescence or apoptosis. This cellular behavior is likely an evolutionarily conserved way to avoid cancer. However, SIRT1 depletion also results in increased genome instability and telomeric aberrations that contribute to decreased cell growth. This may be due to redundant reinforcing programs that protect damaged cells from becoming cancerous. SIRT1 is likely to play a direct role in the expression of telomerase [27]. SIRT2 and SIRT6 regulate DNA repair in several ways. DNA repair in humans doesn’t just involve DNA. It occurs within an organized structurally compact environment where both histones and DNA are present. A reorganization of both histones and DNA is necessary to allow the DNA repair machinery to gain access to damaged areas. An additional layer of complexity is added by the need to recruit and activate the machinery of DNA repair. Sirtuins turn on the machinery of DNA repair, create access to damaged areas of DNA, help limit oxidative stress, and regulate the cell cycle to facilitate DNA repair [28]. Can sirtuin activity be enhanced to increase human lifespan? Caloric restriction, exercise, and sirtuin-activating compounds (STACS) increase NAD+ and AMP and decrease circulating macronutrients, including lipids, amino acids, and glucose. In concert with these changes, insulin, GH, and IGF1 production also decrease [29]. ADVERTISEMENT These changes are sensed by SIRT1 and AMPK, which cue the body to shift metabolic processes in favor of energy conservation and self-maintenance. This allows the organism, human or otherwise, to make it through tough times. When environmental factors later change in a way that is more conducive to reproduction, the body shifts its metabolism to favor reproduction [30]. SIRT1 and AMPK regulate these changes in a variety of ways. SIRT1 uses NAD+ as a co-substrate to promote binding and modification of specific proteins associated with DNA repair, mitochondrial biogenesis, stress resistance, stem cell maintenance, autophagy, chromatin modifications, reduced inflammation, translation fidelity, and telomere maintenance [27,29,31]. AMPK activation complements these processes, as it further upregulates NAD+ to support more SIRT1 activity and adjusts how the body expends its energy resources. It does this by upregulating activities that break down unneeded body tissue and downregulating processes that build tissue unnecessarily or support reproduction. The net effect is autophagy: cellular garbage is used for energy. This not only cleans up cells and makes them more fit, it provides them with additional energy for maintaining vital functions. Resources are generally drawn away from energy-consuming processes that support reproduction [20,21,31]. The reason why is straightforward: having kids when you are starving or falling apart is not conducive to their survival. Evidence suggests that longevity benefits may be derived from increased sirtuin activity. If glucose is restricted from human cells, the lifespan of those cells is increased. It is believed that this increase is caused by increased NAD+ and sirtuin activity [20]. One way to restrict glucose from human cells is through caloric restriction. Caloric restriction is the only established intervention that has been shown to increase lifespan in every species, including primates. SIRT1 and SIRT3 are both believed to be essential for the processes associated with these longevity benefits. Caloric restriction and fasting both increase concentrations of SIRT3 and deacetylate many mitochondrial proteins [32]. Strategies for further enhancing sirtuin activity Ketone bodies like beta-hydroxybutyrate are known to promote sirtuin activity, which suggests that a ketogenic diet could increase it. However, not every aspect of a ketogenic diet promotes longevity. A low-carbohydrate diet that promotes lower insulin and glucose levels also tends to favor sirtuin activity [33,34]. Dry saunas, steam rooms, and hot tubs elevate body temperature. Elevated body temperatures drive the production of heat shock proteins, which increase NAD+ and activate sirtuins [35]. Melatonin is customarily used to reset circadian rhythms and induce sleep. It can also activate sirtuins and carries with it additional anti-aging effects. Melatonin is a powerful antioxidant, but high carbohydrate meals and melatonin are not good bedfellows, as melatonin tends to raise glucose [36]. Resveratrol and its chemical cousin, pterostilbene, have been reported to raise sirtuin levels and appear to do so with minimal side effects. 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Sacchetti, “Ketogenic Diet Modulates NAD+-Dependent Enzymes and Reduces DNA Damage in Hippocampus ,” Frontiers in Cellular Neuroscience , vol. 12. 2018 [34] P. Dabke and A. M. Das, “Mechanism of action of ketogenic diet treatment: Impact of decanoic acid and beta—hydroxybutyrate on sirtuins and energy metabolism in hippocampal murine neurons,” Nutrients, vol. 12, no. 8, pp. 1–19, 2020 [35] R. P. Patrick and T. L. Johnson, “Sauna use as a lifestyle practice to extend healthspan,” Exp. Gerontol., vol. 154, p. 111509, 2021 [36] J. C. Mayo, R. M. Sainz, P. González Menéndez, V. Cepas, D.-X. Tan, and R. J. Reiter, “Melatonin and sirtuins: A ‘not-so unexpected’ relationship,” J. Pineal Res., vol. 62, no. 2, p. e12391, Mar. 2017 About the author Stephen Rose  Stephen Rose Chris is one of the writers at Lifespan.io. His interest in regenerative medicine and aging emerged as his personal training client base grew older and their training priorities shifted. He started his masters work in Bioengineering at Harvard University in 2013 and is currently completing his PhD at SUNY Polytechnic University in Albany, NY. His dissertation is focused on the role of the senescent cell burden in the development of fibrotic disease. His many interests include working out, molecular gastronomy, architectural design, and herbology.