Paper: Aguilar-Arnal, L., Katada, S., Orozco-Solis, R., and Sassone-Corsi, R. “NAD+-SIRT1 Control of H3K4 Trimethylation through Circadian Deacetylation of MLL1.” Nature Structural & Molecular Biology, (March 9, 2015). doi:10.1038/nsmb.2990.
The circadian clock allows almost all organisms to modulate their behavior and metabolic needs based on the light-dark cycle, which repeats every 24 hours. The clock then affects changes in metabolism by cyclically modulating the expression of genes throughout the day; while sleeping, for example, many genes are turned off, but others, such as genes involved in tissue repair and memory consolidation, are activated (Eckel-Mahan and Storm, 2009). The circadian clock is able to cause these effects by modulating genome-wide changes in transcription levels, sometimes via epigenetic modifications (Fang and Lazar, 2012).
One specific modification, trimethylation at Histone H3, Lysine 4 (H3K4), is known to activate transcription of a variety of genes throughout the body. Importantly for the circadian clock, mixed-lineage leukemia 1 (MLL1—an enzyme, not a cancer) trimethylates H3K4 (H3K4me3) of Dbp, a known promoter of circadian-regulated genes; this trimethylation helps to recruit CLOCK and BMAL1, which are two important transcription factors in the circadian cycle that cause changes in expression in almost all circadian-controlled genes throughout the entire 24-hour cycle.
Yet this single trimethylation event is insufficient to finely control the activity of so many genes, and it has been proposed that the circadian cycle might be able to recruit other epigenetic modifiers, particularly sirtuins (SIRTs) to modulate genetic changes. SIRT1, a deacetylase, is believed to have a major effect on the transcription of genes, as it has previously been shown that destruction of SIRT1 somehow prevents the rhythmic fluctuation of gene expression. This is important because this cyclic activation of downstream products implies that there could be a previously unknown link between SIRT1 and the circadian cycle. Interestingly, limiting the concentration of SIRT1’s cofactor, NAD+, can be used to regulate SIRT1 activity; NAD+ concentration in the brain fluctuates greatly over a 24-hour period and its concentration is regulated rhythmically by CLOCK-BMAL1 dimerization interaction, giving further evidence of a potential link between SIRT1 and the circadian cycle.
It thus seems likely that epigenetic modifications, especially deacetylation, are important for normal circadian rhythms; however, the mechanisms through which the circadian clock is able to modulate these epigenetic changes are currently unknown. Here the authors specifically study the activity of MLL1, the H3K4me3 of Dbp (and other genes), to determine if, and through what mechanism, the circadian clock is able to control MLL1 activity, and the subsequent downstream expression of circadian-regulated genes.
Since previous studies have shown a link between histone acetylation levels and H3K4me3, and thus gene activation, the authors first investigate a connection between MLL1 and SIRT1. The authors inhibit SIRT1 via in vitro inhibitors and in vivo mice Sirt1 knockouts, and ChIP assays show that H3K4 trimethylation levels increase in the Dbp region (promoter for master-regulatory genes of circadian clock), and on other genes that are regulated by the circadian cycle; trimethylation of non-circadian genes is not affected. This suggests that SIRT1 does affect H3K4me3 levels, but only in a specific set of genes, and the authors next show that changes in NAD+ concentration, SIRT1’s cofactor, can also affect these levels. Unsurprisingly, the addition of NAD+ and B-NMN (NAD precursor) decreases both H3K4me3 of Dbp and transcriptional activity of known MLL1 downstream targets. These two experiments suggest that changes to H3K4me3 levels can be directly attributed to SIRT1, likely via MLL1. Co-Immunoprecipitation assays further show that SIRT1 does bind to MLL1, and assays conducted at various time points in the circadian cycle show that this interaction is time-dependent, peaking consistently at certain times. SIRT1-MLL1 dimerization levels are then shown to be highest when H3K4me3 levels at Dbp are lowest, showing that SIRT1 binds to MLL1, controlling its activity and decreasing trimethylation of crucial MLL1 targets; these targets, when activated by MLL1, would regulate the normal cyclic expression of numerous other genes throughout the body.
The authors hypothesize that this MLL-SIRT1 interaction is not a simple binding interaction, but instead a deacetylation event. After demonstrating that MLL1 has acetylation domains, the use of specific combinations of HDAC inhibitors show that only sirtuins, and not other HDACs, could be responsible for MLL1 deacetylation; deacetylation assays demonstrate that SIRT1 is the only sirtuin capable of deacetylating MLL1. After identifying K1130 and K1133 as acetylation sites on MLL1 via Mass Spectrometry, the authors show that the presence of normal SIRT1, but not mutated or pharmacologically inhibited SIRT1, decreases acetylation of these specific residues both in vitro and in mouse livers. The deacetylation at these residues is shown to be cyclic, as maximum deacetylation occurs during periods of greatest SIRT1-MLL1 interaction, as demonstrated via Western Blotting in Sirt1 knockout mice; maximum MLL1 deacetylation also corresponds to near-peak NAD+ levels, showing interplay between metabolism and epigenetics, as NAD+ is able to regulate the epigenetic modifications caused by MLL1 via SIRT1.
Finally, and most importantly, the authors show in vivo that the presence of SIRT1, but not mutated SIRT1, reduces the activation of the Dbp promoter, and also reduces the expression of downstream circadian genes in a dose dependent manner. The authors have thus shown that deacetylation of MLL1 by SIRT1 downregulates Dbp, and Dbp’s targets, which is highly significant because these downstream genes have been previously shown to regulate the circadian clock throughout the entire body. This pathway involving SIRT1 and MLL1 is most seen clearly in Figure 5, which nicely summarizes the overall mechanistic results uncovered by the paper; the figure shows not only the interaction between SIRT1 and MLL1, but also the subsequent increase in the translation and expression of clock controlled genes.
Taken together, this study shows not only that SIRT1 is able to deacetylate MLL1, but also that these interactions decrease MLL1-mediated H3K4me3 of the Dbp promoter. The deacetylation of MLL1 prevents its normal activity, so Dbp cannot be trimethylated, and the subsequent recruitment of CLOCK, which is required for the transcriptional activation of a huge variety of other genes throughout the entire body, cannot occur. Since SIRT1 can be cyclically controlled via NAD+ levels, rhythmicity can be induced not only in SIRT1, but also in trimethylation of Dbp, and all of CLOCK’s and Dbp’s downstream genes. This is highly significant because the authors have established a previously unknown mechanistic link between epigenetic modifications and the cyclic oscillations in metabolism that are required for life; these findings support the idea that changes to NAD+ levels can actually cause changes in transcriptional expression of a wide variety of other genes. This is particularly interesting because H3K4me3 is known to promote transcriptional consistency (the stable and frequent transcription of genes) in most genes, and such robust expression of circadian genes (ie those influenced by CLOCK), is correlated to increased life span. If SIRT1 activity decreases H3K4me3 of upstream circadian genes, then circadian genes may have reduced transcriptional consistency, potentially decreasing lifespan. The authors thus suspect that better understanding the activity of SIRT1 and NAD+ may provide important information about aging and death, and possibly new treatment options related to diseases of aging, as well as sleep-related disorders.
References:
Eckel-Mahan, K., and Storm, D. “Circadian Rhythms and Memory: Not so Simple as Cogs and Gears.” EMBO Reports 10.6 (2009): 584–591. doi:10.1038/embor.2009.123
Feng, D., and Lazar, M. “Clocks, Metabolism, and the Epigenome.” Molecular Cell 47, no. 2 (July 27, 2012): 158–67. doi:10.1016/j.molcel.2012.06.026.
Hi Mike! Awesome job tackling a challenging article. I have a few broader implication questions. There are several things that can throw off or shift circadian rhythms for humans such as jet lag and puberty. How does the metabolic regulation laid out in this article relate to or account for this? In the case of jet lag, are these shifts caused by the changes in NAD+ levels continuing to occur on the same schedule, regardless of potential shifts in the time on the clock? What do you think eventually shifts in the metabolic pathway that corrects the jet lagged feeling? Could you hypothesize which point in the pathway may be disrupted or shifted in the case of puberty (the dimerization of Sirt1 and MLL1, the deacetylation, or the NAD+ levels?)? Is it possible that people with sleep regulation disorders have metabolic dis-regulation of this pathway, perhaps due to some kind of mutation that would prevent Sirt1 from deacetylating MLL1? This kind of dis-regulation can be inconvenient and challenging, leaving people feeling exhausted. Is it possible that there is a place in this pathway that could be a target for pharmaceutical companies for drugs that could lessen the time it takes to overcome jet lag or aid people with sleep regulation disorders? Thanks!
Hi Kelly,
Those are interesting questions. The entire circadian clock is actually regulated in the superchiasmatic nucleus (SCN) of the hypothalamus, which is the master regulator of the entire circadian clock, and thus all downstream effects of the clock. The SCN receives direct enervation from the eye, allowing these few cells to adjust to exterior light/dark cycles, thus syncing the entire clock. The SCN can then cause the normal effects seen throughout the circadian cycle, including cyclic changes in NAD levels. If the SCN is improperly regulated due to jetlag, then the downstream changes to NAD concentration will also be incorrect, and likely off by several hours until the SCN is able to catch up to local time. Interestingly, decreased SIRT1 activity can be correlated to longer periods of jet lag in mice, but SIRT1 overexpression prevents this effect. Since we know that SIRT1 activity is so directly controlled by NAD+ concentration, it seems likely that NAD+ levels do play a crucial role in jet lag. These effects, though, are likely caused farther upstream, as if the SCN is slow to adapt, then NAD+ levels are correspondingly behind local time, making SIRT1 activity, and its related effects on gene expression, also incorrect based on the exterior time in the new country.
As evidenced by the current lack of treatment, the literature hasn’t quite figured out exactly how jet lag works, or how the clock manages to eventually catch up. The most recent thinking, though, is that jet lag is actually correlated to arginine vasopressin (AVP), as AVP antagonists in mice were shown to combat the effects of jetlag. AVP deficient mice are able to keep time very precisely (like a normal circadian clock) yet their clock is actually capable of making huge jumps in time, which would be required to sync with the new light-dark conditions in a new country. The authors monitored downstream gene expression and found that the clock can quickly ‘jump’ ahead or back to match the new light/dark pattern in the new country, and since this allows for normal gene expression and regulation, the effects of jet lag could be stopped.
As for sleep disorders, which are also a temporary symptom of jet lag, it is possible that patients could have a mutation in SIRT1, preventing deacetylation of MLL1. It does seem more likely, though, that the real reason is far downstream of SIRT1, as those with sleep regulation disorders do not show dysregulation in other major metabolic pathways; if the patient did have a SIRT1 mutation, sleep disturbances would be the least of the patient’s problems, as the deacetylation of MLL1 affects literally thousands of genes in a huge variety of metabolic processes, and SIRT1 has been implicated in a wide variety of functions that are crucial to human health. The authors of the arginine vasopressin study did hypothesize in their discussion, though, that AVP may also be involved in long-term sleep problems, and AVP antagonists could serve as a future treatment for sleep disorders. Since the SCN does mediate normal sleep-wake cycles in the body, an AVP antagonist that helps cure jet lag may also help cure sleep problems, as the circadian cycle could be reset; this would force (or trick) the body into realizing that the exterior environment is dark, and that the body should prepare to sleep.
The link between SIRT1 and puberty, though, is much less well defined. Thee dimerization or deacetylation are again unlikely be affected, as normal SIRT1 activity is needed to all of the changes in gene expression that occur throughout the day. Although teenagers do experience changes in the circadian clock (ie go to bed later, sleeping fewer hours), the prevention of SIRT1 activity probably is not the cause, as teenagers generally do not experience jet-lag type symptoms for several years. However, SIRT1 has been loosely implicated with the PcG complex, which is a group of transcriptional repressors that are involved in genome-wide changes in puberty; this link however, has not been deeply explored. It seems likely that SIRT1 and PcG are able to interact, and this may allow either SIRT1 or PcG to slightly modify their activity, causing the activation (or suppression) of different genes involved in puberty, which could then cause the changes in sleep in teenagers.
Belden, William J., and Jay C. Dunlap. “Aging Well with a Little Wine and a Good Clock.” Cell 153, no. 7 (June 20, 2013): 1421–22. doi:10.1016/j.cell.2013.05.055.
Chang, Hung-Chun, and Leonard Guarente. “SIRT1 Mediates Central Circadian Control in the SCN by a Mechanism That Decays with Aging.” Cell 153, no. 7 (June 20, 2013): 1448–60. doi:10.1016/j.cell.2013.05.027.
Lomniczi, Alejandro, Hollis Wright, and Sergio R. Ojeda. “Epigenetic Regulation of Female Puberty.” Frontiers in Neuroendocrinology 36 (January 2015): 90–107. doi:10.1016/j.yfrne.2014.08.003.
Yamaguchi, Yoshiaki, Toru Suzuki, Yasutaka Mizoro, Hiroshi Kori, Kazuki Okada, Yulin Chen, Jean-Michel Fustin, et al. “Mice Genetically Deficient in Vasopressin V1a and V1b Receptors Are Resistant to Jet Lag.” Science 342, no. 6154 (October 4, 2013): 85–90. doi:10.1126/science.1238599.
Good summary, Mike! Thanks for your analysis and insights. I was particularly interested in the fact that the authors linked changing the concentration of NAD+ with sirtuin activity. This makes sense intuitively because NAD+ is required for SIRT1 to function. Additionally, if NAD+ concentration is changing over a 24 hour time period, does this mean that certain metabolic pathways are more active during different times of day? There are some metabolic pathways that are more NAD-intensive than others (ie. involve more redox chemistry), so maybe these pathways are more performed more frequently at times of day when NAD+ concentration is high? Just a few thoughts – nice job.
Hi Zach,
You definitely bring up a good point! The effects of rhythmicity in NAD levels extend far beyond sirtuins, and have been implicated in a wide variety of other pathways. It was previously thought that NAD concentrations were stable in the body, yet research over the last few years has debunked this idea, and instead shown that its concentration actually varies quite dramatically over a 24-hour period; the circadian clock causes these oscillations. Thus, essentially any pathway that involves the use of a NAD cofactor is affected by these cyclic changes, including the core metabolic pathways like the Krebs cycle and ETC, which could be slowed in response to decreasing NAD levels in the cell. Beyond the obvious examples, everything pathway that uses NAD, from insulin secretion to granulocyte colony-stimulating factor, which is essential for differentiation of some white blood cells, can be effected by the circadian clock.
Imai, Shin-ichiro. “‘Clocks’ in the NAD World: NAD as a Metabolic Oscillator for the Regulation of Metabolism and Aging.” Biochimica et Biophysica Acta 1804, no. 8 (August 2010): 1584–90. doi:10.1016/j.bbapap.2009.10.024.
Hi Mike! Awesome job with an interesting paper. I have a question about the implications of this paper. I see that you say that maximum MLL1 deacetylation occurs near peak NAD+ levels. I wonder then, if dietary supplementation with nicotinamide could help repair a faulty circadian rhythm, or, if the circadian rhythm is already “normal,” if it could throw it off in some way. I think that the metabolic/chromatin remodeling connection that is revealed in this paper is perhaps its most intriguing facet- I don’t often think of epigenetics and metabolism being related at a regulatory level. This is kind of straying off-topic, but I wonder if this regulatory connection exists in certain metabolic disorders- perhaps the absence (or presence) of some metabolic products could affect epigenetics in such a way as to cause expression of genes that cause symptoms. I would be interested in reading more papers that make that kind of connection.
Hi Gabbie,
Using NAD+ supplementation would definitely be an interesting experiment to conduct. I’m not positive that dietary supplementation would work to drastically increase NAD levels in the brain, but a few previous studies have demonstrated that intranasal administration of NAD, or NAD precursors, is effective, as the NAD is able to cross the blood brain barrier. If NAD levels are directly causing a problem with the circadian clock, then this type of supplementation would be useful, as a patient could just take a dose of NAD whenever levels in the brain are supposed to be increasing. Since the NAD could directly enter the brain, it could quickly have an effect and cause normal activity of SIRT1 and MLL1. NAD has been shown to decline with age, and there are changes in the circadian cycle with age, so it definitely seems like a promising method for treatment (1).
As for your second question, there is definitely a lot of work currently being conducted. There is almost definitely a link between metabolism, epigenetics, and disease, but the interconnection between these areas has not yet been thoroughly investigated in many cases. However, in the area of cancer, it is known that accumulations of succinate or fumarate, which can be caused by mutations in succinate dehydrogenase or fumarate hydratase, modifies the behavior of various dioxygenases. Some of these affected dioxygenases are required for DNA and histone methylation, and specific changes in their activity have been linked to tumorigenesis (2).
1: Imai, Shin-ichiro, and Leonard Guarente. “NAD+ and Sirtuins in Aging and Disease.” Trends in Cell Biology 24, no. 8 (August 2014): 464–71. doi:10.1016/j.tcb.2014.04.002.
2: Kaelin Jr., William G., and Steven L. McKnight. “Influence of Metabolism on Epigenetics and Disease.” Cell 153, no. 1 (March 28, 2013): 56–69. doi:10.1016/j.cell.2013.03.004.
Hey mike! Great work on outlining the paper, and thank you for the thorough introductory; it was much better than the paper’s!
The constant regulation of the circadian rhythms is very interesting. I took from the paper that the main point was activation of SIRT1 by NAD+. The authors concluded that this finding has implications in therapy and aging diseases. Could you speculate as to how therapy would work? The trouble I’m having with an application is that NAD is present as a cofactor in so many metabolic pathways. I can’t see the solution to circadian disregulation as simply increasing the concentration of the cofactor, because it could put stress on, or shift equilibria of other systems. Furthermore, let’s say we looked into a treatment that was able to selectively increase NAD concentrations strictly in the liver and it was able to make its way to SIRT1 in the circadian pathway… The paper states that NAD is required for activation of SIRT1, but does this also mean that high endogenous NAD is enough to induce activation of the circadian pathway? If you wouldn’t mind speculating with me, or correcting me if I’m misunderstanding how the pathway works, it would be much appreciated!
Again, good work!
Hi Ryan,
While the authors do link the results of their paper to a potential treatment, it is obviously important to remember that the study was conducted in mice, not humans, and that diseases of aging are a multifaceted conditions, and likely do not have one specific cause that could be so simply corrected. With that in mind, it is known that a disease like Alzheimer’s has been linked to global changes in both methylation and acetylation of histones of various genes; thus, SIRT1 could serve as a potential treatment method. If, for example, global acetylation levels of a particular histone are too high, it may be possible to increase the activity of SIRT1, which acts as a deacetylase, to correct this problem. Since acetylation and methylation can directly influence gene transcription, the activation of SIRT1 could hopefully return the body to normal gene expression levels that are not associated with disease. Since SIRT1 only targets acetylation at certain histones, it might be necessary to consider modifying the activity of other SIRTs, or even other categories of HATs, as well.
It would be important to investigate, though, if this type of treatment is actually viable, as SIRT1 can control the function of many genes. You are definitely right to be concerned about stressing other pathways and systems, as an increase in SIRT1 activity has been correlated with lung cancer, prostate cancer, and leukemias (1). Thus, the side effects of using SIRT1 may be too great for it to be a practical treatment method, and it may be more practical to look for targets further downstream, which might cause fewer side effects.
The control of SIRT1 is naturally complex, which is not entirely surprising for an enzyme that plays a role in the regulation of so many different processes. Regulating NAD levels is just one way that SIRT1 can be controlled, and it seems to act more like a negative regulator; the absence of the cofactor prevents enzyme activity. The presence of NAD supplementation itself has been shown to cause some small effects on SIRT1 activity, but there are other regulatory controls present, such as allosteric binders and some protein-protein interactions. For example, it is known that NMNAT, which is involved with NAD salvage, is known to bind to SIRT1 and affect its activity (2). There are numerous other ways that SIRT1 can be both up and down-regulated, so a simple change in NAD would not necessarily be enough to cause any downstream changes in gene expression.
1: Calvanese, Vincenzo, Ester Lara, Arnold Kahn, and Mario F. Fraga. “The Role of Epigenetics in Aging and Age-Related Diseases.” Ageing Research Reviews 8, no. 4 (October 2009): 268–76. doi:10.1016/j.arr.2009.03.004.
2: Zhang, Tong, Jhoanna G. Berrocal, Kristine M. Frizzell, Matthew J. Gamble, Michelle E. DuMond, Raga Krishnakumar, Tianle Yang, Anthony A. Sauve, and W. Lee Kraus. “Enzymes in the NAD+ Salvage Pathway Regulate SIRT1 Activity at Target Gene Promoters.” Journal of Biological Chemistry 284, no. 30 (July 24, 2009): 20408–17. doi:10.1074/jbc.M109.016469.
Hey Mike,
Great paper, and great overview!
I was curious as to how specific the NAD+ peak mediated response in SIRT1 was, specifically as compared to other electron transporters such as FAD. Alternatively, what circumstances do you believe would need to be extant before the body came to utilize concentrations of another electron transporter in the event of a prolonged lack of change regarding NAD+ concentration?
Hey Anthony,
Since SIRT1 uses NAD as a cofactor, other electron transporters would not affect deacetylation activity. SIRT1 physically binds to NAD, so something like FAD would not be able to bind, and thus would not affect the enzyme. If NAD+ levels simply remained constant, than this regulatory control mechanism would cease to function normally, and SIRT1 would be up-regulated, as NAD is normally used to turn the enzyme off.
Hi Mike! While reading this, I was wondering if the NAD+ concentration fluctuates just based on metabolic activity, or if there is a way that the body sequesters this cofactor so that it cannot be used at certain times. Is it a simple depletion of NAD+ over a normal time period, or does the body actively control these levels? Since NAD+ is used for a number of different reactions in the body, is this a widespread phenomenon or is it localized to certain parts of the body? Do you think there could be some kind of dietary supplementation to help those who suffer from sleep related diseases or even those suffering from jet lag? Thanks for choosing an interesting paper and writing a great summary!
Hi Matt,
Those are really good questions. The concentration of NAD fluctuates simply based on the circadian clock, which functions by controlling transcription levels; specifically, the clock can turn on or off genes that synthesize NAMPT, which catalyzes the rate-limiting step of NAD. My understanding of the cyclic changes in concentration is that the body regulates NAD concentrations at the transcription level, and not directly at the degradation level. If, for example, the body wants to downregulate SIRT1, it would slow the transcription of genes that synthesize NAD, and this has been shown to be enough to effectively limit the activity of other sirtuins (1). SIRT1 could probably remain temporarily active until NAD+ levels fall, and then would remain inactive or less active until the circadian clock genes up-regulated the production of NAD.
As you mention, the control of NAD is definitely critical throughout metabolic pathways in the body, and needs to be carefully controlled. NAD levels obviously fluctuate in the brain, and other studies have show that its concentration varies in the liver as well (2). I couldn’t find much information about NAD oscillation in other peripheral tissues, but this may simply be due to a lack of active research in this area. Since NAD obviously plays such an important role in metabolism, though, I would speculate that NAD levels do remain relatively constant in other parts of the body. The changes in NAD in the brain are probably effective enough to cause the system-wide changes seen during the 24-hour cycle, likely because regulators like SIRT1 are able to effect such great change on numerous pathways. Thus, these changes in the liver and the brain are probably sufficient to control metabolism, without needing to regulate NAD levels in every cell of the body.
Modifying the circadian clock could definitely be a future treatment that helps with jet lag. Although supplementation or SIRT1 or NAD would probably be problematic because they interact with so many different pathways, other researchers have shown that arginine vasopressin (AVP) antagonists are able to treat both sleep diseases and jet lag in mice. This was an extremely interesting study because it showed that the mice could keep perfect time via their circadian clock, but simple AVP antagonist supplementation actually permitted the clock to jump forward (or back), and sync with the exterior light/dark conditions in the new location. Thus, the levels of NAD production could be normalized based on the new time, allowing SIRT1 to function normally, allowing for normal transcription of downstream genes; this would prevent any symptoms of jet lag from developing. In a similar manner, an AVP supplement could force the circadian clock into realizing that it is dark outside, and thus that the body should prepare to sleep. The supplement could force the clock to jump forward to a time that would trigger the molecular basis of sleep, and thus would help treat long-term sleep disorders (3).
1: Peek, Clara Bien, Alison H. Affinati, Kathryn Moynihan Ramsey, Hsin-Yu Kuo, Wei Yu, Laura A. Sena, Olga Ilkayeva, et al. “Circadian Clock NAD+ Cycle Drives Mitochondrial Oxidative Metabolism in Mice.” Science 342, no. 6158 (November 1, 2013): 1243417. doi:10.1126/science.1243417.
2: Mohawk, Jennifer A., Carla B. Green, and Joseph S. Takahashi. “CENTRAL AND PERIPHERAL CIRCADIAN CLOCKS IN MAMMALS.” Annual Review of Neuroscience 35 (2012): 445–62. doi:10.1146/annurev-neuro-060909-153128.
3: Yamaguchi, Yoshiaki, Toru Suzuki, Yasutaka Mizoro, Hiroshi Kori, Kazuki Okada, Yulin Chen, Jean-Michel Fustin, et al. “Mice Genetically Deficient in Vasopressin V1a and V1b Receptors Are Resistant to Jet Lag.” Science 342, no. 6154 (October 4, 2013): 85–90. doi:10.1126/science.1238599.
Hey Mike,
Very interesting and unique article, which I enjoyed. The part I found most intersting was the portion in the introduction where the authors stated “Remarkably, NAD+ oscillation is dictated by CLOCK–BMAL1, which directly controls the gene Nampt, encoding the nicotinamide phosphoribosyltransferase (NAMPT) enzyme, which catalyzes the rate-limiting step in the NAD+ salvage.” The authors also stated that pharmocalogical inhbition of SIRT1 appears to alter circadian rhythyms. This seems like an incredibly useful drug for individuals forced to work a night shift, but the target of the drug, to me, seems one step removed, in that the drug targets a protein that functions to turn off another protein (SIRT1 controls acetylation of NAMPT which encodes for nicotinamide phosphoribosyltransferase) rather than just targeting the functional protein itself. I was wondering if you think think that targeting nicotinamide phosphoribosyltransferase might be more effective in altering the cricadian cycle than targeting SIRT1? If not, are there any reasons to not target nicotinamide phosphoribosyltransferase or to suggest that targeting SIRT1 might be more effective?
I think you are definitely correct that some of the pathways from this article could serve as a treatment for those working night shifts, and, as others have pointed out, those that are jet-lagged or suffer from long-term sleep problems. I do agree that targeting NAMPT may be more effective than targeting SIRT1, especially because NAMPT is a much smaller target. SIRT1 indirectly controls activity of thousands of pathways, and modifying SIRT1 activity has been shown to cause various diseases, so any type of life-long modification of SIRT1 seems like a poor treatment plan. Thus, NAMPT could serve as a slightly more attractive target. However, targeting NAMPT also presents problems because this protein mediates the synthesis of NAD+; NAMPT converts nicotinamide to nicotinamide mononucleotide (NMN), but this is the major way that NAD is synthesized in the mammalian body. So while this should be an effective way to down-regulated NAD production, this type of treatment might cause system wide changes in metabolism due to the loss of this important cofactor. Thus, targeting NAMPT and SIRT1, while effective, would probably not be the best therapeutic option to alter the circadian rhythm. Instead, other research groups have targeted molecules that are involved in downstream pathways that have less of a global regulation function. These smaller pathways can still have great effects on the circadian clock, but do so with fewer side effects. A lot of studies have shown that arginine vasopressin antagonists are highly successful at treating sleep related disorders by literally allowing the clock to ‘jump’ forward, which would be effective here to treat people that do shift work (1). Other possible targets include prokinetictin2 and the famous melatonin, which all regulate the circadian clock (2); even though these molecules do not operate directly on the pathways from this paper, they would probably serve as a more useful target/treatment method than modifying SIRT1 or MLL1 directly.
1: Yamaguchi, Yoshiaki, Toru Suzuki, Yasutaka Mizoro, Hiroshi Kori, Kazuki Okada, Yulin Chen, Jean-Michel Fustin, et al. “Mice Genetically Deficient in Vasopressin V1a and V1b Receptors Are Resistant to Jet Lag.” Science 342, no. 6154 (October 4, 2013): 85–90. doi:10.1126/science.1238599.
2: Toh, KL. “Basic Science Review on Circadian Rhythm Biology and Circadian Sleep Disorders.” Ann Acad Med Sing 37, no 8 (August, 2008)
Hey Mike – great analysis on a pretty dense article. My question pertains to your final sentence, where you say that more insight into these phenomena may lead to a greater understanding of diseases of aging. We know that oxidative damage occurs naturally and has been linked to diseases of aging, and since NAD+ is the oxidized form of NADH, is there a correlation here? The authors correlate NAD+ concentration to SIRT activities. Do you think that an oxidative environment resulting from the presence of ROS’s would increase concentrations of NAD+ and mess with these sirtuins and eventually circadian rhythm? I’d love to hear your input.
Hi Besher,
That is definitely an interesting idea, and relates pretty well to everything we have been learning about. The authors don’t really address the idea of redox balance or ROS, but instead use the idea of transcriptional consistency to explain how this may affect diseases of aging. With increasing age, genes are actually transcribed much less frequently and contain more errors, and this lack of consistent and stable transcript has been linked to causing many problems, including diseases of aging. I still think you raise an interesting point though, as NAD+/NADH can have some effect on the on the oxidative status of the cell. It has been shown in mice that the NAD:NADH ratio actually decreases significantly in many tissues with increasing age, and this decrease is accompanied by increased DNA damage, increased (undesirable) protein modifications, and decreased SIRT1 activity (1). This does seem to imply that NAD not only effects SIRT1 activity, but can also have an effect on aging and genetic damage, which could lead to diseases of aging. I don’t think that NAD itself is causing the damage here, as it seems that NADH may be more problematic, or perhaps it is the ratio between the two that is most critical. We do also know that the redox balance is tightly controlled with the cell and is affected by a wide variety of molecules, so it would be difficult to implicate oxidative damage directly to NAD; however, there does appear to be some evidence to suggest a link between the two, and it will be interesting to see if this becomes an area of further research.
Interestingly, NAD+ has actually been shown to have a protective effect on axons the brain, and such protective effects have been loosely correlated to decreased prevalence of diseases of aging. Using RNAi knockdown, a previous study has shown that SIRT1 mediates some of the protective effects of NAD+, so it is actually possible that an oxidized environment that increases concentration of NAD+ could actually be helpful in the brain (2). If SIRT1, for example, became more active, then there would be less trimethylation activity by MLL1, and causing fewer downstream genes to become activated; this could definitely cause changes in the circadian clock because the rhythmic expression of genes would be effected, but this could be somewhat positive, as some of these genes that are now turned off may actually cause neuronal damage, leading to diseases of aging.
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p>1: Braidy, Nady, Gilles J. Guillemin, Hussein Mansour, Tailoi Chan-Ling, Anne Poljak, and Ross Grant. “Age Related Changes in NAD+ Metabolism Oxidative Stress and Sirt1 Activity in Wistar Rats.” PLoS ONE 6, no. 4 (April 26, 2011): e19194. doi:10.1371/journal.pone.0019194.
2: Belenky, Peter, Katrina L. Bogan, and Charles Brenner. “NAD+ Metabolism in Health and Disease.” Trends in Biochemical Sciences 32, no. 1 (January 2007): 12–19. doi:10.1016/j.tibs.2006.11.006.