NAD ⁺ and SIRT3 control microtubule dynamics and reduce susceptibility to antimicrotubule agents

Abstract

Nicotinamide adenine dinucleotide (NAD + ) is an endogenous enzyme cofactor and cosubstrate that has effects on diverse cellular and physiologic processes, including reactive oxygen species generation, mitochondrial function, apoptosis, and axonal degeneration. A major goal is to identify the NAD + -regulated cellular pathways that may mediate these effects. Here we show that the dynamic assembly and disassembly of microtubules is markedly altered by NAD + . Furthermore, we show that the disassembly of microtubule polymers elicited by microtubule depolymerizing agents is blocked by increasing intracellular NAD + levels. We find that these effects of NAD + are mediated by the activation of the mitochondrial sirtuin sirtuin-3 (SIRT3). Overexpression of SIRT3 prevents microtubule disassembly and apoptosis elicited by antimicrotubule agents and knockdown of SIRT3 prevents the protective effects of NAD + on microtubule polymers. Taken together, these data demonstrate that NAD + and SIRT3 regulate microtubule polymerization and the efficacy of antimicrotubule agents.N icotinamide adenine dinucleotide (NAD + ) is an endogenous dinucleotide that is present in the cytosol, nucleus, and mitochondria. Athough it serves an important role as a redox cofactor in metabolism, NAD + is also a substrate for several families of enzymes, including the poly(ADP ribose) polymerases and the sirtuin deacetylase enzymes (reviewed in refs. 1 and 2). The level of intracellular NAD + is regulated by many factors, including diet and energy status (3), axonal injury (4), DNA damage (5), and certain disease states (6), suggesting that NAD + -dependent signaling is dynamically modulated in diverse contexts.NAD + -dependent signaling can be induced by treatment of cells with exogenous NAD + , which increases intracellular NAD + levels and results in diverse effects in cells and animals. These effects include enhanced oxygen consumption and ATP production (7), as well as protection from genotoxic stress and apoptosis (3). Mice treated with nicotinamide riboside, a NAD + precursor that is metabolized into NAD + , have enhanced oxidative metabolism, increased insulin sensitivity, and protection from high-fat diet-induced obesity (8). These results demonstrate that NAD + -dependent pathways can enhance metabolic function and improve a variety of disease phenotypes.An NAD + -regulated pathway also inhibits axonal degeneration elicited by axonal transection (4). Treatment of axons with 5-20 mM NAD + markedly delays the axon degenerative process (9). Additionally, animals that express the Wallerian degeneration slow (Wld S ) protein, a fusion of the NAD + biosynthetic enzyme Nicotinamide mononucleotide adenylyl transferase 1 and Ube4a, exhibit markedly delayed degeneration of the distal axonal fragment after axonal transection (10), and expression of Wld S mitigates disease phenotypes in several neurodegenerative disease models (11)(12)(13)(14). Thus, understanding the intracellular pathways regulated by NAD + may be important for understanding the pa...

Poly(ADP-ribose) polymerase 1 is a novel target to promote axonal regeneration

“…Indeed, it was recently reported that in neurons, PARP1-dependent NAD + depletion inhibits the activity of the NAD + -dependent sirtuin 1 (43), which is critical for axonogenesis and neurite growth (49)(50)(51)(52). Likewise, the dynamic modulation of microtubules, essential to growth cone motility highly depends on NAD + levels through activation of mitochondrial sirtuin 3 (53). As such, a decline in sirtuin activity due to PARP1-dependent NAD + depletion might have a profound impact on neurite growth in an inhibitory environment.…”

SIRT3, PP2A and TTP protein stability in the presence of TNF‐α on vincristine‐induced apoptosis of leukaemia cells

“…Transfection of pCMV3‐His‐SIRT3 reduces the microtubule‐destabilizing effect of VCR in leukaemia cells. Consistent with this, prior studies have revealed that transfection with a SIRT3‐expressed vector reduced the capability of vinblastine to induce disassembly of microtubule polymers in breast cancer MCF‐7 cells . Notably, Wang et al found that treatment of U937 cells with a SIRT3 inhibitor, brRESV, down‐regulated SIRT3 expression, but did not induce microtubule destabilization and mitotic arrest.…”

“…Harkcom et al found that overexpression of SIRT3 suppresses vinblastine‐induced microtubule destabilization and G 2 /M arrest. Thus, the involvement of SIRT3 in VCR cytotoxicity was analysed.…”

Subcellular localization and mitotic interactome analyses identify SIRT4 as a centrosomally localized and microtubule associated protein

“…described for the tumor suppressor RITA (RBP-J and tubulin-associated protein) that interacts with HDAC6 and thereby modulates levels of K40-acetylated -tubulin and microtubule dynamics 71 . Lastly, the NAD + -SIRT3 axis has been also implicated in the regulation of microtubule dynamics and chromosomal alignment during mitosis 32,33 . However, this function of SIRT3 is likely mitochondrial-based, given that SIRT3 is predominantly found in mitochondria ( Fig.…”

“…SIRT3 regulates, in a direct or indirect (i.e. mitochondria-dependent) manner, microtubule dynamics and chromosomal alignment during mitosis by currently unknown mechanism(s) 32,33 . SIRT4 could also play an extramitochondrial role in microtubule dynamics, given that SIRT4 interacts with Leucinerich protein 130 (LRP130) 24,34 , a multi-domain and dual-function protein that binds to the microtubule-associated protein MAP1S and integrates mitochondrial transport and the microtubule cytoskeleton in interphase 35 .…”