Intracellular nicotinamide adenine dinucleotide promotes TNF-induced necroptosis in a sirtuin-dependent manner.

Cellular necrosis has long been regarded as an incidental and uncontrolled form of cell death. However, a regulated form of cell death termed necroptosis has been identified recently. Necroptosis can be induced by extracellular cytokines, pathogens and several pharmacological compounds, which share the property of triggering the formation of a RIPK3-containing molecular complex supporting cell death. Of interest, most ligands known to induce necroptosis (including notably TNF and FASL) can also promote apoptosis, and the mechanisms regulating the decision of cells to commit to one form of cell death or the other are still poorly defined. We demonstrate herein that intracellular nicotinamide adenine dinucleotide (NAD(+)) has an important role in supporting cell progression to necroptosis. Using a panel of pharmacological and genetic approaches, we show that intracellular NAD(+) promotes necroptosis of the L929 cell line in response to TNF. Use of a pan-sirtuin inhibitor and shRNA-mediated protein knockdown led us to uncover a role for the NAD(+)-dependent family of sirtuins, and in particular for SIRT2 and SIRT5, in the regulation of the necroptotic cell death program. Thus, and in contrast to a generally held view, intracellular NAD(+) does not represent a universal pro-survival factor, but rather acts as a key metabolite regulating the choice of cell demise in response to both intrinsic and extrinsic factors.Cell Death and Differentiation advance online publication, 22 May 2015; doi:10.1038/cdd.2015.60.SCOPUS: ar.jinfo:eu-repo/semantics/publishe

Protein Kinase C Epsilon Promotes Cerebral Ischemic Tolerance Via Modulation of Mitochondrial Sirt5

Sirtuin 5 (SIRT5) is a mitochondrial-localized NAD(+)-dependent lysine desuccinylase and a major regulator of the mitochondrial succinylome. We wanted to determine whether SIRT5 is activated by protein kinase C epsilon (PKCε)-mediated increases in mitochondrial Nampt and whether SIRT5 regulates mitochondrial bioenergetics and neuroprotection against cerebral ischemia. In isolated mitochondria from rat cortical cultures, PKCε activation increased SIRT5 levels and desuccinylation activity in a Nampt-dependent manner. PKCε activation did not lead to significant modifications in SIRT3 activity, the major mitochondrial lysine deacetylase. Assessments of mitochondrial bioenergetics in the cortex of wild type (WT) and SIRT5−/− mice revealed that SIRT5 regulates oxygen consumption in the presence of complex I, complex II, and complex IV substrates. To explore the potential role of SIRT5 in PKCε-mediated protection, we compared WT and SIRT5−/− mice by employing both in vitro and in vivo ischemia paradigms. PKCε-mediated decreases in cell death following oxygen-glucose deprivation were abolished in cortical cultures harvested from SIRT5−/− mice. Furthermore, PKCε failed to prevent cortical degeneration following MCAO in SIRT5−/− mice. Collectively this demonstrates that SIRT5 is an important mitochondrial enzyme for protection against metabolic and ischemic stress following PKCε activation in the brain