Abstract
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.
Highlights
Aberrant energy metabolism following cerebral ischemia/reperfusion induces mitochondrial impairments such as reduced respiration, free radical generation, and release of proapoptotic factors[1,2]
The main goal of this study was to investigate whether PKCε is involved in regulating SIRT3 and Sirtuin 5 (SIRT5) and whether these sirtuins are involved in PKCε-mediated neuroprotection following cerebral ischemia
To test the hypothesis that PKCε is a common pathway in ischemic preconditioning (IPC), resveratrol, and AMPK neuroprotection, rat neuronal-astrocyte cortical cultures were preconditioned by exposure for 1 hour to either oxygen-glucose deprivation (OGD) to induce IPC, resveratrol (25 μM), or the AMPK activator AICAR (0.5 mM), with or without the selective Tat-conjugated PKCε inhibitor, εV1–2 (100 nM)
Summary
Aberrant energy metabolism following cerebral ischemia/reperfusion induces mitochondrial impairments such as reduced respiration, free radical generation, and release of proapoptotic factors[1,2]. Our laboratory has previously shown that protein kinase C epsilon (PKCε), a serine/threonine kinase isoform of the PKC family, confers wide-scale mitochondrial protection and is an important signaling pathway in the induction of neuroprotection against otherwise lethal ischemic injury[4,5,6,7]. PKCε activity influences a number of downstream signaling pathways that affect mitochondrial processes associated with ischemic neuroprotection. A major regulator of sirtuins is the enzyme, nicotinamide phosphoribosyl transferase (Nampt), which enhances sirtuin activity by increasing NAD+ levels[16]. Whether mitochondrial-localized sirtuins are regulated by PKCε-Nampt pathway or involved in ischemic neuroprotection has not been investigated. The main goal of this study was to investigate whether PKCε is involved in regulating SIRT3 and SIRT5 and whether these sirtuins are involved in PKCε-mediated neuroprotection following cerebral ischemia
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