Bioenergetic and autophagic control by Sirt3 in response to nutrient deprivation in mouse embryonic fibroblasts.
Bioenergetic and autophagic control by Sirt3 in response to nutrient deprivation in mouse embryonic fibroblasts.
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DOI：
10.1042/bj20130414
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发表时间：
2013-09-01
期刊：
影响因子：
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通讯作者：
Zhang J
中科院分区：
文献类型：
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作者：
Liang Q;Benavides GA;Vassilopoulos A;Gius D;Darley-Usmar V;Zhang J
Sirtuin 3（Sirt 3）是位于线粒体的NAD依赖性脱乙酰酶。Sirt 3在小鼠肌肉和肝脏中的表达通过饥饿而增加，这可以防止氧化应激和蛋白质损伤的饥饿依赖性增加。受损的蛋白质和细胞器依赖于自噬来清除，这对细胞存活至关重要，但Sirt 3的作用尚不清楚。为了检验这一点，我们使用Sirt 3敲除（KO）小鼠胚胎成纤维细胞，并发现在基础条件下，Sirt 3 KO细胞与野生型（WT）细胞相比表现出增加的自噬通量。响应于营养剥夺，WT和KO细胞均表现出增加的基础和ATP相关的线粒体呼吸，表明能量需求增加。两种细胞均表现出较低水平的磷酸化mTOR和较高的自噬通量，其中KO细胞表现出较低的最大线粒体呼吸和储备能力以及比WT细胞更高水平的自噬。在饥饿条件下，KO细胞比WT细胞表现出更高的磷酸化-JNK和磷酸化-c-Jun。然而，在Sirt 3 KO细胞中抑制JNK活性并不影响LC 3-I和LC 3-II水平，表明Sirt 3调节的自噬不依赖于JNK途径。与WT细胞相比，响应于营养剥夺，Sirt 3 KO细胞中的半胱天冬酶3活化和细胞死亡显著更高。通过氯喹抑制自噬加剧WT和Sirt 3 KO细胞中的细胞死亡，并且通过3-甲基腺嘌呤抑制自噬加剧Sirt 3 KO细胞中的细胞死亡。这些数据表明，营养缺乏诱导的自噬在细胞存活中起保护作用，Sirt 3降低了对增强自噬的需求并改善了细胞生物能量学。
Sirtuin 3 (Sirt3) is an NAD-dependent deacetylase localized to mitochondria. Sirt3 expression is increased in mouse muscle and liver by starvation, which could protect against the starvation-dependent increase in oxidative stress and protein damage. Damaged proteins and organelles depend on autophagy for removal and this is critical for cell survival but the role of Sirt3 is unclear. To examine this, we used Sirt3 knockout (KO) mouse embryonic fibroblast cells, and found that under basal conditions, Sirt3 KO cells exhibited increased autophagy flux compared to Wildtype (WT) cells. In response to nutrient deprivation, both WT and KO cells exhibited increased basal and ATP linked mitochondrial respiration, indicating an increased energy demand. Both cells exhibited lower levels of phosphorylated mTOR, and higher autophagy flux, with KO cells exhibiting lower maximal mitochondrial respiration and reserve capacity and higher levels of autophagy than WT cells. KO cells exhibit higher phospho-JNK and phospho-c-Jun than WT cells under starvation conditions. However, inhibition of JNK activity in Sirt3 KO cells did not affect LC3-I and LC3-II levels, indicating the Sirt3-regulated autophagy is independent of the JNK pathway. Caspase 3 activation and cell death are significantly higher in Sirt3 KO cells compared to WT cells in response to nutrient deprivation. Inhibition of autophagy by chloroquine, exacerbated cell death in both WT and Sirt3 KO cells, and by 3-methyadenine exacerbated cell death in Sirt3 KO cells. These data suggest that nutrient deprivation-induced autophagy plays a protective role in cell survival, and Sirt3 decreases the requirement for enhanced autophagy and improves cellular bioenergetics.