Regulation of Autophagy by Sphingosine Kinase 1 and Its Role in Cell Survival during Nutrient Starvation

Abstract

The sphingolipid ceramide induces macroautophagy (here called autophagy) and cell death with autophagic features in cancer cells. Here we show that overexpression of sphingosine kinase 1 (SK1), an enzyme responsible for the production of sphingosine 1-phosphate (S1P), in MCF-7 cells stimulates autophagy by increasing the formation of LC3-positive autophagosomes and the rate of proteolysis sensitive to the autophagy inhibitor 3-methyladenine. Autophagy was blocked in the presence of dimethylsphingosine, an inhibitor of SK activity, and in cells expressing a catalytically inactive form of SK1. In SK1 wt -overexpressing cells, however, autophagy was not sensitive to fumonisin B1, an inhibitor of ceramide synthase. In contrast to ceramide-induced autophagy, SK1(S1P)-induced autophagy is characterized by (i) the inhibition of mammalian target of rapamycin signaling independently of the Akt/protein kinase B signaling arm and (ii) the lack of robust accumulation of the autophagy protein Beclin 1. In addition, nutrient starvation induced both the stimulation of autophagy and SK activity. Knocking down the expression of the autophagy protein Atg7 or that of SK1 by siRNA abolished starvation-induced autophagy and increased cell death with apoptotic hallmarks. In conclusion, these results show that SK1(S1P)-induced autophagy protects cells from death with apoptotic features during nutrient starvation.

Sphingosine-1-phosphate phosphohydrolase-1 regulates ER stress-induced autophagy

“…Overexpression of sphingosine kinase 1 (SphK1), which increases S1P, has been shown to induce autophagy. 11 It was thus of interest to investigate the role in the autophagic process of the counteracting enzyme, SPP1, which decreases S1P levels. To this end, human breast adenocarcinoma MCF7 cells were transfected with specific siRNA targeted to SPP1, which markedly reduced its mRNA (480%) without altering the expression of other enzymes that regulate S1P levels, including S1P phosphatase-2 (SPP2), S1P lyase (Figure 1a), and SphK1 and SphK2 (data not shown).…”

“…Elevation of intracellular S1P by overexpression of SphK1, like downregulation of SPP1, has previously been reported to cause autophagy. 11 Although this was also characterized by a lack of robust upregulation of Beclin-1, inhibition of mTOR signaling was suggested to be involved. 11 On the other hand, autophagy was not altered in S1P lyase-null MEFs, despite increased S1P and concomitant elevation of both sphingosine and ceramide.…”

“…11 Although this was also characterized by a lack of robust upregulation of Beclin-1, inhibition of mTOR signaling was suggested to be involved. 11 On the other hand, autophagy was not altered in S1P lyase-null MEFs, despite increased S1P and concomitant elevation of both sphingosine and ceramide. 36 As the phosphatase and the lyase are important for different routes of S1P metabolism, the first for salvage of sphingolipids and the latter for generation of phosphatidylethanolamine, they likely have different functions in regulation of ER homeostasis and autophagy.…”

“…10 Surprisingly, however, increasing endogenous S1P levels by overexpression of sphingosine kinase 1 (SphK1), one of the isoenzymes that produces it, leads to inhibition of the mTOR pathway and autophagic cell survival that protects cells from apoptosis during nutrient starvation. 11 Thus, it has been proposed that ceramide-induced autophagy and S1P-induced autophagy may be mutually exclusive responses and that autophagy is the mechanism by which the sphingolipid rheostat controls cell fate decisions. 7 Examination of the enzymes that regulate the balance between ceramide and S1P may help to resolve these issues.…”

Aspartyl Protease-Mediated Cleavage of BAG6 Is Necessary for Autophagy and Fungal Resistance in Plants

“…Following these results, we further assessed whether chemical induction of autophagy is able to rescue the bag6 mutant phenotype. Leaves of bag6 mutant plants were preinfiltrated with known inducers of autophagy, including trehalose , rapamycin (Sarkar et al, 2009;Xiong and Sheen, 2014), DTT (Pérez-Martín et al, 2014;Zhou et al, 2014), tunicamycin (Liu et al, 2012;Munch et al, 2014), and tamoxifen (Lavieu et al, 2006;Brabec-Zaruba et al, 2007), prior to fungal inoculations. Tunicamycin treatment fully rescued the mutant phenotype to wild-type levels, while trehalose and tamoxifen treatments partially restored this phenotype (>50% of the wild type) (Supplemental Figure 10).…”