Caffeine induces apoptosis by enhancement of autophagy via pi3k-akt-mtor-p70s6k inhibition

BAsic ReseARch PAPeR

Autophagy 7:2, 176-187; February 2011; © 2011 Landes Bioscience

Caffeine induces apoptosis by enhancement of autophagy via PI3K/Akt/mTOR/p70S6K inhibition
shinji saiki,1 Yukiko sasazawa,2 Yoko imamichi,1 sumihiro Kawajiri,1 Takahiro Fujimaki,2 isei Tanida,3 hiroki Kobayashi,2 Fumiaki sato,4 shigeto sato,1 Kei-ichi ishikawa,1 Masaya imoto2 and Nobutaka hattori1,*
1Department of Neurology; Juntendo University school of Medicine; Bunkyo, Tokyo; 2Department of Biosciences and informatics; Faculty of science and Technology; Keio University; Kohoku, Yokohama; 3Department of Biochemistry and cell Biology; National institute of infectious Diseases; shinjyuku, Tokyo; 4 Research institute for Disease of Old Age; Juntendo University school of Medicine; Tokyo, JapanKey words: apoptosis, autophagy, PI3K/Akt/mTOR/p70S6K, ERK1/2, caffeine Abbreviations: PI3K, phosphoinositide-3 kinase; 4E-BP1, eukaryotic initiation factor 4-binding protein 1; ERK, extracellular signal-regulated kinase; mTOR, mammalian target of rapamycin; 3-MA, 3-methyladenine; MEFs, mouse embryonic fibroblasts; p70S6K, 70-kDa ribosomal protein S6 kinase; PI, propidium iodide; MPP+,1-methyl-4-phenylpyridinium

©2 1 Ln e Boc ne 01 a d s i i c. se D n t i r ue o o ds i t. tb
Introduction Caffeine has a diverse range of pharmacological effects.1 In addition to its various effects on the cell cycle and growth arrest, higher (4–10 mM) concentrations of caffeine can induce apoptosis in several cell lines, such as 10 mM caffeine in human neuroblastoma cells,2 4 mM caffeine in humanpancreatic adenocarcinoma cells3 and 5 mM caffeine in human A549 lung adenocarcinoma cells.4 Although caffeine has been reported to modulate cell cycle checkpoints and perturb molecular targets of the cell cycle, the exact mechanism of caffeine-induced apoptosis remains unclear.1 Autophagy is a key mechanism in various physiopathological processes, including tumorigenesis, development, cell death andsurvival.5,6 It has also been shown to have a complex relationship with apoptosis, especially in tumor cell lines.7 Several reports have shown that autophagy not only enhances caspase-dependent cell death, but is also required for it.8 In contrast, it has also been shown that autophagy plays an important role in promoting cell survival against apoptosis.7 Caffeine has been reported to inhibit somekinase activities, including various forms of phosphoinositol-3 kinase and mammalian target of rapamycin (mTOR).9,10 Recently, in food spoilage studies involving yeast, caffeine has been shown to induce a starvation response,11 which is a key regulator of autophagy causing its induction. However, the exact mechanism by which caffeine induces autophagy is still unknown. Here we report that higherconcentrations of caffeine enhance autophagic flux in a dose-dependent manner in various cell lines. Furthermore, we show that caffeine-induced autophagy is mainly dependent on PI3K/Akt/mTOR/p70S6 signaling and eventually results in apoptosis.

caffeine is one of the most frequently ingested neuroactive compounds. All known mechanisms of apoptosis induced by caffeine act through cell cyclemodulation or p53 induction. it is currently unknown whether caffeine-induced apoptosis is associated with other cell death mechanisms, such as autophagy. herein we show that caffeine increases both the levels of microtubule-associated protein 1 light chain 3-ii and the number of autophagosomes, through the use of western blotting, electron microscopy and immunocytochemistry techniques. Phosphorylatedp70 ribosomal protein s6 kinase (Thr389), s6 ribosomal protein (ser235/236), 4e-BP1 (Thr37/46) and Akt (ser473) were significantly decreased by caffeine. in contrast, eRK1/2 (Thr202/204) was increased by caffeine, suggesting an inhibition of the Akt/mTOR/p70s6K pathway and activation of the eRK1/2 pathway. Although insulin treatment phosphorylated Akt (ser473) and led to autophagy suppression,...