Neuroscience Letters 516 (2012) 105–109 Contents lists available at SciVerse ScienceDirect Neuroscience Letters jou rn al h om epage: www.elsevier.com/locate/neulet Insulin-like growth factor-1 induces the phosphorylation of PRAS40 via the PI3K/Akt signaling pathway in PC12 cells Haitao Wang a,1 , Qishan Zhang a,1 , Lang Zhang a , Peter J. Little b , Xiaochun Xie a , Qian Meng a , Yannan Ren a , Lihua Zhou d , Guoquan Gao d , Remi Quirion c , Wenhua Zheng a,∗ a State Key Laboratory of Ophthalmology, Zhongshan Ophthalmic Center and Neurophamacology, School of Pharmaceutical Sciences, Sun Yat-sen University, Guanzhou, China b Discipline of Pharmacy, School of Medical Sciences and Diabetes Complications Group, Health Innovations Research Institute, RMIT University, Bundoora, VIC 3083, Australia c Douglas Mental Health University Institute, McGill University, Montreal, Canada d Zhongshan School of Medicine, Sun Yat-Sen University, Guangzhou 510006, China a r t i c l e i n f o Article history: Received 12 July 2011 Received in revised form 13 February 2012 Accepted 26 March 2012 Keywords: IGF-1 PRAS40 PI3K/Akt MAPK Neuronal cells survival a b s t r a c t Insulin-like growth factor-1 (IGF-1) is a polypeptide tropic factor that plays an important role in the sur- vival and differentiation of both neuronal and non-neuronal cells. Numerous studies have demonstrated that IGF-1 promotes neuronal cell survival via the PI3K/Akt signaling pathway. Proline-rich Akt substrate of 40 kDa (PRAS40) is a recently discovered downstream target of Akt. However, the relationship between IGF-1 and PRAS40 is not known. In this study, we characterized the phosphorylation of PRAS40 induced by IGF-1 in PC12 cells and explored the signaling pathway responsible for the effect of IGF-1. IGF-1 induced the phosphorylation of Akt at Thr473 and PRAS40 at Thr246 in PC12 cells. The phosphorylation of Akt and PRAS40 induced by IGF-1 (100 ng/ml) was inhibited by the phosphatidylinositide 3-kinase (PI3K) specific inhibitor LY294002 (50 M), while no inhibitory effect was observed for a MAPK kinase pathway specific inhibitor PD98059 nor a p38 MAPK inhibitor PD169316, suggesting that the phosphorylation of PRAS40 induced by IGF-1 is mediated by the PI3K pathway in PC12 cells and primary cultured neu- rons. In further support this hypothesis, an Akt kinase specific inhibitor, Akt inhibitor VIII, attenuated IGF-1-induced phosphorylation of PRAS40 at the concentration that blocked the phosphorylation of Akt induced by IGF-1. Taken together, these data demonstrate that IGF-1 stimulates the phosphorylation of PRAS40 at Thr246 in neuronal cells and the effect of IGF-1 is mediated, at least in part, by the PI3K/Akt signaling pathway. © 2012 Elsevier Ireland Ltd. All rights reserved. 1. Introduction Insulin-like growth factor-1 (IGF-1) is one of the most impor- tant stimulators of cell growth [16], and a potent inhibitor of programmed cell death in both neuronal and non-neuronal cells [8,13]. IGF-1 binds to IGF-1 receptors causing receptor auto-phosphorylation and activation of receptor tyrosine kinase activity. Phosphorylation sites on the IGF-1 receptor bind various adapter molecules that contribute to the activation of down-stream signaling. IGF-1 stimulates the growth and survival of cells primar- ily through the activation of the phosphatidylinositide 3-kinase (PI3K)/Akt (PKB) and the Extracellular Signal-Regulated Kinase ∗ Corresponding author at: State Key Laboratory of Ophthalmology, Zhongshan Ophthalmic Center and Neuropharmacology, School of Pharmaceutical Sciences, Sun Yat-Sen University, East Waihuan Road, Higher Education Mega Center, Guangzhou 510006, Guangdong, China. Tel.: +86 20 39943027; fax: +86 20 39943027. E-mail address: whzheng123@gmail.com (W. Zheng). 1 Authors contributed equally to this work. (ERK) branch of the Mitogen Activated Protein Kinase (MAPK) sig- naling pathway [7,18]. Akt is one of the most important targets of PI3K and is an important serine/threonine protein kinase mediating cell sur- vival induced by many factors, including IGF-1. The activation of PI3K induced by growth factors and other stimuli causes increased levels of phosphatidylinositol 3,4-diphosphate (PIP2) and phosphatidylinositol-3,4,5-triphosphate (PIP3) in target cells. PIP3 recruits Akt to the plasma membrane where it is phosphorylated at positions Thr308 and Ser473 by PIP3-dependent kinase (PDK)-1 and PDK-2, respectively. The phosphorylation of Thr308 and Ser473 of Akt by PDK1 and PDK2 fully activates this kinase, which then phosphorylates its many substrates including glycogen-synthase kinase-3 (GSK-3), the Bcl-2 family member Bad [4], caspase-9 [2], nuclear factor-B [10,11] and the winged-helix family of transcrip- tion factors [6]. In this context a newly identified target of Akt is the proline-rich Akt substrate of 40 kDa (PRAS40), the activation of which promotes cell survival [9,17]. PRAS40, a small cytoplasmic protein, was recently identi- fied as an Akt substrate and a raptor-binding protein that is 0304-3940/$ – see front matter © 2012 Elsevier Ireland Ltd. All rights reserved. http://dx.doi.org/10.1016/j.neulet.2012.03.068