Phosphoproteomics Analysis of Insulin-like Growth Factor-1 Signaling in Neuronal Cells

Phosphorylation is a key post-translational modification that can impact the function of a protein and the outcome of cell signaling pathways. Using phosphoproteomics to characterize the phosphorylation changes downstream of trophic factor signaling is important for better understanding the pleiotropic actions of this class of molecules. Insulin-like growth factor-1 (IGF-1) is a trophic factor that can influence cellular growth and differentiation, and IGF-1 signaling in the brain has been linked to cognitive processes. While its main signaling molecules are well characterized, we sought to perform a more in-depth and unbiased analysis of the IGF-1 phosphoproteome in neuronal cells. To obtain insight into the IGF-1 signaling pathway in neuronal cells, we performed a quantitative mass spectrometric phosphoproteomics analysis in rat pheochromocytoma (PC-12) cells. Our results illustrate a diverse phosphoproteome downstream of IGF-1, with insight into novel phosphoprotein sites, such as Plcβ3 (Ser¹¹⁰⁵), that likely influence IGF-1 signaling in neuronal cells. We also identify a robust upregulation of the Rho GTPase cycle and an activation of synaptogenesis signaling downstream of IGF-1. These results pave the way for more targeted studies on specific phosphoprotein sites, which will facilitate a better understanding of how these phosphorylation events impact IGF-1-related signaling outcomes in neuronal cells.