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. 2012 May 14;7(5):e36911. doi: 10.1371/journal.pone.0036911

Figure 2. MeCh-induced protein phosphorylation in diverse cellular contexts.

Figure 2

(A) Proportional diagrams illustrating the number of specifically enriched and identified phosphoproteins, basally occurring, and induced by MeCh stimulation (10 nM to 100 µM) of control or CMP-treated cells. (B) Selective protein immunoprecipitation (IP) and determination of MeCh-induced G protein-coupled receptor kinase interacting ArfGAP 1 (GIT1) serine and threonine phosphorylation, with antisera immunoblot (IB), in MeCh-stimulated control-state SH-SY5Y cells. The associated histogram (representing mean ± SEM data from three independent immunoprecipitations) indicates the extent of immunoprecipitated protein serine and threonine phosphorylation. Panels (C) and (D) indicate similar data from control-state cells for the MeCh-induced phosphorylation of G protein regulated inducer of neurite outgrowth 1 (GPRIN1) and p21 protein (Cdc42/Rac)-activated kinase 1 (PAK1). (E) Selective protein immunoprecipitation and determination of MeCh-induced microtubule-associated protein 2 (MAP2) serine and threonine phosphorylation, with antisera immunoblot, in MeCh-stimulated CMP-state SH-SY5Y cells. Panels (F) and (G) indicate similar data from CMP-state cells for the MeCh-induced phosphorylation of reticulon-4 (RTN4) and GRB2-associated binding protein 2 (GAB2). Statistical analysis was performed on three independent experiments using GraphPad Prism version 5.02 with a Student’s t-test (p<0.05, *; p<0.01, **; p<0.001, ***).