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. Author manuscript; available in PMC: 2021 May 1.
Published in final edited form as: J Thromb Haemost. 2020 Mar 5;18(5):1183–1196. doi: 10.1111/jth.14748

Figure 5: PDK1 regulates eIF4E activation and binding to an mRNA cap homologue in activated platelets.

Figure 5:

A, B Washed human platelets were untreated or pretreated with the PDK1 inhibitor BX-795 (1μM) for 5 min, followed by stimulation with 2-methylthio-ADP (2MeSADP) (50nM) under stirring condition for 15 min. The platelets were lysed and part of the lysate was separated and used as loading control before adding m7 GTP-Sepharose beads. The platelet lysate was incubated with m7 GTP-Sepharose beads. The beads were eluted to collect the eIF4E protein. Samples were analyzed for eIF4E binding using western blot analysis. This experiment is a representation of three experiments (**P < 0.05). C. Washed human platelets were left alone or pre-treated with the GSK3β inhibitor SB216763, followed by stimulation with 2MeSADP. Cell lysates were immunoprecipitated with 4EBP1 and samples were probed for total eIF4E using an anti-eIF4E mouse mAb. D. Washed human platelets were left alone or pre-treated with the GSK3β inhibitor SB216763 for 5 min, followed by stimulation with 2MeSADP (50 nM) for 5 min. Western blots were then probed for phosphorylated 4EBP1(T37) and eIF4E (S209). The western blots shown are representative of three independent experiments (* P< 0.05).