In vivo role of the PIF-binding docking site of PDK1 defined by knock-in mutation

AUTOR(ES)

Collins, Barry J.

FONTE

Oxford University Press

RESUMO

PKB/Akt, S6K, SGK and RSK are mediators of responses triggered by insulin and growth factors and are activated following phosphorylation by 3-phosphoinositide-dependent protein kinase-1 (PDK1). To investigate the importance of a substrate-docking site in the kinase domain of PDK1 termed the ‘PIF-pocket’, we generated embryonic stem (ES) cells in which both copies of the PDK1 gene were altered by knock-in mutation to express a form of PDK1 retaining catalytic activity, in which the PIF-pocket site was disrupted. The knock-in ES cells were viable, mutant PDK1 was expressed at normal levels and insulin-like growth factor 1 induced normal activation of PKB and phosphorylation of the PKB substrates GSK3 and FKHR. In contrast, S6K, RSK and SGK were not activated, nor were physiological substrates of S6K and RSK phosphorylated. These experiments establish the importance of the PIF-pocket in governing the activation of S6K, RSK, SGK, but not PKB, in vivo. They also illustrate the power of knock-in technology to probe the physiological roles of docking interactions in regulating the specificity of signal transduction pathways.

Documentos Relacionados

• The PIF-binding pocket in PDK1 is essential for activation of S6K and SGK, but not PKB
• The in vivo role of PtdIns(3,4,5)P3 binding to PDK1 PH domain defined by knockin mutation
• Knock-In Mouse Models of Huntington's Disease
• High resolution crystal structure of the human PDK1 catalytic domain defines the regulatory phosphopeptide docking site
• A phosphoserine-regulated docking site in the protein kinase RSK2 that recruits and activates PDK1