Regulation of 3-Phosphoinositide-dependent Protein Kinase-1 (PDK1) by Src Involves Tyrosine Phosphorylation of PDK1 and Src Homology 2 Domain Binding

• Published in: The Journal of biological chemistry 2008-01, Vol.283 (3), p.1480-1491
• Main Authors: Yang, Keum-Jin, Shin, Sanghee, Piao, Longzhen, Shin, Eulsoon, Li, Yuwen, Park, Kyeong Ah, Byun, Hee Sun, Won, Minho, Hong, Janghee, Kweon, Gi Ryang, Hur, Gang Min, Seok, Jeong Ho, Chun, Taehoon, Brazil, Derek P., Hemmings, Brian A., Park, Jongsun
• Format: Article
• Language: English
• Subjects: 3-Phosphoinositide-Dependent Protein Kinases; Cell Line; Disease; Enzyme Activation - drug effects; Enzyme Stability - drug effects; HSP90 Heat-Shock Proteins - antagonists & inhibitors; Humans; Leupeptins - pharmacology; Models, Biological; Mutant Proteins - metabolism; Phosphorylation - drug effects; Phosphotyrosine - metabolism; Proteasome Inhibitors; Protein Binding - drug effects; Protein Transport - drug effects; Protein-Serine-Threonine Kinases - metabolism; Proto-Oncogene Proteins c-crk - metabolism; Proto-Oncogene Proteins pp60(c-src) - chemistry; Proto-Oncogene Proteins pp60(c-src) - metabolism; Recombinant Fusion Proteins - metabolism; src Homology Domains; Subcellular Fractions - enzymology
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M706361200

3-Phosphoinositide-dependent protein kinase-1 (PDK1) appears to play a central regulatory role in many cell signalings between phosphoinositide-3 kinase and various intracellular serine/threonine kinases. In resting cells, PDK1 is known to be constitutively active and is further activated by tyrosine phosphorylation (Tyr9 and Tyr373/376) following the treatment of the cell with insulin or pervanadate. However, little is known about the mechanisms for this additional activation of PDK1. Here, we report that the SH2 domain of Src, Crk, and GAP recognized tyrosine-phosphorylated PDK1 in vitro. Destabilization of PDK1 induced by geldanamycin (a Hsp90 inhibitor) was partially blocked in HEK 293 cells expressing PDK1-Y9F. Co-expression of Hsp90 enhanced PDK1-Src complex formation and led to further increased PDK1 activity toward PKB and SGK. Immunohistochemical analysis with anti-phospho-Tyr9 antibodies showed that the level of Tyr9 phosphorylation was markedly increased in tumor samples compared with normal. Taken together, these data suggest that phosphorylation of PDK1 on Tyr9, distinct from Tyr373/376, is important for PDK1/Src complex formation, leading to PDK1 activation. Furthermore, Tyr9 phosphorylation is critical for the stabilization of both PDK1 and the PDK1/Src complex via Hsp90-mediated protection of PDK1 degradation.