Threonine 34 phosphorylation by phosphoinositide-dependent protein kinase 1 facilitates dissociation of Akt from the plasma membrane.
Akt is a key mediator of cell proliferation, survival and metabolism. After translocation to the membrane and phosphorylation at T308 and S473, the activated Akt dissociates from the plasma membrane to cytoplasm, which is an important step to phosphorylate its downstream targets. In addition to its central role in regulating the kinase activity, phosphorylation of T308 in the kinase loop has been reported to be necessary for this dissociation process. However, it is not clear whether the membrane detachment requires further mechanisms. In the present report, we demonstrate that membrane dissociation of Akt requires phosphoinositide-dependent protein kinase 1 (PDK1) which directly phosphorylates not only T308 but also T34 in the pleckstrin homology (PH) domain. Like T308, T34 was phosphorylated in a phosphatidylinositol 3,4,5-trisphosphate- and phosphatidylserine-dependent manner. Phosphorylation of T34 also occurred in cells following growth factor stimulation, concurrently with T308 phosphorylation. Moreover, when T34 was mutated to aspartic acid (T34D) to mimic its phosphorylation, Akt-membrane association assessed by surface plasmon resonance spectroscopy was significantly reduced. In cells, this mutation impaired the IGF-induced Akt membrane translocation and subsequent phosphorylation at T308 and S473. Taken together, our results demonstrate that T34 phosphorylation by PDK1 promotes the membrane dissociation of activated Akt for its downstream action through attenuating membrane binding affinity. This membrane dissociation mechanism offers a new insight for Akt activation process and provides a potential new target for controlling the Akt-dependent cellular processes.
Laboratory of Molecular Signaling, National Institute of Alcohol Abuse and Alcoholism, National Institutes of Health, Bethesda, MD 20892-9410, United States.
Bill X
BX
Huang
Laboratory of Molecular Signaling, National Institute of Alcohol Abuse and Alcoholism, National Institutes of Health, Bethesda, MD 20892-9410, United States.
Rachel
R
Lee
Laboratory of Molecular Signaling, National Institute of Alcohol Abuse and Alcoholism, National Institutes of Health, Bethesda, MD 20892-9410, United States.
Mohammed
M
Akbar
Laboratory of Molecular Signaling, National Institute of Alcohol Abuse and Alcoholism, National Institutes of Health, Bethesda, MD 20892-9410, United States. Electronic address: hykim@nih.gov.
Hee-Yong
HY
Kim
D000818:Animals; D045744:Cell Line, Tumor; D002462:Cell Membrane / Q000201:enzymology; D051379:Mice; D010766:Phosphorylation; D011485:Protein Binding; D011499:Protein Processing, Post-Translational; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D021381:Protein Transport; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism*; D000081382:Pyruvate Dehydrogenase Acetyl-Transferring Kinase; D013002:Somatomedins / Q000502:physiology; D013912:Threonine / Q000378:metabolism
C000631433:Pdk1 protein, mouse; D000081382:Pyruvate Dehydrogenase Acetyl-Transferring Kinase; D013002:Somatomedins; D013912:Threonine; D017346:Protein Serine-Threonine Kinases; D051057:Proto-Oncogene Proteins c-akt
Akt activation; Membrane translocation; PDK1; Protein kinase B; Threonine 34