PDK1 Protein Phosphorylation at Thr354 by Murine Protein Serine-Threonine Kinase 38 Contributes to Negative Regulation of PDK1 Protein Activity

摘要

Murine protein serine-threonine kinase 38 (MPK38) is a member of the AMP-activated protein kinase-related serine/threonine kinase family, which acts as cellular energy sensors. In this study, MPK38-induced PDK1 phosphorylation was examined to elucidate the biochemical mechanisms underlying phosphorylation-dependent regulation of 3-phosphoinositide-dependent protein kinase-1 (PDK1) activity. The results showed that MPK38 interacted with and inhibited PDK1 activity via Thr³⁵⁴ phosphorylation. MPK38-PDK1 complex formation was mediated by the amino-terminal catalytic kinase domain of MPK38 and the pleckstrin homology domain of PDK1. This activity was dependent on insulin, a PI3K/PDK1 stimulator, as well as various apoptotic stimuli, including TNF-α, H2O2, thapsigargin, and ionomycin. MPK38 inhibited PDK1 activity in a kinase-dependent manner and alleviated PDK1-mediated suppression of TGF-β (or ASK1) signaling, probably via the phosphorylation of PDK1 at Thr³⁵⁴. In addition, MPK38-mediated inhibition of PDK1 activity was accompanied by the modulation of PDK1 binding to its positive and negative regulators, serine/threonine kinase receptor-associated protein and 14-3-3, respectively. Together, these findings suggest an important role for MPK38-mediated phosphorylation of PDK1 in the negative regulation of PDK1 activity.