AMPK-mediated phosphorylation of murine p27 at T197 promotes binding of 14-3-3 proteins and increases p27 stability.
The tuberous sclerosis complex 2 (Tsc2) gene product, tuberin, acts as a negative regulator of mTOR signaling, and loss of tuberin function leads to tumors of the brain, skin, kidney, heart, and lungs. Previous studies have shown that loss of tuberin function affects the stability and subcellular localization of the cyclin-dependent kinase inhibitor (CKI) p27, although the mechanism(s) by which tuberin modulates p27 stability has/have not been elucidated. Previous studies have also shown that AMP-activated protein kinase (AMPK), which functions in an energy-sensing pathway in the cell, becomes activated in the absence of tuberin. Here we show that in Tsc2-null tumors and cell lines, AMPK activation correlates with an increase in p27 levels, and inhibition of AMPK signaling decreases p27 levels in these cells. In addition, activation of AMPK led to phosphorylation of p27 at the conserved terminal threonine residue of murine p27 (T197) in both in vitro kinase assays and in cells. Phosphorylation of p27 at T197 led to increased interaction between p27 and 14-3-3 proteins and increased the protein stability of p27. Furthermore, activation of AMPK signaling promoted the interaction between p27 and 14-3-3 proteins and increased the stability of the p27 protein in a manner that was dependent on T197. These data identify a conserved mechanism for the regulation of p27 stability via phosphorylation at the terminal threonine (mT197/hT198) and binding of 14-3-3 proteins, which when AMPK is activated results in stabilization of the p27 protein.
Department of Carcinogenesis, The University of Texas M. D. Anderson Cancer Center-Research Division, Smithville, Texas 78957, USA.
John D
JD
Short
Ruhee
R
Dere
Kevin D
KD
Houston
Sheng-Li
SL
Cai
Jinhee
J
Kim
Judith M
JM
Bergeron
Jianjun
J
Shen
Jiyong
J
Liang
Mark T
MT
Bedford
Gordon B
GB
Mills
Cheryl Lyn
CL
Walker
D048948:14-3-3 Proteins / Q000378:metabolism*; D000263:Adenylate Kinase / Q000378:metabolism*; D000818:Animals; D002460:Cell Line; D050760:Cyclin-Dependent Kinase Inhibitor p27 / Q000378:metabolism*; D006801:Humans; D051379:Mice; D010766:Phosphorylation; D011485:Protein Binding; D015398:Signal Transduction
D048948:14-3-3 Proteins; D050760:Cyclin-Dependent Kinase Inhibitor p27; D000263:Adenylate Kinase