This Article Full Text Full Text (PDF) Data Supplement All Versions of this Article: 117/7/952    most recent CIRCULATIONAHA.107.744490v1 Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Request Permissions Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Xie, Z. Articles by Zou, M.-H. Search for Related Content PubMed PubMed Citation Articles by Xie, Z. Articles by Zou, M.-H. Pubmed/NCBI databases Compound via MeSH Substance via MeSH Hazardous Substances DB METFORMIN HYDROCHLORIDE Related Collections Type 1 diabetes Type 2 diabetes Other Vascular biology Related Article

(Circulation. 2008;117:952-962.)
© 2008 American Heart Association, Inc.

Molecular Cardiology

Phosphorylation of LKB1 at Serine 428 by Protein Kinase C- Is Required for Metformin-Enhanced Activation of the AMP-Activated Protein Kinase in Endothelial Cells

Zhonglin Xie, MD, PhD; Yunzhou Dong, PhD; Roland Scholz, BS; Dietbert Neumann, PhD; Ming-Hui Zou, MD, PhD

From the Division of Endocrinology and Diabetes, Department of Medicine, University of Oklahoma Health Sciences Center, Oklahoma City (Z.X., Y.D., M.Z.); and Institute of Cell Biology, ETH Zurich, Zurich, Switzerland (R.S., D.N.).

Correspondence to Ming-Hui Zou, MD, PhD, Division of Endocrinology and Diabetes, Department of Medicine, University of Oklahoma Health Sciences Center, BSEB 325, 941 Stanton L. Young Blvd, Oklahoma City, OK 73104. E-mail ming-hui-zou{at}ouhsc.edu

Received October 9, 2007; accepted December 14, 2007.

Background— Metformin, one of most commonly used antidiabetes drugs, is reported to exert its therapeutic effects by activating AMP-activated protein kinase (AMPK); however, the mechanism by which metformin activates AMPK is poorly defined. The objective of the present study was to determine how metformin activates AMPK in endothelial cells.

Methods and Results— Exposure of human umbilical vein endothelial cells or bovine aortic endothelial cells to metformin significantly increased AMPK activity and the phosphorylation of both AMPK at Thr172 and LKB1 at Ser428, an AMPK kinase, which was paralleled by increased activation of protein kinase C (PKC)-, as evidenced by increased activity, phosphorylation (Thr410/403), and nuclear translocation of PKC-. Consistently, either pharmacological or genetic inhibition of PKC- ablated metformin-enhanced phosphorylation of both AMPK-Thr172 and LKB1-Ser428, suggesting that PKC- might act as an upstream kinase for LKB1. Furthermore, adenoviral overexpression of LKB1 kinase-dead mutants abolished but LKB1 wild-type overexpression enhanced the effects of metformin on AMPK in bovine aortic endothelial cells. In addition, metformin increased the phosphorylation and nuclear export of LKB1 into the cytosols as well as the association of AMPK with LKB1 in bovine aortic endothelial cells. Similarly, overexpression of LKB1 wild-type but not LKB1 S428A mutants (serine replaced by alanine) restored the effects of metformin on AMPK in LKB1-deficient HeLa-S3 cells, suggesting that Ser428 phosphorylation of LKB1 is required for metformin-enhanced AMPK activation. Moreover, LKB1 S428A, like kinase-dead LKB1 D194A, abolished metformin-enhanced LKB1 translocation as well as the association of LKB1 with AMPK in HeLa-S3 cells. Finally, inhibition of PKC- abolished metformin-enhanced coimmunoprecipitation of LKB1 with both AMPK1 and AMPK2.

Conclusions— We conclude that PKC- phosphorylates LKB1 at Ser428, resulting in LKB1 nuclear export and hence AMPK activation.

---

 

CLINICAL PERSPECTIVE

Related Article:

Clinical Summaries
Circulation 2008 117: 857-859. [Extract] [Full Text]

This article has been cited by other articles:

				R. M. Memmott and P. A. Dennis LKB1 and Mammalian Target of Rapamycin As Predictive Factors for the Anticancer Efficacy of Metformin J. Clin. Oncol., December 1, 2009; 27(34): e226 - e226. [Full Text] [PDF]

				L. Zhou, S. S. Deepa, J. C. Etzler, J. Ryu, X. Mao, Q. Fang, D. D. Liu, J. M. Torres, W. Jia, J. D. Lechleiter, et al. Adiponectin Activates AMP-activated Protein Kinase in Muscle Cells via APPL1/LKB1-dependent and Phospholipase C/Ca2+/Ca2+/Calmodulin-dependent Protein Kinase Kinase-dependent Pathways J. Biol. Chem., August 14, 2009; 284(33): 22426 - 22435. [Abstract] [Full Text] [PDF]

				J. R. Ussher, J. S. Jaswal, C. S. Wagg, H. E. Armstrong, D. G. Lopaschuk, W. Keung, and G. D. Lopaschuk Role of the atypical protein kinase C{zeta} in regulation of 5'-AMP-activated protein kinase in cardiac and skeletal muscle Am J Physiol Endocrinol Metab, August 1, 2009; 297(2): E349 - E357. [Abstract] [Full Text] [PDF]

				B. Fisslthaler and I. Fleming Activation and Signaling by the AMP-Activated Protein Kinase in Endothelial Cells Circ. Res., July 17, 2009; 105(2): 114 - 127. [Abstract] [Full Text] [PDF]

				Z. Xie, Y. Dong, J. Zhang, R. Scholz, D. Neumann, and M.-H. Zou Identification of the Serine 307 of LKB1 as a Novel Phosphorylation Site Essential for Its Nucleocytoplasmic Transport and Endothelial Cell Angiogenesis Mol. Cell. Biol., July 1, 2009; 29(13): 3582 - 3596. [Abstract] [Full Text] [PDF]

				V. W. Dolinsky, A. Y.M. Chan, I. Robillard Frayne, P. E. Light, C. Des Rosiers, and J. R.B. Dyck Resveratrol Prevents the Prohypertrophic Effects of Oxidative Stress on LKB1 Circulation, March 31, 2009; 119(12): 1643 - 1652. [Abstract] [Full Text] [PDF]

				F. C. Denison, N. J. Hiscock, D. Carling, and A. Woods Characterization of an Alternative Splice Variant of LKB1 J. Biol. Chem., January 2, 2009; 284(1): 67 - 76. [Abstract] [Full Text] [PDF]

				S. Fogarty and D. G. Hardie C-terminal Phosphorylation of LKB1 Is Not Required for Regulation of AMP-activated Protein Kinase, BRSK1, BRSK2, or Cell Cycle Arrest J. Biol. Chem., January 2, 2009; 284(1): 77 - 84. [Abstract] [Full Text] [PDF]

				S. Roos, O. Lagerlof, M. Wennergren, T. L. Powell, and T. Jansson Regulation of amino acid transporters by glucose and growth factors in cultured primary human trophoblast cells is mediated by mTOR signaling Am J Physiol Cell Physiol, January 1, 2009; 297(3): C723 - C731. [Abstract] [Full Text] [PDF]

				S. Liangpunsakul, S.-E. Wou, Y. Zeng, R. A. Ross, H. N. Jayaram, and D. W. Crabb Effect of ethanol on hydrogen peroxide-induced AMPK phosphorylation Am J Physiol Gastrointest Liver Physiol, December 1, 2008; 295(6): G1173 - G1181. [Abstract] [Full Text] [PDF]

				F. Lan, J. M. Cacicedo, N. Ruderman, and Y. Ido SIRT1 Modulation of the Acetylation Status, Cytosolic Localization, and Activity of LKB1: POSSIBLE ROLE IN AMP-ACTIVATED PROTEIN KINASE ACTIVATION J. Biol. Chem., October 10, 2008; 283(41): 27628 - 27635. [Abstract] [Full Text] [PDF]

				J. Zhang, Z. Xie, Y. Dong, S. Wang, C. Liu, and M.-H. Zou Identification of Nitric Oxide as an Endogenous Activator of the AMP-activated Protein Kinase in Vascular Endothelial Cells J. Biol. Chem., October 10, 2008; 283(41): 27452 - 27461. [Abstract] [Full Text] [PDF]

				H. C. Choi, P. Song, Z. Xie, Y. Wu, J. Xu, M. Zhang, Y. Dong, S. Wang, K. Lau, and M.-H. Zou Reactive Nitrogen Species Is Required for the Activation of the AMP-activated Protein Kinase by Statin in Vivo J. Biol. Chem., July 18, 2008; 283(29): 20186 - 20197. [Abstract] [Full Text] [PDF]

				X. Hou, S. Xu, K. A. Maitland-Toolan, K. Sato, B. Jiang, Y. Ido, F. Lan, K. Walsh, M. Wierzbicki, T. J. Verbeuren, et al. SIRT1 Regulates Hepatocyte Lipid Metabolism through Activating AMP-activated Protein Kinase J. Biol. Chem., July 18, 2008; 283(29): 20015 - 20026. [Abstract] [Full Text] [PDF]