GSK-3β controls autophagy by modulating LKB1-AMPK pathway in prostate cancer cells

  • Prostate. 2016 Feb;76(2):172-83. doi: 10.1002/pros.23106.
Aijing Sun  1  2 ,  Changlin Li  2 ,  Ruibao Chen  2 ,  Yiling Huang  2  3 ,  Qi Chen  4 ,  Xiangjun Cui  5 ,  Huafeng Liu  6 ,  J Brantley Thrasher  2 ,  Benyi Li  2  3  5  6
Affiliations
  • 1. Department of Pathology, Shaoxing University School of Medicine, Shaoxing, China.
  • 2. Department of Urology, University of Kansas Medical Center, Kansas City, Kansas.
  • 3. Department of Pathology, China Three Gorges University College of Medicine, Yichang, China.
  • 4. Department of Pharmacology, Toxicology and Therapeutics, University of Kansas Medical Center, Kansas City, Kansas.
  • 5. Department of Clinical Immunology and Rheumatology, Yichang Renmin Hospital, China Three Gorges University, Yichang, China.
  • 6. Department of Internal Medicine and Kidney Institute, The Affiliated Hospital, Guangdong Medical College, Zhanjiang, China.
PMID: 26440826 DOI: 10.1002/pros.23106
Abstract

Background: Glycogen synthase kinase 3β (GSK3B, GSK-3β) is a multi-functional protein kinase involved in various cellular processes and its activity elevates after serum deprivation. We have shown that inhibition of GSK-3β activity triggered a profound autophagic response and subsequent necrotic cell death after serum deprivation in prostate Cancer cells. In this study, we dissected the mechanisms involved in GSK-3β inhibition-triggered Autophagy.

Methods: Prostate Cancer PC-3 and DU145 cells were used in the study. Multiple GSK-3β specific inhibitors were used including small chemicals TDZD8, Tideglusib, TWS119, and peptide L803-mts. Western blot assay coupled with phospho-specific antibodies were used in detecting signal pathway activation. ATP levels were assessed with ATPLite kit and HPLC methods. Autophagy response was determined by evaluating Microtubule-associated proteins 1A/1B light chain 3B (LC3B) processing and p62 protein stability in Western blot assays. Immunofluorescent microscopy was used to detect LKB1 translocation.

Results: Inhibition of GSK-3β activity resulted in a significant decline of cellular ATP production, leading to a significant increase of AMP/ATP ratio, a strong trigger of AMP-activated protein kinase (AMPK) activation in prostate Cancer PC-3 cells. In parallel with increased LC-3B biosynthesis and p62 protein reduction, the classical sign of Autophagy induction, AMPK was activated after inhibition of GSK-3β activity. Further analysis revealed that Liver kinase B1 (LKB1) but not Calcium/calmodulin-dependent protein kinase kinase β (CaMKKβ) is involved in AMPK activation and Autophagy induction triggered by GSK-3β inhibition. Meanwhile, GSK-3β inhibition promoted LKB1 translocation from nuclear to cytoplasmic compartment and enhanced LKB1 interaction with its regulatory partners Mouse protein-25 (MO25) and STE20-related adaptor (STRAD).

Conclusions: In conclusion, our data suggest that GSK-3β plays an important role in controlling Autophagy induction by modulating the activation of LKB1-AMPK pathway after serum deprivation.

Keywords
AMPK; GSK-3β; LKB1; autophagy; serum deprivation.
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