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

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.

AMPK/GSK3β/β‐catenin cascade‐triggered overexpression of CEMIP promotes migration and invasion in anoikis‐resistant prostate cancer cells by enhancing metabolic reprogramming

“…AMPK signaling inhibitor, dorsomorphin (compound C) 2HCl (Selleck Chemicals, Houston, TX, USA), was dissolved in sterile double-distilled water, whereas GSK3b inhibitor, TDZD-8 (Selleck Chemicals), and glycolysis inhibitor, 2-deoxy-D-glucose (2-DG; Selleck Chemicals), were dissolved in DMSO. Anoikisresistant PC-3 or DU145 cells were treated with 4 or 2 mM dorsomorphin, respectively, for 24 h (Supplemental Information 3), and anoikis-resistant PCa cells were treated with 10 mM TDZD-8 for 24 h or 10 mM 2-DG for 48 h (25,26). Isometric double-distilled water or DMSO were used as negative control (NC).…”

Clinically relevant GSK‑3β inhibitor 9‑ING‑41 is active as a single agent and in combination with other antitumor therapies in human renal cancer

“…An increase of intracellular glucose storage and induction of autophagy have been demonstrated in 9-ING-41-treated renal cancer cells (20). Inhibition of GSK-3 triggers an autophagic response in prostate (31), pancreatic (32) and renal (20) cancer. Pal et al (20) have demonstrated that GSK-3 inhibition by 9-ING-41 affects energy homeostasis and triggers a pro-survival autophagic response in renal cancer cells.…”

FAM48A mediates compensatory autophagy induced by proteasome impairment

“…Several reports have suggested that this compensatory autophagy is induced by a stress response elicited during proteasome impairment. Mitochondrial ROS production and depletion of amino acids during proteasome dysfunction cause the activation of autophagy through sensing via AMPK and mTORC1, respectively (Jiang et al, ; Min et al, ; Sun et al, ). Proteasome impairment also induces endoplasmic reticulum (ER) stress, which then triggers transcriptional up‐regulation of autophagy‐related genes (Brüning, Rahmeh, & Friese, ; Høyer‐Hansen & Jäättelä, ; Senft & Ronai, ).…”