title: Destabilization of heterologous proteins mediated by the GSK3β phosphorylation domain of the β-catenin protein.
abstract: OBJECTIVE
  Wnt/β-catenin signaling plays important roles in development and cellular processes. The hallmark of canonical Wnt signaling activation is the stabilization of β-catenin protein in cytoplasm and/or nucleus. The stability of β-catenin is the key to its biological functions and is controlled by the phosphorylation of its amino-terminal degradation domain. Aberrant activation of β-catenin signaling has been implicated in the development of human cancers. It has been recently suggested that GSK3βmay play an essential role in regulating global protein turnover. Here, we investigate if the GSK3β phosphorylation site-containing degradation domain of β-catenin is sufficient to destabilize heterologous proteins.
  
  
  RESULTS
  We engineer chimeric proteins by fusing β-catenin degradation domain at the N- and/or C-termini of the enhanced green fluorescent protein (eGFP). In both transient and stable expression experiments, the chimeric GFP proteins exhibit a significantly decreased stability, which can be effectively antagonized by lithium and Wnt1. An activating mutation in the destruction domain significantly stabilizes the fusion protein. Furthermore, GSK3 inhibitor SB-216763 effectively increases the GFP signal of the fusion protein. Conversely, the inhibition of Wnt signaling with tankyrase inhibitor XAV939 results in a decrease in GFP signal of the fusion proteins, while these small molecules have no significant effects on the mutant destruction domain-GFP fusion protein.
  
  
  CONCLUSIONS
  Our findings strongly suggest that the β-catenin degradation domain may be sufficient to destabilize heterologous proteins in Wnt signaling-dependent manner. It is conceivable that the chimeric GFP proteins may be used as a functional reporter to measure the dynamic status of β-catenin signaling, and to identify potential anticancer drugs that target β-catenin signaling.
authors: The First Affiliated Hospital, Chongqing Medical University, Chongqing, China.
authors: Yuhan
authors: Y
authors: Kong
authors: Hongyu
authors: H
authors: Zhang
authors: Xian
authors: X
authors: Chen
authors: Wenwen
authors: W
authors: Zhang
authors: Chen
authors: C
authors: Zhao
authors: Ning
authors: N
authors: Wang
authors: Ningning
authors: N
authors: Wu
authors: Yunfeng
authors: Y
authors: He
authors: Guoxin
authors: G
authors: Nan
authors: Hongmei
authors: H
authors: Zhang
authors: Sheng
authors: S
authors: Wen
authors: Fang
authors: F
authors: Deng
authors: Zhan
authors: Z
authors: Liao
authors: Di
authors: D
authors: Wu
authors: Junhui
authors: J
authors: Zhang
authors: Xinyue
authors: X
authors: Qin
authors: Rex C
authors: RC
authors: Haydon
authors: Hue H
authors: HH
authors: Luu
authors: Tong-Chuan
authors: TC
authors: He
authors: Lan
authors: L
authors: Zhou
mesh_terms: D000595:Amino Acid Sequence; D002460:Cell Line; D004791:Enzyme Inhibitors / Q000494:pharmacology; D038362:Glycogen Synthase Kinase 3 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D000071679:Glycogen Synthase Kinase 3 beta; D006801:Humans; D007211:Indoles / Q000494:pharmacology; D008094:Lithium / Q000494:pharmacology; D008301:Maleimides / Q000494:pharmacology; D008969:Molecular Sequence Data; D010766:Phosphorylation; D055550:Protein Stability; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism*; D060449:Wnt Signaling Pathway / Q000187:drug effects; D051155:Wnt1 Protein / Q000235:genetics / Q000378:metabolism; D051176:beta Catenin / Q000235:genetics / Q000378:metabolism*
chemical_list: D004791:Enzyme Inhibitors; D007211:Indoles; D008301:Maleimides; D011993:Recombinant Fusion Proteins; C417521:SB 216763; D051155:Wnt1 Protein; C495246:Wnt1 protein, mouse; D051176:beta Catenin; D008094:Lithium; C000605506:GSK3B protein, human; D000071679:Glycogen Synthase Kinase 3 beta; C000605507:Gsk3b protein, mouse; D038362:Glycogen Synthase Kinase 3