TRAF6 inhibits colorectal cancer metastasis through regulating selective autophagic CTNNB1/ß-catenin degradation and is targeted for GSK3B/GSK3ß-mediated phosphorylation and degradation.

ABSTRACT

Aberrant CTNNB1 signaling is one of the fundamental processes in cancers, especially colorectal cancer (CRC). Here, we reported that TRAF6, an E3 ubiquitin ligase important for inflammatory signaling, inhibited epithelial-mesenchymal transition (EMT) and CRC metastasis through driving a selective autophagic CTNNB1 degradation machinery. Mechanistically, TRAF6 interacted with MAP1LC3B/LC3B through its LC3-interacting region 'YxxL' and catalyzed K63-linked polyubiquitination of LC3B. The K63-linked ubiquitination of LC3B promoted the formation of the LC3B-ATG7 complex and was critical to the subsequent recognition of CTNNB1 by LC3B for the selective autophagic degradation. However, TRAF6 was phosphorylated at Thr266 by GSK3B in most clinical CRC, which triggered K48-linked polyubiquitination and degradation of TRAF6 and thereby attenuated its inhibitory activity towards the autophagy-dependent CTNNB1 signaling. Clinically, decreased expression of TRAF6 was associated with elevated GSK3B protein levels and activity and reduced overall survival in CRC patients. Pharmacological inhibition of GSK3B activity stabilized the TRAF6 protein, promoted CTNNB1 degradation, and effectively suppressed EMT and CRC metastasis. Thus, targeting TRAF6 and its pathway may be meaningful for treating advanced CRC. Abbreviations AMBRA1 autophagy and beclin 1 regulator 1; AOM azoxymethane; ATG5 autophagy related 5; ATG7 autophagy related 7; Baf A1 bafilomycin A1; BECN1 beclin 1; CoIP co-immunoprecipitation; CQ chloroquine; CRC colorectal cancer; CTNNB1/ß-catenin catenin beta 1; DSS dextran sodium sulfate; EMT epithelial-mesenchymal transition; FBS fetal bovine serum; GFP green fluorescent protein; GSK3B/GSK3ß glycogen synthase kinase 3 beta; IgG Immunoglobulin G; IHC immunohistochemistry; LIR LC3-interacting region; MAP1LC3B/LC3B microtubule associated protein 1 light chain 3 beta; RFP red fluorescent protein; RT room temperature; shRNA short hairpin RNA; siRNA small interfering RNA; TRAF6 TNF receptor-associated factor 6; WT wild-type; ZEB1 zinc finger E-box binding homeobox 1.