title: Ubiquitin Ligase, Fbw7, Targets CDX2 for Degradation via Two Phosphodegron Motifs in a GSK3β-Dependent Manner.
abstract: Drosophila caudal-related homeobox transcription factor 2 (CDX2) drives differentiation of the intestinal epithelium. Loss of CDX2 expression has been reported in several colorectal cancers and cancer cell lines with a potential inverse correlation between CDX2 levels and tumor stage. Ubiquitination of CDX2 leading to its downregulation has been implicated in several studies; however, the E3 ubiquitin ligases involved in CDX2 ubiquitination have largely remained unknown. Here, it is mechanistically determined that the E3 ubiquitin ligase Fbw7 promotes CDX2 ubiquitination and degradation through two phosphodegron motifs present within CDX2 in a GSK3β-dependent manner leading to its reduced expression and function in colon cancer cells. Fbw7, through its WD domain, interacted with CDX2 both in a heterologous HEK293T cell system and in colon cancer cells. GSK3β was also present in the same complex as determined by coimmunoprecipitation. Furthermore, overexpression of both Fbw7 or GSK3β down regulated endogenous CDX2 expression and function; however, both failed to inhibit endogenous CDX2 when either of them were depleted in colon cancer cells. Fbw7-mediated inhibition of CDX2 expression also led to reduced CDX2 transactivation and growth arrest of colon cancer cells. Both GSK3β and Fbw7 degraded mutant-CDX2 having either of the Cdc4-phosphodegron (CPD) motifs disrupted (CDX2-S60A or CDX-S281A), but were unable to degrade mutant-CDX2 having both CPDs disrupted (CDX2-S60,64,281A).
  
  
  
  Taken together, these findings demonstrate that Fbw7 negatively regulates CDX2 expression in a GSK3β-dependent manner through two CPDs present in CDX2. Mol Cancer Res; 14(11); 1097-109. ©2016 AACR.
authors.affiliation: Biochemistry Division, CSIR-Central Drug Research Institute (CSIR-CDRI), Lucknow, India.
authors.forename: Yogesh
authors.initials: Y
authors.lastname: Kumar
authors.affiliation: Biochemistry Division, CSIR-Central Drug Research Institute (CSIR-CDRI), Lucknow, India.
authors.forename: Nidhi
authors.initials: N
authors.lastname: Shukla
authors.affiliation: Biochemistry Division, CSIR-Central Drug Research Institute (CSIR-CDRI), Lucknow, India.
authors.forename: Gatha
authors.initials: G
authors.lastname: Thacker
authors.affiliation: Biochemistry Division, CSIR-Central Drug Research Institute (CSIR-CDRI), Lucknow, India.
authors.forename: Isha
authors.initials: I
authors.lastname: Kapoor
authors.affiliation: Biochemistry Division, CSIR-Central Drug Research Institute (CSIR-CDRI), Lucknow, India.
authors.forename: Savita
authors.initials: S
authors.lastname: Lochab
authors.affiliation: Department of Radiotherapy, King George's Medical University, Lucknow, India.
authors.forename: Madan Lal Brahma
authors.initials: ML
authors.lastname: Bhatt
authors.affiliation: Division of Endocrinology and Center for Research in Anabolic Skeletal Targets in Health and Illness (ASTHI), CSIR-Central Drug Research Institute (CSIR-CDRI), Lucknow, India.
authors.forename: Naibedya
authors.initials: N
authors.lastname: Chattopadhyay
authors.affiliation: Biochemistry Division, CSIR-Central Drug Research Institute (CSIR-CDRI), Lucknow, India.
authors.forename: Sabyasachi
authors.initials: S
authors.lastname: Sanyal
authors.affiliation: Biochemistry Division, CSIR-Central Drug Research Institute (CSIR-CDRI), Lucknow, India. arun3vedi@cdri.res.in.
authors.forename: Arun Kumar
authors.initials: AK
authors.lastname: Trivedi
mesh_terms: D020816:Amino Acid Motifs; D000071616:CDX2 Transcription Factor / Q000737:chemistry / Q000378:metabolism*; D018938:Caco-2 Cells; D018797:Cell Cycle Proteins / Q000737:chemistry / Q000378:metabolism*; D045744:Cell Line, Tumor; D003110:Colonic Neoplasms / Q000378:metabolism*; D044783:F-Box Proteins / Q000737:chemistry / Q000378:metabolism*; D000073979:F-Box-WD Repeat-Containing Protein 7; D000071679:Glycogen Synthase Kinase 3 beta / Q000378:metabolism*; D045325:HCT116 Cells; D057809:HEK293 Cells; D019073:HT29 Cells; D006801:Humans; D010766:Phosphorylation; D000072417:Protein Domains; D015398:Signal Transduction; D044767:Ubiquitin-Protein Ligases / Q000737:chemistry / Q000378:metabolism*; D054875:Ubiquitination
chemical_list: D000071616:CDX2 Transcription Factor; C491270:CDX2 protein, human; D018797:Cell Cycle Proteins; D044783:F-Box Proteins; D000073979:F-Box-WD Repeat-Containing Protein 7; C513273:FBXW7 protein, human; D044767:Ubiquitin-Protein Ligases; C000605506:GSK3B protein, human; D000071679:Glycogen Synthase Kinase 3 beta