E3 ubiquitin ligase Fbw7 negatively regulates granulocytic differentiation by targeting G-CSFR for degradation

• Published in: Biochimica et biophysica acta 2013-12, Vol.1833 (12), p.2639-2652
• Main Authors: Lochab, Savita, Pal, Pooja, Kapoor, Isha, Kanaujiya, Jitendra Kumar, Sanyal, Sabyasachi, Behre, Gerhard, Trivedi, Arun Kumar
• Format: Article
• Language: English
• Subjects: Animals; Cell Cycle Proteins - metabolism; Cell Differentiation; Cell Line; F-Box Proteins - metabolism; F-Box-WD Repeat-Containing Protein 7; Fbw7; G-CSFR; Gene Knockdown Techniques; Glycogen Synthase Kinase 3 - metabolism; Glycogen Synthase Kinase 3 beta; Granulocytes - cytology; Granulocytes - metabolism; GSK3β; Humans; Kinetics; Mice; Mutant Proteins - metabolism; Proteasome Endopeptidase Complex - metabolism; Protein Binding; Proteolysis; Receptors, Granulocyte Colony-Stimulating Factor - metabolism; STAT3 Transcription Factor - metabolism; Ubiquitin - metabolism; Ubiquitin-Protein Ligases - metabolism; Ubiquitin–proteasome degradation
• ISSN: 0167-4889; 0006-3002; 1879-2596
• DOI: 10.1016/j.bbamcr.2013.06.018

Tight control between activation and attenuation of granulocyte colony stimulating factor receptor (G-CSFR) signaling is essential to regulate survival, proliferation and differentiation of myeloid progenitor cells. Previous studies demonstrated negative regulation of G-CSFR through endosomal–lysosomal routing and ubiquitin–proteasome mediated degradation. However, very few E3 ubiquitin ligases are known to target G-CSFR for ubiquitin–proteasome pathway. Here we identified F-box and WD repeat domain-containing 7 (Fbw7), a substrate recognizing component of Skp–Cullin–F box (SCF) E3 ubiquitin Ligase physically associates with G-CSFR and promotes its ubiquitin-mediated proteasomal degradation. Our data shows that Fbw7 also interacts with and degrades G-CSFR-T718 (a truncated mutant of G-CSFR found in severe congenital neutropenia/acute myeloid leukemia (SCN/AML patients)) though at a quite slower rate compared to G-CSFR. We further show that glycogen synthase kinase 3 beta (GSK3β), like Fbw7 also targets G-CSFR and G-CSFR-T718 for degradation; however, Fbw7 and GSK3β are interdependent in targeting G-CSFR/G-CSFR-T718 for degradation because they are unable to degrade G-CSFR individually when either of them is knocked down. We further show that Fbw7 mediated downregulation of G-CSFR inhibits signal transducer and activator of transcription 3 (STAT3) phosphorylation which is required for G-CSF dependent granulocytic differentiation. In addition, our data also shows that inhibition of Fbw7 restores G-CSFR signaling leading to enhanced STAT3 activity resulting in massive granulocytic differentiation. These data indicate that Fbw7 together with GSK3β negatively regulates G-CSFR expression and its downstream signaling. •The E3 ubiquitin ligase Fbw7 targets G-CSFR for degradation.•Degradation of truncated G-CSFR-T718 by Fbw7 is quite slower than G-CSFR.•GSK3β and Fbw7 are interdependent in targeting both forms of G-CSFR for degradation.•Inhibition of Fbw7 restores G-CSFR signaling leading to enhanced differentiation.