Investigation of G6PD as a potential substrate of CRBN

Abstract

Cereblon(CRBN), initially discovered as a gene related to autosomal recessive non-syndromic mental retardation, functions as a substrate receptor for Cullin 4-ring E3 ubiquitin ligase complex(known as CRL4CRBN). CRL4CRBN mediates ubiquitination of substrates and proteasomal degradation. Over past years, various substrates of CRBN have been identified and researched molecular mechanisms related to diverse biological functions. Additionally, CRBN recruits the neosubstrates mediated by immunomodulatory drugs(IMiDs) including thalidomide. Previously, the role of CRBN was revealed as a regulator of AMP-activated protein kinase(AMPK) involving in energy metabolism, inflammatory response and redox homeostasis. However, correlation of CRBN and glucose-6-phosphate dehydrogenase(G6PD), the housekeeping enzyme catalyzing the first reaction of pentose phosphate pathway(PPP), remains still unknown. G6PD functions in anti-oxidant system to protect the cells from oxidative damage by maintaining level of NADPH and glutathione. To investigate the correlation of G6PD and CRBN, the effect of CRBN in G6PD regulation was examined. Protein level of G6PD was higher as CRBN was deficient in liver and skeletal muscle tissue of mice and also in primary MEF cells. Contrastively, G6PD protein level was decreased by exogenous CRBN. However, mRNA level of G6PD was not affected by CRBN knockout, indicated to post-translational regulation of G6PD. I confirmed the direct binding of two proteins in endogenous level. Furthermore, it was verified that N-terminus of G6PD related to binding with CRBN using domain deletion mutants of G6PD. To explore the CRBN-dependent regulation of G6PD induced by high glucose, as previous reported, I inspected the effect of CRBN on G6PD level in different glucose condition. When CRBN was suppressed in cell, down-regulation of G6PD mitigated in high glucose condition. In conclusion, these results indicate that CRBN is involved in regulation of G6PD.