Inhibition of the NEDD8 Conjugation Pathway by shRNA to UBA3, the Subunit of the NEDD8-Activating Enzyme, Suppresses the Growth of Melanoma Cells

Fang Cheng,Li Ruo-Hong,Hao Chen,Lei Zhang,Jian-Fang Sun,Yi Liu

doi:10.7314/apjcp.2012.13.1.057

Abstract

Neural precursor cell-expressed developmentally down-regulated 8 (NEDD8), a ubiquitin-like protein, mainly functions through covalent ligation to cullin proteins. Conjugation of NEDD8 with cullins can promote ubiquitination, which plays a critical role in the degradation of many proteins. UBA3 is the subunit of NEDD8- activating enzyme which is one of the keys for NEDD8 linkage to cullin proteins. Previous research showed NEDD8 conjugation to be up-regulated in highly proliferative cell lines. In the present study, up-regulated NEDD8 conjugation was observed in melanoma cell lines by Western blot analysis. After down-regulation with a RNAi to UBA3, proliferation of M14 was suppressed in vitro and in vivo. In conclusion, up-regulated NEDD8 conjugation may be involved in the development of melanoma. Interference in this pathway might offera promising method for melanoma therapy.

Highlights

Ubiquitin (Ub) is responsible for labeling proteins which are degraded by the 26s proteasome (Hershko, 2005; Hjerpe et al, 2008)
Neural precursor cell-expressed developmentally down-regulated 8 (NEDD8) conjugation in melanoma cells According to previous report (Kamitani et al, 1997; Huang et al, 2009), NEDD8 conjugation showed a series of bands in a ladder-like pattern in which the band of 90 kDa, consistent with the molecular mass of NEDD8cullins conjugation, was detected in all mammalian cells
Western blot was performed to analyze the amount of NEDD8 conjugation in melanoma cells A375, M14, MV3 and melanocytes

Summary

Introduction

Ubiquitin (Ub) is responsible for labeling proteins which are degraded by the 26s proteasome (Hershko, 2005; Hjerpe et al, 2008). Up-regulated NEDD8 conjugation was observed in melanoma cell lines by Western blot analysis. For it could promote the process of ubiquitination which is correlative closely with apoptosis, cell-cycle regulation, the conjugation of NEDD8 to cullins may be involved in the development of tumors. Western blot was performed to analyze the amount of NEDD8 conjugation in melanoma cells A375, M14, MV3 and melanocytes.

Results

Conclusion