The Neddylation-Cullin 2-RBX1 E3 Ligase Axis Targets Tumor Suppressor RhoB for Degradation in Liver Cancer

Abstract

The neddylation-cullin-RING E3 ligase (CRL) pathway has recently been identified as a potential oncogenic event and attractive anticancer target; however, its underlying mechanisms have not been well elucidated. In this study, RhoB, a well known tumor suppressor, was identified and validated with an iTRAQ-based quantitative proteomic approach as a new target of this pathway in liver cancer cells. Specifically, cullin 2-RBX1 E3 ligase, which requires NEDD8 conjugation for its activation, interacted with RhoB and promoted its ubiquitination and degradation. In human liver cancer tissues, the neddylation-CRL pathway was overactivated and reversely correlated with RhoB levels. Moreover, RhoB accumulation upon inhibition of the neddylation-CRL pathway for anticancer therapy contributed to the induction of tumor suppressors p21 and p27, apoptosis, and growth suppression. Our findings highlight the degradation of RhoB via the neddylation-CRL pathway as an important molecular event that drives liver carcinogenesis and RhoB itself as a pivotal effector for anticancer therapy targeting this oncogenic pathway. Molecular & Cellular

Radiosensitization by the investigational NEDD8-activating enzyme inhibitor MLN4924 (pevonedistat) in hormone-resistant prostate cancer cells

“…The siRNA oligonucleotides are as follows: si-WEE1 (5′–GAGGCUGGAUGGAUGCAUUUU–3′) [19], si-p21 (5′–GUGGACAGCGAGCAGCUGAUU–3′) [20], si-p27 (5′–CCGACGAUUCUUCUACUCA–3′) [33], si-UBC12 (5′–GGGCUUCUACAAGAGUGGGAAGUUU–3′) [34] and si-Control (5′–UUCUCCGAACGUGUCACGUUU– 3′) [35]. The oligoes were purchased from GenePharma (Shanghai, China).…”

The CRL3BTBD9 E3 ubiquitin ligase complex targets TNFAIP1 for degradation to suppress cancer cell migration

“…Protein extraction, digestion, and label-free quantification in HepG2 cells HepG2 extracts were prepared according to our published methods. 29 Label-free quantification was carried out by the Beijing Proteome Research Center (Beijing, China).…”

“…In vivo ubiquitination assays TNFAIP1 ubiquitination analysis, including MLN4924 treatment, transfection with siRNA against ROC1 or Cul3, and transfection with sgRNA against BTBD9, was performed in liver and lung cancer cells following our published methods. 29 Transwell migration assays For cell migration assays, TNFAIP1-or BTBD9-knockdown cells were added to the upper chambers. DMEM containing 20% FBS was added to the lower chambers.…”

The Protein Neddylation Inhibitor MLN4924 Suppresses Patient-Derived Glioblastoma Cells via Inhibition of ERK and AKT Signaling

“…The most important substrate of neddylation is Cullin-RING E3 ligase (CRL), which is a multicomponent ubiquitin ligase that regulates the turnover of several proteins with major roles in cancer biology such as p21, p27, and NF-k-B inhibitor alpha [13]. Recently, increasing experimental evidence strongly suggest that the protein neddylation modification process is overexpressed in various human cancers [14][15][16][17]. These findings indicate that inhibition of protein neddylation can be utilized as an important strategy for the treatment of cancer.…”