N-myc downstream-regulated gene 2 promotes the protein stability of estrogen receptor beta via inhibition of ubiquitin-protein ligase E3A to suppress colorectal cancer.

Abstract

N-myc downstream-regulated gene 2 (NDRG2) and estrogen receptor beta (ERβ) both play key roles in cellular differentiation in colorectal cancer (CRC). Previous studies have demonstrated that ERβ co-locates with and directly transactivates NDRG2. However, the effect of NDRG2 on ERβ and its underlying mechanism remain largely unknown. Our aim of the study is to explore the effect of NDRG2 on ERβ and their contributions to progression of CRC. The Cancer Genome Atlas (TCGA) database was first utilized to validate the clinical significance of ERβ and NDRG2 in CRC. MTT and scratch migration assays were carried out to verify the role of ERβ and NDRG2 in CRC cells. Western blotting and polymerase chain reaction were performed to analyze the effect of NDRG2 on ERβ, and an immunoprecipitation assay was conducted to explore the protein-protein interaction. ERβ and NDRG2 were both found to be significantly down-regulated in tumor tissues from the TCGA-CRC database. NDRG2 was also observed to enhance the protein stability of ERβ while could not change messenger RNA (mRNA) level of ESR2 (encoding ERβ). A positive relationship was found to exist between the two proteins in CRC cells, with NDRG2 prolonging the half-life of ERβ and improving its nuclear translocation. Through detecting expression of ERβ downstream genes (such as TP53 and JNK) and performing related function experiment, we demonstrated that NDRG2 could promote transcriptional activation of ERβ target genes and enhance the function of tumor suppressors when the ERβ agonist diarylpropionitrile (DPN). The immunoprecipitation assay showed that NDRG2 could affect the complex components of ubiquitin-protein ligase E3A (UBE3A, known as E6AP) and ERβ, reducing the ubiquitin-mediated proteasome degradation of ERβ. In the current study, we found that NDRG2 could bind with UBE3A to hinder the binding of UBE3A with ERβ. Moreover, a positive feedback loop was discovered between NDRG2 and ERβ, which provides a novel insight and therapeutic target for CRC.