Hypermethylation of SHISA3 Promotes Nasopharyngeal Carcinoma Metastasis by Reducing SGSM1 Stability

Abstract

Altered DNA methylation is a key feature of cancer, and aberrant methylation is important in nasopharyngeal carcinoma (NPC) development. However, the methylation mechanisms underlying metastasis of NPC remain unclear. Analyzing data from public databases and conducting our own experiments, we report here that promoter hypermethylation of SHISA3 is common and contributes to the downregulation of this gene in many types of tumors, including NPC. SHISA3 suppressed NPC cell invasion and metastasis in vitro and in vivo by impeding the E3 ubiquitin ligase tripartite motif containing 21 (TRIM21)-mediated ubiquitination and degradation small G protein signaling modulator 1 (SGSM1) and by inhibiting the MAPK pathway activation. Silencing SGSM1 abrogated the inhibitory effect of SHISA3 on NPC cell migration and invasion. This newly identified SHISA3-TRIM21-SGSM1 axis could be a novel therapeutic target in NPC.Significance: These findings highlight the mechanism by which a newly identified tumor suppressor SHISA3 suppresses invasion and metastasis of nasopharyngeal carcinoma. Migration and invasion assaysCells (5 Â 10 4 or 1 Â 10 5 ) suspended in 200 mL of serum-free medium were plated into transwell chambers (8-mm pores, Corning) precoated with (migration assay) or without (invasion assay) Matrigel (BD Biosciences) cultured at 37 C for 12 hours or 24 hours. The cells attached to the bottom surface of the insert filter were fixed with methyl alcohol, stained with crystal violet, and counted in 10 random fields of view per well. Cell viability assayCells (1 Â 10 3 ) were seeded into 96-well plates and incubated for indicated time periods (1, 2, 3, 4, and 5 days). On the indicated days, CCK-8 (10 mL; Dojindo) were added per well following incubation for 2 hours, and the 450 nm absorbance of each well was read on a spectrophotometer. Colony formation assayCells (0.3 Â 10 3 ) were seeded into 6-well plates and cultured for 1 week (CNE2 cells) or 2 weeks (SUNE1 cells). The colonies were quantified after fixation with methyl alcohol and stained with crystal violet.

Hypermethylation of UCHL1 Promotes Metastasis of Nasopharyngeal Carcinoma by Suppressing Degradation of Cortactin (CTTN)

“…Methylation modification plays an important role in NPC metastasis, but the mechanisms remain poorly understood. We have previously reported that hypermethylation of the HOPX, RAB37, and SHISA3 genes might serve as molecular biomarkers to predict NPC metastasis [17,26,27]. Therefore, the mechanisms and significance of methylation regulation in NPC are worthy to be study in greater detail.…”

Multifaceted Roles of DNA Methylation in Neoplastic Transformation, from Tumor Suppressors to EMT and Metastasis

“…When SHISA3 is silenced, the increased SGSM1 polyubiquitylation and degradation abrogates one of the brakes impinging on the MAPK pathway. Accordingly, SHISA3 is downregulated in various tumor types, including nasopharyngeal carcinoma (NPC) in which the SHISA3 promoter hypermethylation correlates with the SHISA3 ability to suppress the NPC in vitro invasion and in vivo lymph node metastasis [96].…”

Melatonin reverses nasopharyngeal carcinoma cisplatin chemoresistance by inhibiting the Wnt/β-catenin signaling pathway

“…Total RNA was extracted using TRIzol regent (Life Technology, Carlsbad, USA) and converted to cDNA using a M-MLV reverse transcriptase kit (Promega). Quantitative PCR was performed and analyzed as described previously [41]. The primer sequences are shown in Supplementary Table 2.…”