The ubiquitin ligase CHIP modulates cellular behaviors of gastric cancer cells by regulating TRAF2

Cancer Cell International

Hao Chen¹  Jun Zhou²  Jingjing Xu²  Zhili Shan³  Hengying Yang³  Xiaojun Zhou³  Hanjue Dai⁴  Feng Guo⁵ 

[1] Department of Oncology, The Second People’s Hospital of Taizhou, 225500, Taizhou, China;grid.429222.d, Center for Clinical Laboratory, The First Affiliated Hospital of Soochow University, 215006, Suzhou, China;grid.429222.d, Department of General Surgery, The First Affiliated Hospital of Soochow University, 215006, Suzhou, China;grid.430455.3, Oncology center, Changzhou Second People’s Hospital Affiliated Nanjing Medical University, 213003, Changzhou, China;grid.440227.7, Department of Oncology, Nanjing Medical University Affiliated Suzhou Hospital, Baita West Road 16, 215001, Suzhou, China;

关键词: Gastric cancer;    CHIP;    TRAF2;    Cellular behaviors;    Prognostic factor;   

DOI  :  10.1186/s12935-019-0832-z

来源: publisher

【 摘 要 】

BackgroundCHIP is an E3 ubiquitin ligase that plays contrast roles in diverse human malignancies, depending on its targets. To date, the mechanisms underlying the function of CHIP in gastric cancer remains unclear. Here, we aim to further clarify the effects of CHIP on the development and progression of gastric cancer and explore its potential target.MethodsStably transfected CHIP-shRNA and TRAF2-shRNA AGS gastric cancer cell lines were established using Lipofectamine 2000. Cell growth was measured by an xCelligence real-time monitoring system and colony formation assay. Cell proliferation was detected using CCK-8, Ki-67, or CFSE assays. Apoptosis was detected by TUNEL assay or Annexin V/PI-staining followed by flow cytometric analysis. Cell cycle distribution was detected by PI-staining followed by flow cytometric analysis. Cell migration and invasion abilities were measured by a real-time xCelligence system, Transwell insert, and scratch assays. The expression of cell cycle-related proteins, apoptosis-related proteins, AKT, ERK, NF-κB signaling subunits, MMP2, MMP9, and Integrin β-1 were detected by Western blotting analysis. NF-κB DNA-binding capability was quantified using an ELISA-based NF-κB activity assay. Gastric cancer tissue microarray was analyzed to investigate the expression of both CHIP and TRAF2, and their clinical significance.ResultsThe CHIP-silencing in the AGS cells was oncogenic evidenced by the appearance of capable of anchorage-independent growth. The CHIP-silencing significantly enhanced the AGS cell proliferation capability likely due to the induced phosphorylation of ERK. The CHIP-silencing significantly inhibited apoptosis due to increased expression of Bcl-2. The CHIP-silencing promoted the AGS cell migration and invasion abilities, likely by regulating the expression of Integrin β-1. TRAF2 expression was markedly decreased in the CHIP-overexpressing cells at protein level, but not at mRNA level. The TRAF2-silencing markedly inhibited the proliferation ability of the AGS cells, the defected cell proliferation and enhanced apoptosis were involved in. The TRAF2-silencing also attenuated the cell migration and invasion capacities of the AGS cells. Furthermore, the expression of CHIP was downregulated while the expression of TRAF2 was upregulated in gastric cancer tissues. TRAF2 expression is independent prognostic factors of gastric cancer. The expression of CHIP and TRAF2 was negatively correlated in the gastric cancer tissue. Lower CHIP or higher TRAF2 was significantly linked to shorter overall survival in gastric cancer patients.ConclusionsTRAF2 influenced diverse aspects of cellular behavior of gastric cancer cells, including cell growth, migration, and invasion, which was in contrast to the functions of CHIP. TRAF2 could be considered as an independent prognostic factor in gastric cancer patients. It is possible that TRAF2 was a substrate of CHIP and CHIP regulated the TRAF2/NF-κB axis, which modulated diverse cellular behaviors in the AGS gastric cancer cells.

【 授权许可】

CC BY