LncRNA GAS5 regulates redox balance and dysregulates the cell cycle and apoptosis in malignant melanoma cells

Long Chen,Qiao Zhang,Huixin Yang,Zihan Yi,Yuqian Li,Yuechun Zhu,Yingmin Kuang,Yuye Yang,Zhe Yang,Qiaoqiao Han,Lu Jiang

doi:10.1007/s00432-018-2820-4

Abstract

PurposeClinical outcomes for advanced malignant melanoma (MM) are often poor due to tumor invasiveness, metastasis, recurrence, and multidrug resistance.MethodsWe investigated whether apoptosis, cell cycle regulation, oxidative status, and redox balance were altered by changes in the expression of the long noncoding RNA, growth arrest-specific transcript 5 (GAS5), in MM cells.ResultsAnalysis of clinical samples from MM patients showed that the rate of reduced GAS5 expression, relative to that in adjacent noncancerous tissues, was significantly lower for tumors from patients with advanced disease (76.6%, P < 0.001), as evidenced by larger tumor size, higher TNM stage, and higher incidences of ulceration and metastasis (P < 0.001 for all). Cell culture experiments showed that siRNA-mediated knockdown of GAS5 increased the viability of A375-GAS5si cells. Flow cytometry and western blotting showed that GAS5 knockdown increased MM cell proliferation by inducing G1/S cell cycle progression through increases in Cyclin D1, CDK4, and p27 expression (P < 0.05 for all) and by inhibiting apoptosis through an increase in Bcl-2 expression (P < 0.001). Knockdown of GAS5 also increased levels of superoxide anion (P < 0.01), NADP+(P < 0.001), and oxidized glutathiones (P < 0.01) through increases in NOX4 expression (P < 0.001), G6PD expression (P < 0.01), and NOX activity (P < 0.05), and RNA co-immunoprecipitation showed that GAS5 induced these changes through a physical interaction between GAS5 and the G6PD protein.ConclusionsOur findings show GAS5 contributes to regulation of the apoptosis, cell cycle, homeostasis of reactive oxygen species, and redox balance in MM cells, and suggest that reduced GAS5 expression contributes to disease progression in MM patients.

Highlights

The incidence of malignant melanoma (MM) has increased in recent decades, especially in economically developed regions (Siegel et al 2017; Torre et al 2015)
We found that 76.6% of the MM tumor samples exhibited reduced growth arrest-specific transcript 5 (GAS5) expression, compared to that in adjacent noncancerous tissues (P < 0.001), and a significantly greater number of patients with Breslow thickness > 4 cm, TNM stage > II, ulceration, and lymph node metastasis exhibited reduced levels of GAS5 in their MM tissues (P < 0.001 for all)
Our findings show GAS5 contributes to regulation of the apoptosis, cell cycle, homeostasis of reactive oxygen species, and redox balance in MM cells, and suggest that reduced GAS5 expression contributes to disease progression in MM patients

Summary

Introduction

The incidence of malignant melanoma (MM) has increased in recent decades, especially in economically developed regions (Siegel et al 2017; Torre et al 2015). Despite the availability of a number of treatment options, including surgery, targeted chemotherapy, immunotherapy, and radiotherapy, the 5-year survival rate for stage-IV MM remains < 18% (Fruehauf and Trapp 2008; Maverakis et al 2015), and recently developed therapeutic agents for MM. Elevated levels of reactive oxygen species (ROS) occur in rapidly proliferating tumor cells as a result of increased ATP requirements, hypoxia, and oncogene-mediated signaling (Gill et al 2016; Szatrowski and Nathan 1991). Radiotherapy and some anticancer drugs stimulate apoptosis by increasing ROS levels beyond tolerable thresholds (Gill et al 2016; Gorrini et al 2013), but the mechanisms underlying the recalcitrance of MM to such treatments remain unclear (Cesi et al 2017; Gorrini et al 2013). A better understanding of the regulation of ROS production in MM cells and the identification of its links to apoptotic and non-apoptotic cell death mechanisms are needed

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