Abstract
Epstein-Barr virus-associated gastric cancer (EBVaGC) accounts for about 10% of all gastric cancer cases and has unique pathological and molecular characteristics. EBV encodes a large number of microRNAs, which actively participate in the development of EBV-related tumors. Here, we report that EBV-miR-BART3-3p (BART3-3p) promotes gastric cancer cell growth in vitro and in vivo Moreover, BART3-3p inhibits the senescence of gastric cancer cells induced by an oncogene (RASG12V) or chemotherapy (irinotecan). LMP1 and EBNA3C encoded by EBV have also been reported to have antisenescence effects; however, in EBVaGC specimens, LMP1 expression is very low, and EBNA3C is not expressed. BART3-3p inhibits senescence of gastric cancer cells in a nude mouse model and inhibits the infiltration of natural killer cells and macrophages in tumor by altering the senescence-associated secretory phenotype (SASP). Mechanistically, BART3-3p directly targeted the tumor suppressor gene TP53 and caused down-regulation of p53's downstream target, p21. Analysis from clinical EBVaGC samples also showed a negative correlation between BART3-3p and TP53 expression. It is well known that mutant oncogene RASG12V or chemotherapeutic drugs can induce senescence, and here we show that both RASG12V and a chemotherapy drug also can induce BART3-3p expression in EBV-positive gastric cancer cells, forming a feedback loop that keeps the EBVaGC senescence at a low level. Our results suggest that, although TP53 is seldom mutated in EBVaGC, its expression is finely regulated such that EBV-encoded BART3-3p may play an important role by inhibiting the senescence of gastric cancer cells.
Highlights
Epstein–Barr virus–associated gastric cancer (EBVaGC) accounts for about 10% of all gastric cancer cases and has unique pathological and molecular characteristics
It is well known that mutant oncogene RASG12V or chemotherapeutic drugs can induce senescence, and here we show that both RASG12V and a chemotherapy drug can induce BART3-3p expression in EBV-positive gastric cancer cells, forming a feedback loop that keeps the EBVaGC senescence at a low level
TP53 is seldom mutated in EBVaGC, its expression is finely regulated such that EBV-encoded BART3-3p may play an important role by inhibiting the senescence of gastric cancer cells
Summary
To find the EBV BART microRNAs that may regulate p53, we searched all the BART microRNA seed sequences and found that EBV BART3-3p has several binding sites that can interact with TP53. Considering that the binding sites of BART3-3p and TP53 mRNA predicted by bioinformatics were located in the TP53 CDS region, we cotransfected BART3-3p mimics and TP53 expression vector GFP-p53, which lacked the TP53 3Ј-UTR and 5Ј-UTR, into SGC7901 and AGS cells and found that GFP-p53 was inhibited (Fig. 1B). The luciferase activity of the WT TP53 CDS but not the mutant CDS was significantly reduced by BART3-3p but not by negative control mimics (Fig. 1H) These results strongly suggest that BART3-3p directly binds to the TP53 CDS region and inhibits its transcription. The CCK8 assay showed the same function of BART3-3p inhibitor in cell proliferation in SGC7901 LV-b3-3p and AGS LV-b3-3p cells (Fig. 3F) These results suggest that BART3-3p promotes the proliferation of GC cells both in vitro and in vivo.
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