DDX19A promotes gastric cancer cell proliferation and migration by activating the PI3K/AKT pathway

Abstract

Background: DEAD-box RNA helicase 19 A (DDX19A) is overexpressed in cervical squamous cell carcinoma. However, its role in gastric cancer remains unclear. The present study aimed to explore the role and underlying mechanism of DDX19A in the development of gastric cancer.

Methods: The expression of DDX19A in gastric cancer and paracancerous tissues was evaluated through quantitative polymerase chain reaction, western blotting, and immunohistochemical staining. The biological functions of DDX19A in gastric cancer were determined using CCK8, plate colony-forming, and Transwell migration assays. The specific mechanism of DDX19A in gastric cancer cells was studied using western blotting, RNA-binding protein immunoprecipitation, mRNA half-life detection, and nuclear and cytoplasmic RNA isolation.

Results: DDX19A was highly expressed in gastric cancer and positively associated with malignant clinicopathological features and poor prognosis. Additionally, DDX19A promoted gastric cancer cell proliferation, migration, and epithelial-mesenchymal transition phenotypes. Mechanistically, DDX19A activated the PI3K/AKT pathway by upregulating phosphatidylinositol-3-kinase (PIK3CA) expression. Furthermore, DDX19A interacted with PIK3CA mRNA, stabilized it, and facilitated its export from the nucleus.

Conclusions: Our study reveals a novel mechanism whereby DDX19A promotes the proliferation and migration of gastric cancer cells by enhancing the stability and nuclear export of PIK3CA mRNA, thereby activating the PI3K/AKT pathway.

Keywords: DDX19A; Gastric cancer; Nuclear export; PI3K/AKT; PIK3CA.

Fig. 1

DDX19A expression is upregulated in GC, and high DDX19A expression is associated with poor prognosis. (A) DDX19A mRNA levels as detected using RT-qPCR in 24 paired fresh frozen GC tissues. (B) DDX19A expression level in 408 GC samples and 211 normal controls from the GEPIA database. (C) DDX19A protein levels as detected by western blotting in 24 paired fresh frozen GC tissues. (D) Immunohistochemical results of DDX19A (200×). (a) Normal gastric tissue; (b) high-medium differentiated GC tissue; (c) poorly differentiated GC tissue. (E) Kaplan–Meier curves for overall survival analysis by DDX19A expression in patients with GC. *p < 0.05, **p < 0.01. GC, gastric cancer; STAD, stomach adenocarcinoma

Fig. 2

DDX19A induces proliferation and migration of GC cells. (A) Analysis of DDX19A expression levels in GC cell lines and a normal gastric epithelium cell line (GES-1) by western blotting. (B) Silencing and overexpression efficiency assessed using western blotting and RT-qPCR analyses. (C) Growth curves of MGC-803 and AGS cells after transfection with DDX19A siRNA or DDX19A-overexpressing plasmid assessed using CCK8 assays. (D) Colony formation after depletion or overexpression of DDX19A assessed through plate cloning experiments. (E) GC cell migration assessed through Transwell migration assay. *p < 0.05, **p < 0.01, ***p < 0.001

Fig. 3

DDX19A knockdown inhibits tumor growth in vivo. (A) Efficiency of DDX19A knockdown. (B) Images of tumors removed from mice from the sh-DDX19A and sh-NC groups. (C) Images of nude mice from the sh-DDX19A and sh-NC groups. (D) Average tumor weight of xenografts. (E) Tumor growth curves.*p < 0.05, **p < 0.01, ***p < 0.001

Fig. 4

DDX19A regulates EMT markers in GC cells. *p < 0.05, **p < 0.01, ***p < 0.001

Fig. 5

DDX19A activates PI3K/AKT signaling pathway. *p < 0.05, **p < 0.01, ***p < 0.001, ^nsnot statistically significant

Fig. 6

PIK3CA inhibitor MLN1117 attenuates the promotion effect of DDX19A overexpression on GC cells. (A) Colony formation and (B) migration assays with AGS cells transfected with a DDX19A-overexpressing plasmid and incubated with the PIK3CA inhibitor MLN1117 (10 µM). (C) Expression of proteins involved in PI3K/AKT signaling in AGS cells transfected with a DDX19A-overexpressing plasmid and treated with DMSO or the PIK3CA inhibitor MLN1117 (10 µM). *p < 0.05, **p < 0.01, ***p < 0.001

Fig. 7

DDX19A promotes nuclear export and stabilizes PIK3CA mRNA. (A) Effect of DDX19A on PIK3CA mRNA expression. (B, C) Interaction between DDX19A and PIK3CA in AGS cells overexpressing Flag-DDX19A assessed using RIP assay, western blotting, and RT-qPCR. (D) Effects of DDX19A knockdown (left) and overexpression (right) on PIK3CA mRNA stability at the indicated time points measured using RT-qPCR. (E) PIK3CA mRNA levels in the nuclear and cytoplasmic fractions of DDX19A knockdown MGC-803 cells (left) and DDX19A overexpression AGS cells (right) assayed using RT-qPCR. *p < 0.05, **p < 0.01, ***p < 0.001, ^nsnot statistically significant