hnRNPA1 promotes the metastasis and proliferation of gastric cancer cells through WISP2-guided Wnt/β-catenin signaling pathway

Abstract

The main cause of gastric cancer (GC)-related death is due to malignant cell unregulated distant metastasis and proliferation. Heterogeneous nuclear ribonucleoprotein A1 (hnRNPA1) has been shown to play an important role in carcinogenesis and the development of metastasis in several tumors. However, its downstream regulatory mechanism in GC is not well defined. Our study aims to investigate the function and regulatory mechanism of hnRNPA1 in GC. We analyzed the differential expression of hnRNPA1 in gastric cancer and paired adjacent normal tissues in the TCGA database. Kaplan-Meier analysis was employed for survival assessment. The expressions of hnRNPA1 in GC cells were measured by qRT-PCR and Western blot. Transwell assay, CCK8 and colony formation assay were used to detect the effect of hnRNPA1 on the metastasis and proliferation ability of GC cells. Additionally, Western blotting was performed to examine the expression of proteins related to the Wnt/β-catenin signaling pathway as well as epithelial-mesenchymal transition (EMT), while further investigations were carried out to explore potential regulatory mechanisms. The results showed that hnRNPA1 was highly expressed differentially in GC over normal gastric tissue. Knocking down hnRNPA1 inhibited the metastasis and proliferation of human gastric cancer cells. Overexpression of hnRNPA1 significantly enhanced the metastatic potential and proliferative capacity of human GC cells. Further mechanism exploration revealed that knocking down hnRNPA1 inhibited the Wnt/β-catenin signaling pathway and WNT1 inducible signaling pathway protein-2 (WISP2), an activator of the Wnt/β-catenin signaling pathway. Whereas overexpression of hnRNPA1 had the opposite effects. Our results demonstrated that hnRNPA1 promoted metastasis and proliferation of GC cells by activating Wnt/β-catenin signaling pathway via WISP2. hnRNPA1 may serve as a potential biomarker and novel therapeutic targets for GC.

Keywords: AGS; Gastric cancer; HnRNPA1; Mechanism of cancer progression; WISP2; Wnt/β-catenin signaling pathway.

Fig. 1

Differential expression and prognostic analysis of hnRNPA1. A Comparative analysis of hnRNPA1 expression in 33 pairs of GC tissues and adjacent normal tissues, based on data from the TCGA database. B Analysis of hnRNPA1 expression in gastric cancer with and without distant metastasis using GSE26901 database. C Analysis of OS of GC patients through TCGA database using Kaplan-Meier Plotter. D Analysis of PPS of GC patients through TCGA database using Kaplan-Meier Plotter. E hnRNPA1 mRNA and protein table in human normal gastric mucosal epithelial cells (GES-1) and GC cells (AGS, HGC27, and MKN45). *P < 0.05; **P < 0.01; ***P < 0.001

Fig. 2

hnRNPA1 can promote the metastasis and proliferation of GC cells. A The levels of hnRNPA1 mRNA and protein in AGS cells subjected to transfection with sihnRNPA1 or si-NC were quantified using qRT-PCR and Western blot experiments. B The mRNA and protein expression of hnRNPA1 in AGS cells transfected with OE-hnRNPA1 or pcDNA3.1 was determined by qRT-PCR and western blot. C The migration and invasion abilities of AGS cells transfected with sihnRNPA1 or si-NC were assessed by Transwell migration and invasion assay. D The migration and invasion abilities of AGS cells transfected with OE-hnRNPA1 or pcDNA3.1 were determined by Transwell migration and invasion assay. E Proliferation ability in AGS cells of sihnRNPA1 or si-NC was evaluated by CCK-8 assay and colony formation assay. F Proliferation ability in AGS cells of OE-hnRNPA1 or pcDNA3.1 was determined by CCK-8 assay and colony formation assay. G The expression of EMT-related markers in AGS cells transfected with sihnRNPA1 or si-NC was detected by western blot assay. *P < 0.05; **P < 0.01; ***P < 0.001

Fig. 3

hnRNPA1 can regulate Wnt/β-catenin signaling pathway in GC cells through WISP2. A The expression of Wnt/β-catenin signaling pathway-related markers was determined by western blot in AGS cells transfected with sihnRNPA1 or si-NC. B The expression of Wnt/β-catenin signaling pathway-related markers was determined by western blot in AGS cells transfected with OE-hnRNPA1 or pcDNA3.1. C The mRNA and protein expression of WISP2 in AGS cells transfected with sihnRNPA1 or si-NC was determined by qRT-PCR and western blot. D The mRNA and protein expression of WISP2 in AGS cells transfected with OE-hnRNPA1 or pcDNA3.1 was determined by qRT-PCR and western blot. *P < 0.05; **P < 0.01; ***P < 0.001

Fig. 4

Knockdown of WISP2 inhibited the metastasis and proliferation of AGS cells. A Detection of WISP2 mRNA and protein levels in AGS cells subjected to transfection with siWISP2 or si-NC by qRT-PCR and western blot experiments. B Evaluation of the migratory and invasive capacities of AGS cells post-transfection with siWISP2 or si-NC, conducted via Transwell migration and invasion assays. C The proliferation ability in AGS cells transfected with siWISP2 or si-NC was detected by CCK-8 assay and colony formation assay. *P < 0.05; **P < 0.01; ***P < 0.001

Fig. 5

hnRNPA1 promotes GC cell metastasis and proliferation through WISP2. A The migration and invasion abilities of AGS cells co-transfected with sihnRNPA1 or si-NC, OE-WISP2 or pcDNA3.1 were detected by Transwell migration and invasion assay. B The proliferation ability of AGS cells co-transfected with sihnRNPA1 or si-NC, OE-WISP2 or pcDNA3.1 was determined by CCK-8 assay and colony formation assay. *P < 0.05; **P < 0.01; ***P < 0.001