Dickkopf-related protein 1/cytoskeleton-associated protein 4 signaling activation by Helicobacter pylori-induced activator protein-1 promotes gastric tumorigenesis via the PI3K/AKT/mTOR pathway

Abstract

Gastric cancer (GC) is a common malignant tumor with high incidence and mortality rates globally, especially in East Asian countries. Helicobacter pylori (H. pylori) infection is a significant and independent risk factor for GC. However, its underlying mechanism of action is not fully understood. Dickkopf-related protein (DKK) 1 is a Wnt signaling antagonist, and cytoskeleton-associated protein (CKAP) 4 is a newly identified DKK1 receptor. Recent studies found that the binding of DKK1 to CAKP4 mediated the procancer signaling of DKK1 inde-pendent of Wnt signaling. We hypothesize that H. pylori-induced activation of DKK1/CKAP4 signaling contributes to the initiation and progression of GC. To investigate the interaction of H. pylori infection, DKK1 and CAKP4 in GC, as well as the underlying molecular mechanisms. RNA sequencing was used to identify differentially expressed genes (DEGs) between H. pylori-infected and uninfected primary GC cells. Gain- and loss-of-function experiments were performed to verify the H. pylori-induced upregulation of activator protein-1 (AP-1) in GC cells. A dual-luciferase reporter assay and co-immunoprecipitation were used to determine the binding of AP-1 to the DKK1 promoter and DKK1 to CKAP4. Western blotting and immunohistochemistry detected the expression of DKK1, CKAP4, and phos-phatidylinositol 3-kinase (PI3K) pathway-related proteins in GC cells and tissues. Functional experiments and tumorigenicity in nude mice detected malignant behavior of GC cells in vitro and in vivo. We identified 32 DEGs between primary GC cells with and without H. pylori infection, including JUN, fos-like antigen-1 (FOSL1), and DKK1, and confirmed that the three proteins and CKAP4 were highly expressed in H. pylori-infected GC cells, H. pylori-infected gerbil gastric tissues, and human GC tissues. JUN and FOSL1 form AP-1 to transcriptionally activate DKK1 expression by binding to the DKK1 promoter. Activated DKK1 bound to CKAP4, but not the most common Wnt coreceptor low-density lipoprotein receptor-related protein 5/6, to promote GC cell growth, colony formation, migration, invasion, and xenograft tumor growth in nude mice. All these effects were driven by activation of the PI3K/AKT/mammalian target of rapamycin (mTOR) pathway. Targeting the PI3K signaling pathway by LY294002 inhibited DKK1-mediated CKAP4/PI3K signaling activity and the malignant behavior of GC cells. H. pylori induces JUN and FOSL1 expression to form AP-1, which transcriptionally activates DKK1. Binding of DKK1 to KAKP4 contributes to gastric tumorigenesis via the PI3K/AKT/mTOR pathway.