PKNOX1 acts as a transcription factor of DHH and promotes the progression of stomach adenocarcinoma by regulating the Hedgehog signalling pathway

Abstract

Background: This study explored the effects and potential mechanism by which PBX/knotted 1 homeobox 1 (PKNOX1) may exacerbate stomach adenocarcinoma (STAD). Methods: For the in silico analysis, we examined TCGA-PKNOX1 expression using the UALCAN website, as well as its expression patterns in the GSE172032 and GSE174237 datasets, obtained from the GEO database. The associated patient survival curves, were analysed via the KMplot webtool. In vitro, we measured cell viability, proliferation, migration, and invasion using cell counting kit-8, colony formation, wound healing, and cell migration assays, respectively. Real time qPCR and western blotting assessed the mRNA and protein levels of PKNOX1, Snail, vimentin, N-cadherin, E-cadherin, desert hedgehog (DHH), cyclin D2, glioma-associated oncogene homolog 1, and smoothened. Gene Set Enrichment Analysis was performed using LinkedOmics webtools and the clusterProfiler package in R. Dual-luciferase reporter assay was used to examine the interactions of PKNOX1 with DHH, and of TEA domain transcription factor 4 (TEAD4) with PKNOX1. Results: PKNOX1 was highly expressed in STAD and linked to poor patient survival. Downregulation of PKNOX1 inhibited STAD cell viability, proliferation, migration, invasion, and epithelial-mesenchymal transition. Upregulation of TEAD4 promoted colony formation and migration, while these effects were reversed by PKNOX1 depletion. Furthermore, PKNOX1 regulated the activation of the hedgehog signalling pathway at the gene level, as we identified PKNOX1 to be a putative transcription factor for DHH that promotes its expression. Conclusion: Our results show that PKNOX1 acts as a candidate transcription factor for DHH and facilitates STAD development by regulating the hedgehog signalling pathway.