Muscarinic receptor agonist-induced βPix binding to β-catenin promotes colon neoplasia

Abstract

Abstract M 3 muscarinic receptors (M 3 R) modulate β-catenin signaling and colon neoplasia. CDC42/RAC guanine nucleotide exchange factor, βPix, binds to β-catenin in colon cancer cells, augmenting β-catenin transcriptional activity. Using in silico, in vitro, and in vivo approaches, we explored whether these actions are regulated by M 3 R. At the invasive fronts of murine and human colon cancers, we detected co-localized nuclear expression of βPix and β-catenin in stem cells overexpressing M 3 R. Using immunohistochemistry, immunoprecipitation, proximity ligand, and fluorescent cell sorting assays in human tissues and established and primary human colon cancer cell cultures, we detected time-dependent M 3 R agonist-induced cytoplasmic and nuclear association of βPix with β-catenin. βPix knockdown attenuated M 3 R agonist-induced human colon cancer cell proliferation, migration, invasion, and expression of PTGS2 , the gene encoding cyclooxygenase-2, a key player in colon neoplasia. Overexpressing βPix dose-dependently augmented β-catenin binding to the transcription factor TCF4. In a murine model of sporadic colon cancer, advanced neoplasia was attenuated in conditional knockout mice with intestinal epithelial cell deficiency of βPix. Expression levels of β-catenin target genes and proteins relevant to colon neoplasia, including c-Myc and Ptgs2, were reduced in colon tumors from βPix-deficient conditional knockout mice. Targeting the M 3 R/βPix/β-catenin axis may have therapeutic potential.