The Cross-Talk Between M2 Muscarinic Receptors and Notch-1/EGFR Pathways is Mediated by Mir-34a-5p in U87 Glioblastoma Cell Line

Glioblastoma (GBM) is the most aggressive human brain tumour. The high growth potential and decreased susceptibility to apoptosis of the glioma cells is mainly dependent on genetic amplifications or mutations of oncogenic or pro-apoptotic genes, respectively. We have previously shown that the activation of the M2 acetylcholine muscarinic receptors inhibited cell proliferation and induced apoptosis in two GBM cell lines and cancer stem cells. The aim of this study was to delve into the molecular mechanisms underlying the M2-mediated cell proliferation arrest. Exploiting U87MG and U251MG cell lines as model systems, we evaluated the ability of M2 receptors to interfere with Notch-1 and EGFR pathways, whose activation promotes GBM proliferation. We demonstrated that the activation of M2 receptors, by agonist treatment, counteracted Notch and EGFR signalling, through different regulatory cascades depending, at least in part, on p53 status. Only in U87MG cells, which mimic p53-wild type GBMs, M2 activation triggers a molecular circuitry, involving p53, Notch-1 and the tumour suppressor mir-34a-5p. This regulatory module negatively controls Notch-1, which affects cell proliferation mainly through the Notch-1/EGFR axis. Our data highlight, for the first time, a molecular circuitry that is deregulated in p53 wild type GBM, based on the cross-talk between M2 receptor and the Notch-1/EGFR pathways, mediated by mir-34a-5p.