The ERK1/2 and mTORC1 Signaling Pathways Are Involved in the Muscarinic Acetylcholine Receptor-Mediated Proliferation of SNU-407 Colon Cancer Cells

ABSTRACT Muscarinic acetylcholine receptors (mAChRs) regulate diverse cellular functions, including cell growth and proliferation, via multiple signaling pathways. Previously, we showed that mAChRs stimulate the MEK1/2‐ERK1/2‐RSK pathway in SNU‐407 colon cancer cells and subsequently promote cell proliferation. In this study, we provide evidence that the PI3K‐Akt‐mTORC1‐S6K1 pathway is activated by mAChRs in SNU‐407 cells and that this pathway is associated with protein biosynthesis and cell proliferation. When the cells were treated with the cholinergic agonist carbachol, Akt was activated in a dose‐ and time‐dependent fashion. This carbachol effect was almost completely blocked by the PI3K inhibitor LY294002, implying that PI3K is responsible for the Akt activation. S6K1, a major downstream target of mTORC1, was also activated by carbachol in a temporal profile similar to that of the Akt activation. This carbachol‐stimulated S6K1 activation was abrogated by LY294002 or the mTORC1 inhibitor rapamycin, supporting the notion that mAChRs mediate S6K1 activation via the PI3K‐Akt‐mTORC1 pathway. We observed that global protein biosynthesis, monitored by puromycin incorporation, was strongly increased by carbachol in an atropine‐sensitive manner. Inhibition experiments indicated that the ERK1/2 and mTORC1 signaling pathways may be involved in carbachol‐stimulated global protein biosynthesis. We also found that treating SNU‐407 cells with LY294002 or rapamycin significantly suppressed carbachol‐stimulated cell proliferation. In the presence of the MEK1/2 inhibitor U0126, cell proliferation was further reduced by rapamycin treatment. Our data thus suggest that both the MEK1/2‐ERK1/2 and mTORC1 pathways play important roles in mAChR‐mediated cell proliferation in SNU‐407 colon cancer cells. J. Cell. Biochem. 117: 2854–2863, 2016. © 2016 Wiley Periodicals, Inc. Muscarinic acetylcholine receptors (mAChRs) activate the PI3K‐Akt‐mTORC1‐S6K1 pathway in SNU‐407 colon cancer cells. This pathway, together with the MEK1/2‐ERK1/2 pathway, is associated with mAChR‐mediated cell proliferation.