title: Interacting post-muscarinic receptor signaling pathways potentiate matrix metalloproteinase-1 expression and invasion of human colon cancer cells.
abstract: M3 muscarinic receptor (M3R) expression is increased in colon cancer; M3R activation stimulates colon cancer cell invasion via cross-talk with epidermal growth factor receptors (EGFR), post-EGFR activation of mitogen-activated protein kinase (MAPK) extracellular signal-related kinase 1/2 (ERK1/2), and induction of matrix metalloproteinase-1 (MMP1) expression.MMP1expression is strongly associated with tumor metastasis and adverse outcomes. Here, we asked whether other MAPKs regulate M3R agonist-inducedMMP1expression. In addition to activating ERK1/2, we found that treating colon cancer cells with acetylcholine (ACh) stimulated robust time- and dose-dependent phosphorylation of p38 MAPK. Unlike ERK1/2 activation, ACh-induced p38 phosphorylation was EGFR-independent and blocked by inhibiting protein kinase C-α (PKC-α). Inhibiting activation of PKC-α, EGFR, ERK1/2, or p38-α/β alone attenuated, but did not abolish ACh-inducedMMP1expression, a finding that predicted potentiating interactions between these pathways. Indeed, ACh-inducedMMP1expression was abolished by incubating cells with either an EGFR or MEK/ERK1/2 inhibitor combined with a p38-α/β inhibitor. Activating PKC-α and EGFR directly with the combination of phorbol 12-myristate 13-acetate (PMA) and EGF potentiatedMMP1gene and protein expression, and cell invasion. PMA- and ACh-inducedMMP1expression were strongly diminished by inhibiting Src and abolished by concurrently inhibiting both p38-α/β and Src, indicating that Src mediates the cross-talk between PKC-α and EGFR signaling. Using siRNA knockdown, we identified p38-α as the relevant p38 isoform. Collectively, these studies uncover novel functional interactions between post-muscarinic receptor signaling pathways that augmentMMP1expression and drive colon cancer cell invasion; targeting these potentiating interactions has therapeutic potential.
authors.affiliation: Division of Gastroenterology and Hepatology, Department of Medicine, University of Maryland School of Medicine, 22 South Greene Street, N3W62, Baltimore, MD 21201-1595, U.S.A.
authors.forename: Anan H
authors.initials: AH
authors.lastname: Said
authors.affiliation: Division of Gastroenterology and Hepatology, Department of Medicine, University of Maryland School of Medicine, 22 South Greene Street, N3W62, Baltimore, MD 21201-1595, U.S.A.
authors.forename: Shien
authors.initials: S
authors.lastname: Hu
authors.affiliation: Division of Gastroenterology and Hepatology, Department of Medicine, University of Maryland School of Medicine, 22 South Greene Street, N3W62, Baltimore, MD 21201-1595, U.S.A.
authors.forename: Ameer
authors.initials: A
authors.lastname: Abutaleb
authors.affiliation: Division of Gastroenterology and Hepatology, Department of Medicine, University of Maryland School of Medicine, 22 South Greene Street, N3W62, Baltimore, MD 21201-1595, U.S.A.
authors.forename: Tonya
authors.initials: T
authors.lastname: Watkins
authors.affiliation: Division of Gastroenterology and Hepatology, Department of Medicine, University of Maryland School of Medicine, 22 South Greene Street, N3W62, Baltimore, MD 21201-1595, U.S.A.|Veterans Affairs Maryland Health Care System, Baltimore, MD 21201-1595, U.S.A.
authors.forename: Kunrong
authors.initials: K
authors.lastname: Cheng
authors.affiliation: Division of Gastroenterology and Hepatology, Department of Medicine, University of Maryland School of Medicine, 22 South Greene Street, N3W62, Baltimore, MD 21201-1595, U.S.A.
authors.forename: Ahmed
authors.initials: A
authors.lastname: Chahdi
authors.affiliation: Division of Gastroenterology and Hepatology, Department of Medicine, University of Maryland School of Medicine, 22 South Greene Street, N3W62, Baltimore, MD 21201-1595, U.S.A.
authors.forename: Panjamurthy
authors.initials: P
authors.lastname: Kuppusamy
authors.affiliation: Division of Gastroenterology and Hepatology, Department of Medicine, University of Maryland School of Medicine, 22 South Greene Street, N3W62, Baltimore, MD 21201-1595, U.S.A.
authors.forename: Neeraj
authors.initials: N
authors.lastname: Saxena
authors.affiliation: Division of Gastroenterology and Hepatology, Department of Medicine, University of Maryland School of Medicine, 22 South Greene Street, N3W62, Baltimore, MD 21201-1595, U.S.A.|Veterans Affairs Maryland Health Care System, Baltimore, MD 21201-1595, U.S.A.
authors.forename: Guofeng
authors.initials: G
authors.lastname: Xie
authors.affiliation: Division of Gastroenterology and Hepatology, Department of Medicine, University of Maryland School of Medicine, 22 South Greene Street, N3W62, Baltimore, MD 21201-1595, U.S.A.|Veterans Affairs Maryland Health Care System, Baltimore, MD 21201-1595, U.S.A.|Marlene and Stewart Greenebaum Comprehensive Cancer Center, University of Maryland School of Medicine, Baltimore, MD, U.S.A.
authors.forename: Jean-Pierre
authors.initials: JP
authors.lastname: Raufman
mesh_terms: D000109:Acetylcholine / Q000494:pharmacology; D018938:Caco-2 Cells; D045744:Cell Line, Tumor; D004815:Epidermal Growth Factor / Q000494:pharmacology; D066246:ErbB Receptors / Q000235:genetics / Q000378:metabolism; D015972:Gene Expression Regulation, Neoplastic; D019073:HT29 Cells; D006801:Humans; D020781:Matrix Metalloproteinase 1 / Q000235:genetics / Q000378:metabolism*; D019950:Mitogen-Activated Protein Kinase 1 / Q000235:genetics / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000235:genetics / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D051571:Protein Kinase C-alpha / Q000235:genetics / Q000378:metabolism; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D043587:Receptor, Muscarinic M3 / Q000235:genetics* / Q000378:metabolism; D015398:Signal Transduction / Q000235:genetics*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D048051:p38 Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D019061:src-Family Kinases / Q000235:genetics / Q000378:metabolism
chemical_list: D047428:Protein Kinase Inhibitors; D034741:RNA, Small Interfering; D043587:Receptor, Muscarinic M3; D004815:Epidermal Growth Factor; C512478:EGFR protein, human; D066246:ErbB Receptors; D019061:src-Family Kinases; D051571:Protein Kinase C-alpha; C535150:MAPK1 protein, human; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D048051:p38 Mitogen-Activated Protein Kinases; C522360:MMP1 protein, human; D020781:Matrix Metalloproteinase 1; D000109:Acetylcholine; D013755:Tetradecanoylphorbol Acetate
keywords: cell invasion; cell signaling; colorectal cancer; matrix metalloproteinases; muscarinic receptors; p38 MAPK