title: Ras-mutant cancer cells display B-Raf binding to Ras that activates extracellular signal-regulated kinase and is inhibited by protein kinase A phosphorylation.
abstract: The small G protein Ras regulates proliferation through activation of the mitogen-activated protein (MAP) kinase (ERK) cascade. The first step of Ras-dependent activation of ERK signaling is Ras binding to members of the Raf family of MAP kinase kinase kinases, C-Raf and B-Raf. Recently, it has been reported that in melanoma cells harboring oncogenic Ras mutations, B-Raf does not bind to Ras and does not contribute to basal ERK activation. For other types of Ras-mutant tumors, the relative contributions of C-Raf and B-Raf are not known. We examined non-melanoma cancer cell lines containing oncogenic Ras mutations and express both C-Raf and B-Raf isoforms, including the lung cancer cell line H1299 cells. Both B-Raf and C-Raf were constitutively bound to oncogenic Ras and contributed to Ras-dependent ERK activation. Ras binding to B-Raf and C-Raf were both subject to inhibition by the cAMP-dependent protein kinase PKA. cAMP inhibited the growth of H1299 cells and Ras-dependent ERK activation via PKA. PKA inhibited the binding of Ras to both C-Raf and B-Raf through phosphorylations of C-Raf at Ser-259 and B-Raf at Ser-365, respectively. These studies demonstrate that in non-melanocytic Ras-mutant cancer cells, Ras signaling to B-Raf is a significant contributor to ERK activation and that the B-Raf pathway, like that of C-Raf, is a target for inhibition by PKA. We suggest that cAMP and hormones coupled to cAMP may prove useful in dampening the effects of oncogenic Ras in non-melanocytic cancer cells through PKA-dependent actions on B-Raf as well as C-Raf.
authors.affiliation: Vollum Institute, Oregon Health and Science University, Portland, Oregon 97239.
authors.forename: Yanping
authors.initials: Y
authors.lastname: Li
authors.affiliation: Vollum Institute, Oregon Health and Science University, Portland, Oregon 97239.
authors.forename: Maho
authors.initials: M
authors.lastname: Takahashi
authors.affiliation: Vollum Institute, Oregon Health and Science University, Portland, Oregon 97239. Electronic address: stork@ohsu.edu.
authors.forename: Philip J S
authors.initials: PJS
authors.lastname: Stork
mesh_terms: D045744:Cell Line, Tumor; D000242:Cyclic AMP / Q000235:genetics / Q000378:metabolism; D017868:Cyclic AMP-Dependent Protein Kinases / Q000235:genetics / Q000378:metabolism*; D004789:Enzyme Activation / Q000235:genetics; D048049:Extracellular Signal-Regulated MAP Kinases / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008175:Lung Neoplasms / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D009154:Mutation; D015689:Oncogene Protein p21(ras) / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation / Q000235:genetics; D011485:Protein Binding / Q000235:genetics; D048493:Proto-Oncogene Proteins B-raf / Q000235:genetics / Q000378:metabolism*; D019908:Proto-Oncogene Proteins c-raf / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction
chemical_list: D000242:Cyclic AMP; C482119:BRAF protein, human; D048493:Proto-Oncogene Proteins B-raf; D019908:Proto-Oncogene Proteins c-raf; D017868:Cyclic AMP-Dependent Protein Kinases; D048049:Extracellular Signal-Regulated MAP Kinases; D015689:Oncogene Protein p21(ras)
keywords: B-Raf; C-Raf; ERK; MAP Kinases (MAPKs); Protein Kinase A (PKA); Raf; Ras
references.citation: Mol Cell Biol. 2002 Jun;22(11):3717-28
references.pmid: 11997508
references.citation: FEBS Lett. 1995 Jan 9;357(3):290-6
references.pmid: 7835430
references.citation: Oncogene. 2006 Oct 12;25(47):6262-76
references.pmid: 16702958
references.citation: Nat Cell Biol. 2010 Dec;12(12):1242-9
references.pmid: 21102438
references.citation: Int Immunol. 2013 Apr;25(4):259-69
references.pmid: 23334952
references.citation: Cell. 1993 Jul 16;74(1):205-14
references.pmid: 8334704
references.citation: Nucleic Acids Res. 2011 Jan;39(Database issue):D945-50
references.pmid: 20952405
references.citation: J Biol Chem. 2000 Sep 15;275(37):28688-94
references.pmid: 10862777
references.citation: Trends Cell Biol. 2002 Jun;12(6):258-66
references.pmid: 12074885
references.citation: Nat Rev Cancer. 2011 Oct 13;11(11):761-74
references.pmid: 21993244
references.citation: Cell. 1997 Apr 4;89(1):73-82
references.pmid: 9094716
references.citation: Mol Cell Biol. 2002 May;22(10):3237-46
references.pmid: 11971957
references.citation: Lung Cancer. 2007 Apr;56(1):51-8
references.pmid: 17275947
references.citation: EMBO J. 2001 Jul 2;20(13):3459-72
references.pmid: 11432833
references.citation: J Biol Chem. 2006 Apr 7;281(14):9238-50
references.pmid: 16452469
references.citation: Mol Pharmacol. 2009 Nov;76(5):1123-9
references.pmid: 19720729
references.citation: PLoS One. 2009 May 27;4(5):e5717
references.pmid: 19492075
references.citation: Br J Cancer. 2005 Jan 17;92(1):131-9
references.pmid: 15597105
references.citation: J Biol Chem. 2000 Nov 3;275(44):34433-41
references.pmid: 10950954
references.citation: Mol Cell Biol. 2010 Aug;30(16):3956-69
references.pmid: 20547757
references.citation: Mol Cell Biol. 1998 Jul;18(7):3947-55
references.pmid: 9632779
references.citation: Mol Cell Biol. 2007 Jan;27(1):31-43
references.pmid: 17074813
references.citation: Nat Struct Mol Biol. 2011 May;18(5):584-91
references.pmid: 21478863
references.citation: Cancer Res. 2006 Oct 1;66(19):9483-91
references.pmid: 17018604
references.citation: J Biol Chem. 2000 Aug 18;275(33):25342-50
references.pmid: 10840035
references.citation: Curr Biol. 2011 Apr 12;21(7):563-8
references.pmid: 21458265
references.citation: J Biol Chem. 2002 Jun 21;277(25):22191-200
references.pmid: 11956184
references.citation: Cancer Biol Ther. 2010 Jan;9(2):111-9
references.pmid: 20009523
references.citation: Biochem Biophys Res Commun. 2010 Aug 27;399(3):313-7
references.pmid: 20674547
references.citation: J Biol Chem. 2003 Aug 8;278(32):29819-23
references.pmid: 12801936
references.citation: Trends Mol Med. 2012 Jan;18(1):27-35
references.pmid: 21962474
references.citation: Mol Cell Biol. 2008 Dec;28(23):7109-25
references.pmid: 18824540
references.citation: Mol Cell Biol. 2002 Aug;22(15):5281-95
references.pmid: 12101225
references.citation: Cell Cycle. 2009 Sep 1;8(17):2676-7
references.pmid: 19690457
references.citation: Mol Endocrinol. 2001 Apr;15(4):614-24
references.pmid: 11266512
references.citation: Nat Genet. 2008 May;40(5):600-8
references.pmid: 18372904
references.citation: Mol Carcinog. 2006 Jun;45(6):397-402
references.pmid: 16652378
references.citation: Mol Cell Biol. 2002 Jul;22(14):4984-96
references.pmid: 12077328
references.citation: Mol Cell Biol. 2006 Mar;26(6):2130-45
references.pmid: 16507992
references.citation: Sci Signal. 2013 Jun 25;6(281):ra51
references.pmid: 23800469
references.citation: Genes Dev. 2007 Dec 15;21(24):3214-31
references.pmid: 18079171
references.citation: Nature. 2010 Mar 18;464(7287):427-30
references.pmid: 20179705
references.citation: Proc Natl Acad Sci U S A. 1993 Nov 1;90(21):10300-4
references.pmid: 7694289
references.citation: EMBO J. 2000 Jun 15;19(12):2900-10
references.pmid: 10856235
references.citation: Cancer Res. 2012 May 15;72(10):2457-67
references.pmid: 22589270
references.citation: Cell. 2010 Jan 22;140(2):209-21
references.pmid: 20141835
references.citation: J Biol Chem. 2005 Nov 18;280(46):38276-89
references.pmid: 16172130
references.citation: Leuk Res. 2003 Jan;27(1):51-6
references.pmid: 12479852
references.citation: Kidney Int. 2003 Jun;63(6):1983-94
references.pmid: 12753285
references.citation: Nature. 2010 Mar 18;464(7287):431-5
references.pmid: 20130576
references.citation: J Biol Chem. 2000 Oct 13;275(41):31921-9
references.pmid: 10931830
references.citation: Neoplasia. 2006 Jan;8(1):1-8
references.pmid: 16533420
references.citation: J Surg Oncol. 2011 May 1;103(6):464-7
references.pmid: 21480237
references.citation: FEBS J. 2005 Jul;272(14):3491-504
references.pmid: 16008550
references.citation: Small GTPases. 2011 Sep;2(5):289-292
references.pmid: 22292133
references.citation: J Clin Oncol. 2004 Nov 15;22(22):4456-62
references.pmid: 15483017
references.citation: Cancer Res. 1989 Sep 1;49(17):4682-9
references.pmid: 2547513
references.citation: Regul Pept. 2002 Nov 15;109(1-3):135-40
references.pmid: 12409225
references.citation: Mol Cell Biol. 1994 Oct;14(10):6522-30
references.pmid: 7935374
references.citation: J Biol Chem. 2003 Jan 31;278(5):3185-96
references.pmid: 12446733
references.citation: Mol Cell Biol. 2010 Feb;30(3):806-19
references.pmid: 19933846
references.citation: Cancer Res. 2002 Dec 1;62(23):6997-7000
references.pmid: 12460918
references.citation: J Biol Chem. 2002 May 24;277(21):18598-604
references.pmid: 11897793
references.citation: Nat Cell Biol. 2004 Aug;6(8):770-6
references.pmid: 15258589
references.citation: J Cell Biochem. 2009 Nov 1;108(4):998-1011
references.pmid: 19725049
references.citation: Oncogene. 2007 Feb 15;26(7):1056-66
references.pmid: 16924241
references.citation: Science. 1993 Nov 12;262(5136):1065-9
references.pmid: 7694366
references.citation: Eur J Neurosci. 2009 Jan;29(2):287-306
references.pmid: 19200235
references.citation: J Biol Chem. 2012 Apr 20;287(17):13633-43
references.pmid: 22393060
references.citation: Mol Endocrinol. 2002 Jul;16(7):1638-51
references.pmid: 12089357