title: Towards the systematic discovery of signal transduction networks using phosphorylation dynamics data.
abstract: BACKGROUND
  Phosphorylation is a ubiquitous and fundamental regulatory mechanism that controls signal transduction in living cells. The number of identified phosphoproteins and their phosphosites is rapidly increasing as a result of recent mass spectrometry-based approaches.
  
  
  RESULTS
  We analyzed time-course phosphoproteome data obtained previously by liquid chromatography mass spectrometry with the stable isotope labeling using amino acids in cell culture (SILAC) method. This provides the relative phosphorylation activities of digested peptides at each of five time points after stimulating HeLa cells with epidermal growth factor (EGF). We initially calculated the correlations between the phosphorylation dynamics patterns of every pair of peptides and connected the strongly correlated pairs to construct a network. We found that peptides extracted from the same intracellular fraction (nucleus vs. cytoplasm) tended to be close together within this phosphorylation dynamics-based network. The network was then analyzed using graph theory and compared with five known signal-transduction pathways. The dynamics-based network was correlated with known signaling pathways in the NetPath and Phospho.ELM databases, and especially with the EGF receptor (EGFR) signaling pathway. Although the phosphorylation patterns of many proteins were drastically changed by the EGF stimulation, our results suggest that only EGFR signaling transduction was both strongly activated and precisely controlled.
  
  
  CONCLUSIONS
  The construction of a phosphorylation dynamics-based network provides a useful overview of condition-specific intracellular signal transduction using quantitative time-course phosphoproteome data under specific experimental conditions. Detailed prediction of signal transduction based on phosphoproteome dynamics remains challenging. However, since the phosphorylation profiles of kinase-substrate pairs on the specific pathway were localized in the dynamics-based network, our method will be a complementary strategy to explore new components of protein signaling pathways in combination with previous methods (including software) of predicting direct kinase-substrate relationships.
authors.affiliation: Institute for Advanced Biosciences, Keio University, Tsuruoka, Yamagata, Japan.
authors.forename: Haruna
authors.initials: H
authors.lastname: Imamura
authors.forename: Nozomu
authors.initials: N
authors.lastname: Yachie
authors.forename: Rintaro
authors.initials: R
authors.lastname: Saito
authors.forename: Yasushi
authors.initials: Y
authors.lastname: Ishihama
authors.forename: Masaru
authors.initials: M
authors.lastname: Tomita
mesh_terms: D004815:Epidermal Growth Factor / Q000378:metabolism; D066246:ErbB Receptors / Q000378:metabolism; D013058:Mass Spectrometry / Q000379:methods; D010766:Phosphorylation; D011506:Proteins / Q000737:chemistry* / Q000378:metabolism; D040901:Proteomics / Q000379:methods*; D015398:Signal Transduction
chemical_list: D011506:Proteins; D004815:Epidermal Growth Factor; D066246:ErbB Receptors
references.citation: Genome Biol. 2007;8(11):R250
references.pmid: 18039369
references.citation: Sci STKE. 2003 Jul 15;2003(191):RE12
references.pmid: 12865499
references.citation: Mol Cell Proteomics. 2009 May;8(5):1061-71
references.pmid: 19136663
references.citation: Mol Cell Proteomics. 2005 Mar;4(3):310-27
references.pmid: 15665377
references.citation: Genome Res. 2003 Nov;13(11):2498-504
references.pmid: 14597658
references.citation: Proteomics. 2009 May;9(10):2764-75
references.pmid: 19405024
references.citation: Cell. 2006 Nov 3;127(3):635-48
references.pmid: 17081983
references.citation: Nucleic Acids Res. 2008 Jan;36(Database issue):D240-4
references.pmid: 17962309
references.citation: Sci Signal. 2008 Sep 02;1(35):ra2
references.pmid: 18765831
references.citation: In Silico Biol. 2007;7(2):141-4
references.pmid: 17688438
references.citation: Science. 2009 Dec 11;326(5959):1502-9
references.pmid: 20007894
references.citation: Biochim Biophys Acta. 2005 Dec 30;1754(1-2):200-9
references.pmid: 16172032
references.citation: Nat Biotechnol. 2005 Nov;23(11):1391-8
references.pmid: 16273072
references.citation: Science. 2002 Dec 6;298(5600):1912-34
references.pmid: 12471243
references.citation: Trends Biotechnol. 2001 May;19(5):178-81
references.pmid: 11301130
references.citation: Genome Biol. 2010 Jan 12;11(1):R3
references.pmid: 20067622
references.citation: Sci Signal. 2010 Jan 12;3(104):ra3
references.pmid: 20068231
references.citation: J Proteome Res. 2008 Apr;7(4):1675-82
references.pmid: 18327897
references.citation: Eur J Biochem. 2001 Oct;268(19):5001-10
references.pmid: 11589691
references.citation: PLoS One. 2009 Aug 26;4(8):e6522
references.pmid: 19707567
references.citation: Trends Biochem Sci. 2005 Jun;30(6):286-90
references.pmid: 15950870
references.citation: Cell Stem Cell. 2009 Aug 7;5(2):214-26
references.pmid: 19664995
references.citation: Proc Natl Acad Sci U S A. 2007 Jan 30;104(5):1488-93
references.pmid: 17242355
references.citation: Nucleic Acids Res. 2009 Jan;37(Database issue):D767-72
references.pmid: 18988627
references.citation: Mol Syst Biol. 2008;4:193
references.pmid: 18463617
references.citation: Proc Natl Acad Sci U S A. 2007 Feb 13;104(7):2193-8
references.pmid: 17287358
references.citation: Mol Cell Proteomics. 2009 Aug;8(8):1908-20
references.pmid: 19429919
references.citation: Nat Rev Mol Cell Biol. 2006 Jun;7(6):391-403
references.pmid: 16723975
references.citation: J Mol Biol. 1999 Dec 17;294(5):1351-62
references.pmid: 10600390
references.citation: Nat Biotechnol. 2001 Apr;19(4):348-53
references.pmid: 11283593
references.citation: Nat Protoc. 2007;2(8):1896-906
references.pmid: 17703201
references.citation: Proc Natl Acad Sci U S A. 2004 Aug 17;101(33):12130-5
references.pmid: 15302935
references.citation: Oncogene. 2005 Apr 18;24(17):2860-70
references.pmid: 15838520
references.citation: Nucleic Acids Res. 2010 Jan;38(Database issue):D142-8
references.pmid: 19843607
references.citation: Trends Biochem Sci. 2000 Dec;25(12):596-601
references.pmid: 11116185
references.citation: Mol Cell. 2008 Sep 26;31(6):777-81
references.pmid: 18922462
references.citation: Proteomics. 2007 Aug;7(16):2751-68
references.pmid: 17703509
references.citation: Mol Cell Proteomics. 2008 Feb;7(2):299-307
references.pmid: 17938405
references.citation: Nature. 2005 Dec 1;438(7068):679-84
references.pmid: 16319894
references.citation: Nucleic Acids Res. 2003 Jul 1;31(13):3635-41
references.pmid: 12824383
references.citation: Cell. 2007 Jun 29;129(7):1415-26
references.pmid: 17570479
references.citation: Mol Cell Proteomics. 2005 Sep;4(9):1240-50
references.pmid: 15951569
references.citation: Brief Funct Genomic Proteomic. 2008 Jan;7(1):17-26
references.pmid: 18270216
references.citation: Proteomics. 2004 Jul;4(7):1985-8
references.pmid: 15221759