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Construction of human activity‐based phosphorylation networks
Authors:Robert H Newman  Jianfei Hu  Hee‐Sool Rho  Zhi Xie  Crystal Woodard  John Neiswinger  Christopher Cooper  Matthew Shirley  Hillary M Clark  Shaohui Hu  Woochang Hwang  Jun Seop Jeong  George Wu  Jimmy Lin  Xinxin Gao  Qiang Ni  Renu Goel  Shuli Xia  Hongkai Ji  Kevin N Dalby  Morris J Birnbaum  Philip A Cole  Stefan Knapp  Alexey G Ryazanov  Donald J Zack  Seth Blackshaw  Tony Pawson  Anne‐Claude Gingras  Stephen Desiderio  Akhilesh Pandey  Benjamin E Turk  Jin Zhang  Heng Zhu  Jiang Qian
Affiliation:1. Department of Pharmacology and Molecular Sciences, Johns Hopkins School of Medicine, , Baltimore, MD, USA;2. Department of Biology, North Carolina Agricultural and Technical State University, , Greensboro, NC, USA;3. Department of Ophthalmology, Johns Hopkins School of Medicine, , Baltimore, MD, USA;4. Center for High‐Throughput Biology, Johns Hopkins School of Medicine, , Baltimore, MD, USA;5. Department of Molecular Biology and Genetics, Johns Hopkins School of Medicine, , Baltimore, MD, USA;6. Department of Biostatistics, Johns Hopkins Bloomberg School of Public Health, , Baltimore, MD, USA;7. The Sidney Kimmel Comprehensive Cancer Center, Johns Hopkins School of Medicine, , Baltimore, MD, USA;8. Institute of Bioinformatics, International Tech Park, , Bangalore, India;9. Hugo W. Moser Kennedy Krieger Institute, , Baltimore, MD, USA;10. Division of Medicinal Chemistry, College of Pharmacy, University of Texas at Austin, , Austin, TX, USA;11. Department of Medicine, University of Pennsylvania School of Medicine, , Philadelphia, PA, USA;12. Nuffield Department of Clinical Medicine, Structural Genomics Consortium, University of Oxford, , Oxford, UK;13. Department of Pharmacology, University of Medicine and Dentistry of New Jersey, , Piscataway, NJ, USA;14. Sol H. Snyder Department of Neuroscience, Johns Hopkins School of Medicine, , Baltimore, MD, USA;15. The McKusick‐Nathans Institute of Genetic Medicine, The Johns Hopkins University School of Medicine, , Baltimore, MD, USA;16. Department of Neurology, Johns Hopkins School of Medicine, , Baltimore, MD, USA;17. Institute of Cell Engineering, Johns Hopkins School of Medicine, , Baltimore, MD, USA;18. Centre for Systems Biology, Samuel Lunenfeld Research Institute, Mount Sinai Hospital Toronto, , ON, Canada;19. Department of Molecular Genetics, University of Toronto, , Toronto, ON, Canada;20. Department of Pharmacology, Yale University School of Medicine, , New Haven, CT, USA
Abstract:The landscape of human phosphorylation networks has not been systematically explored, representing vast, unchartered territories within cellular signaling networks. Although a large number of in vivo phosphorylated residues have been identified by mass spectrometry (MS)‐based approaches, assigning the upstream kinases to these residues requires biochemical analysis of kinase‐substrate relationships (KSRs). Here, we developed a new strategy, called CEASAR, based on functional protein microarrays and bioinformatics to experimentally identify substrates for 289 unique kinases, resulting in 3656 high‐quality KSRs. We then generated consensus phosphorylation motifs for each of the kinases and integrated this information, along with information about in vivo phosphorylation sites determined by MS, to construct a high‐resolution map of phosphorylation networks that connects 230 kinases to 2591 in vivo phosphorylation sites in 652 substrates. The value of this data set is demonstrated through the discovery of a new role for PKA downstream of Btk (Bruton's tyrosine kinase) during B‐cell receptor signaling. Overall, these studies provide global insights into kinase‐mediated signaling pathways and promise to advance our understanding of cellular signaling processes in humans.
Keywords:phosphorylation  signaling networks  systems biology
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