title: The human Na(+)/H(+) exchanger 1 is a membrane scaffold protein for extracellular signal-regulated kinase 2.
abstract: BACKGROUND
  Extracellular signal-regulated kinase 2 (ERK2) is an S/T kinase with more than 200 known substrates, and with critical roles in regulation of cell growth and differentiation and currently no membrane proteins have been linked to ERK2 scaffolding.
  
  
  RESULTS
  Here, we identify the human Na(+)/H(+) exchanger 1 (hNHE1) as a membrane scaffold protein for ERK2 and show direct hNHE1-ERK1/2 interaction in cellular contexts. Using nuclear magnetic resonance (NMR) spectroscopy and immunofluorescence analysis we demonstrate that ERK2 scaffolding by hNHE1 occurs by one of three D-domains and by two non-canonical F-sites located in the disordered intracellular tail of hNHE1, mutation of which reduced cellular hNHE1-ERK1/2 co-localization, as well as reduced cellular ERK1/2 activation. Time-resolved NMR spectroscopy revealed that ERK2 phosphorylated the disordered tail of hNHE1 at six sites in vitro, in a distinct temporal order, with the phosphorylation rates at the individual sites being modulated by the docking sites in a distant dependent manner.
  
  
  CONCLUSIONS
  This work characterizes a new type of scaffolding complex, which we term a "shuffle complex", between the disordered hNHE1-tail and ERK2, and provides a molecular mechanism for the important ERK2 scaffolding function of the membrane protein hNHE1, which regulates the phosphorylation of both hNHE1 and ERK2.
authors.affiliation: Cell and Developmental Biology, Department of Biology, University of Copenhagen, Universitetsparken 13, DK-2100, Copenhagen Ø, Denmark.|Structural Biology and NMR Laboratory, Department of Biology, University of Copenhagen, Ole Maaløes Vej 5, DK-2200, Copenhagen N, Denmark.
authors.forename: Ruth
authors.initials: R
authors.lastname: Hendus-Altenburger
authors.affiliation: Cell and Developmental Biology, Department of Biology, University of Copenhagen, Universitetsparken 13, DK-2100, Copenhagen Ø, Denmark.
authors.forename: Elena
authors.initials: E
authors.lastname: Pedraz-Cuesta
authors.affiliation: Cell and Developmental Biology, Department of Biology, University of Copenhagen, Universitetsparken 13, DK-2100, Copenhagen Ø, Denmark.
authors.forename: Christina W
authors.initials: CW
authors.lastname: Olesen
authors.affiliation: Structural Biology and NMR Laboratory, Department of Biology, University of Copenhagen, Ole Maaløes Vej 5, DK-2200, Copenhagen N, Denmark.
authors.forename: Elena
authors.initials: E
authors.lastname: Papaleo
authors.affiliation: Structural Biology and NMR Laboratory, Department of Biology, University of Copenhagen, Ole Maaløes Vej 5, DK-2200, Copenhagen N, Denmark.
authors.forename: Jeff A
authors.initials: JA
authors.lastname: Schnell
authors.affiliation: Physical and Theoretical Chemistry Laboratory, Department of Chemistry, University of Oxford, South Parks Road, Oxford, OX1 3QZ, UK.
authors.forename: Jonathan T S
authors.initials: JT
authors.lastname: Hopper
authors.affiliation: Physical and Theoretical Chemistry Laboratory, Department of Chemistry, University of Oxford, South Parks Road, Oxford, OX1 3QZ, UK.
authors.forename: Carol V
authors.initials: CV
authors.lastname: Robinson
authors.affiliation: Cell and Developmental Biology, Department of Biology, University of Copenhagen, Universitetsparken 13, DK-2100, Copenhagen Ø, Denmark. SFPedersen@bio.ku.dk.
authors.forename: Stine F
authors.initials: SF
authors.lastname: Pedersen
authors.affiliation: Structural Biology and NMR Laboratory, Department of Biology, University of Copenhagen, Ole Maaløes Vej 5, DK-2200, Copenhagen N, Denmark. bbk@bio.ku.dk.
authors.forename: Birthe B
authors.initials: BB
authors.lastname: Kragelund
mesh_terms: D000595:Amino Acid Sequence; D027682:Cation Transport Proteins / Q000737:chemistry / Q000378:metabolism*; D002460:Cell Line; D004789:Enzyme Activation; D006801:Humans; D064267:Intrinsically Disordered Proteins / Q000737:chemistry / Q000378:metabolism*; D019950:Mitogen-Activated Protein Kinase 1 / Q000737:chemistry / Q000378:metabolism*; D048052:Mitogen-Activated Protein Kinase 3 / Q000737:chemistry / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010766:Phosphorylation; D017510:Protein Folding; D060066:Protein Interaction Maps; D017434:Protein Structure, Tertiary; D000074082:Sodium-Hydrogen Exchanger 1; D017923:Sodium-Hydrogen Exchangers / Q000737:chemistry / Q000378:metabolism*
chemical_list: D027682:Cation Transport Proteins; D064267:Intrinsically Disordered Proteins; C490998:SLC9A1 protein, human; D000074082:Sodium-Hydrogen Exchanger 1; D017923:Sodium-Hydrogen Exchangers; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3
keywords: Intrinsically disordered protein; MAPK; NHE1; NMR; Phosphorylation; Scaffold; Shuffle complex