title: Soybean nodule autoregulation receptor kinase phosphorylates two kinase-associated protein phosphatases in vitro.
abstract: The NARK (nodule autoregulation receptor kinase) gene, a negative regulator of cell proliferation in nodule primordia in several legumes, encodes a receptor kinase that consists of an extracellular leucine-rich repeat and an intracellular serine/threonine protein kinase domain. The putative catalytic domain of NARK was expressed and purified as a maltose-binding or a glutathione S-transferase fusion protein in Escherichia coli. The recombinant NARK proteins showed autophosphorylation activity in vitro. Several regions of the NARK kinase domain were shown by mass spectrometry to possess phosphoresidues. The kinase-inactive protein K724E failed to autophosphorylate, as did three other proteins corresponding to phenotypically detected mutants defective in whole plant autoregulation of nodulation. A wild-type NARK fusion protein transphosphorylated a kinase-inactive mutant NARK fusion protein, suggesting that it is capable of intermolecular autophosphorylation in vitro. In addition, Ser-861 and Thr-963 in the NARK kinase catalytic domain were identified as phosphorylation sites through site-directed mutagenesis. The genes coding for the kinase-associated protein phosphatases KAPP1 and KAPP2, two putative interacting components of NARK, were isolated. NARK kinase domain phosphorylated recombinant KAPP proteins in vitro. Autophosphorylated NARK kinase domain was, in turn, dephosphorylated by both KAPP1 and KAPP2. Our results suggest a model for signal transduction involving NARK in the control of nodule development.
authors.affiliation: Australian Research Council Centre of Excellence for Integrative Legume Research, St. Lucia, Brisbane, Queensland 4072, Australia.
authors.forename: Akira
authors.initials: A
authors.lastname: Miyahara
authors.affiliation: Australian Research Council Centre of Excellence for Integrative Legume Research, St. Lucia, Brisbane, Queensland 4072, Australia.
authors.forename: Tripty A
authors.initials: TA
authors.lastname: Hirani
authors.affiliation: Australian Research Council Centre of Excellence for Integrative Legume Research, Research School of Biological Sciences, Australian National University, Canberra, Australian Capital Territory 2601, Australia.
authors.forename: Marie
authors.initials: M
authors.lastname: Oakes
authors.affiliation: Australian Research Council Centre of Excellence for Integrative Legume Research, St. Lucia, Brisbane, Queensland 4072, Australia.
authors.forename: Attila
authors.initials: A
authors.lastname: Kereszt
authors.affiliation: School of Microbial and Molecular Sciences and Institute for Molecular Bioscience, University of Queensland, St. Lucia, Brisbane, Queensland 4072, Australia and the.
authors.forename: Bostjan
authors.initials: B
authors.lastname: Kobe
authors.affiliation: Australian Research Council Centre of Excellence for Integrative Legume Research, Research School of Biological Sciences, Australian National University, Canberra, Australian Capital Territory 2601, Australia.
authors.forename: Michael A
authors.initials: MA
authors.lastname: Djordjevic
authors.affiliation: Australian Research Council Centre of Excellence for Integrative Legume Research, St. Lucia, Brisbane, Queensland 4072, Australia. Electronic address: p.gresshoff@uq.edu.au.
authors.forename: Peter M
authors.initials: PM
authors.lastname: Gresshoff
mesh_terms: D000595:Amino Acid Sequence; D017360:Arabidopsis / Q000378:metabolism; D020134:Catalytic Domain; D049109:Cell Proliferation; D004926:Escherichia coli / Q000378:metabolism*; D005978:Glutathione / Q000378:metabolism*; D008954:Models, Biological; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D010766:Phosphorylation; D010770:Phosphotransferases / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D011956:Receptors, Cell Surface / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry; D017386:Sequence Homology, Amino Acid; D013025:Glycine max / Q000378:metabolism*
chemical_list: D011956:Receptors, Cell Surface; D011994:Recombinant Proteins; D010770:Phosphotransferases; D017346:Protein Serine-Threonine Kinases; D005978:Glutathione