title: Activation of AtMPK9 through autophosphorylation that makes it independent of the canonical MAPK cascades.
abstract: Mitogen-activated protein kinases (MAPKs) are part of conserved signal transduction modules in eukaryotes that are typically organized into three-tiered kinase cascades. The activation of MAPKs in these pathways is fully dependent on the bisphosphorylation of the TXY motif in the T-loop by the pertinent dual-specificity MAPK kinases (MAPKKs). The Arabidopsis mitogen-activated protein kinase 9 (AtMPK9) is a member of an atypical class of MAPKs. Representatives of this MAPK family have a TDY phosphoacceptor site, a long C-terminal extension and lack the common MAPKK-binding docking motif. In the present paper, we describe multiple in vitro and in vivo data showing that AtMPK9 is activated independently of any upstream MAPKKs but rather is activated through autophosphorylation. We mapped the autophosphorylation sites by MS to the TDY motif and to the C-terminal regulatory extension. We mutated the phosphoacceptor sites on the TDY, which confirmed the requirement for bisphorylation at this site for full kinase activity. Next, we demonstrated that the kinase-inactive mutant form of AtMPK9 is not trans-phosphorylated on the TDY site when mixed with an active AtMPK9, implying that the mechanism of the autocatalytic phosphorylation is intramolecular. Furthermore, we show that in vivo AtMPK9 is activated by salt and is regulated by okadaic acid-sensitive phosphatases. We conclude that the plant AtMPK9 shows similarities to the mammalian atypical MAPKs, such as extracellular-signal-regulated kinase (ERK) 7/8, in terms of an MAPKK-independent activation mechanism.
authors: *Department of Medical Chemistry, Molecular Biology and Pathobiochemistry, Semmelweis University, H-1444 Budapest, POB 260, Hungary.
authors: Szilvia K
authors: SK
authors: Nagy
authors: †Proteomics Research Group, Biological Research Center of the Hungarian Academy of Sciences, Szeged, H-6701, Szeged, POB 521, Hungary.
authors: Zsuzsanna
authors: Z
authors: Darula
authors: *Department of Medical Chemistry, Molecular Biology and Pathobiochemistry, Semmelweis University, H-1444 Budapest, POB 260, Hungary.
authors: Brigitta M
authors: BM
authors: Kállai
authors: ‡School of Biological Sciences, Centre for Systems and Synthetic Biology, Royal Holloway, University of London, Egham, Surrey, TW20 0EX, U.K.
authors: László
authors: L
authors: Bögre
authors: *Department of Medical Chemistry, Molecular Biology and Pathobiochemistry, Semmelweis University, H-1444 Budapest, POB 260, Hungary.
authors: Gábor
authors: G
authors: Bánhegyi
authors: †Proteomics Research Group, Biological Research Center of the Hungarian Academy of Sciences, Szeged, H-6701, Szeged, POB 521, Hungary.
authors: Katalin F
authors: KF
authors: Medzihradszky
authors: §Institute of Plant Biology, Biological Research Centre of the Hungarian Academy of Sciences, H-6701, Szeged, POB 521, Hungary.
authors: Gábor V
authors: GV
authors: Horváth
authors: *Department of Medical Chemistry, Molecular Biology and Pathobiochemistry, Semmelweis University, H-1444 Budapest, POB 260, Hungary.
authors: Tamás
authors: T
authors: Mészáros
mesh_terms: D020816:Amino Acid Motifs; D019943:Amino Acid Substitution; D017360:Arabidopsis / Q000166:cytology / Q000201:enzymology / Q000378:metabolism; D029681:Arabidopsis Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D055162:Biocatalysis; D002478:Cells, Cultured; D004789:Enzyme Activation; D020935:MAP Kinase Signaling System; D020928:Mitogen-Activated Protein Kinases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008958:Models, Molecular; D050505:Mutant Proteins / Q000737:chemistry / Q000378:metabolism; D009842:Oligopeptides / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D010766:Phosphorylation; D018517:Plant Roots / Q000166:cytology / Q000201:enzymology / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D013379:Substrate Specificity; D013912:Threonine / Q000737:chemistry*; D014443:Tyrosine / Q000737:chemistry*
chemical_list: D029681:Arabidopsis Proteins; D050505:Mutant Proteins; D009842:Oligopeptides; D011994:Recombinant Proteins; C097918:threonyl-aspartyl-tyrosine; D013912:Threonine; D014443:Tyrosine; C436293:MPK6 protein, Arabidopsis; C583608:MPK9 protein, Arabidopsis; D020928:Mitogen-Activated Protein Kinases