title: The unique characteristics of HOG pathway MAPKs in the extremely halotolerant Hortaea werneckii.
abstract: HwHog1A/B, Hortaea werneckii homologues of the MAP kinase Hog1 from Saccharomyces cerevisiae, are vital for the extreme halotolerance of H. werneckii. In mesophilic S. cerevisiae, Hog1 is phosphorylated already at low osmolyte concentrations, and regulates expression of a similar set of genes independent of osmolyte type. To understand how HwHog1 kinases activity is regulated in H. werneckii, we studied HwHog1A/B activation in vivo, by following phosphorylation of HwHog1A/B in H. werneckii exposed to various osmolytes, and in vitro, by measuring kinase activities of recombinant HwHog1A, HwHog1B and Hog1ΔC. To this end, highly pure and soluble recombinant Hog1 homologues were isolated from insect cells. Our results demonstrate that HwHog1A/B are, in general, transiently phosphorylated in cells shocked with ≥3 M osmolyte, yet constitutive phosphorylation is observed at extreme NaCl and KCl concentrations. Importantly, phosphorylation profiles differ depending on the osmolyte type. Additionally, phosphorylated recombinant HwHog1A/B show lower specific kinase activities compared to Hog1ΔC. In summary, HOG pathway MAPKs in the extremely halotolerant H. werneckii show unique characteristics compared to S. cerevisiae homologues. The reported findings contribute to defining the key determinants of H. werneckii osmotolerance, which is important for its potential transfer to economically relevant microorganisms and crops.
authors.affiliation: Institute of Biochemistry, Faculty of Medicine, University of Ljubljana, Vrazov Trg 2, SI-1000 Ljubljana, Slovenia.
authors.forename: Anja
authors.initials: A
authors.lastname: Kejžar
authors.affiliation: Institute of Biochemistry, Faculty of Medicine, University of Ljubljana, Vrazov Trg 2, SI-1000 Ljubljana, Slovenia.
authors.forename: Matej
authors.initials: M
authors.lastname: Cibic
authors.affiliation: Department of Chemistry and Molecular Biology, University of Gothenburg, SE-412 96 Gothenburg, Sweden.
authors.forename: Morten
authors.initials: M
authors.lastname: Grøtli
authors.affiliation: Institute of Biochemistry, Faculty of Medicine, University of Ljubljana, Vrazov Trg 2, SI-1000 Ljubljana, Slovenia.
authors.forename: Ana
authors.initials: A
authors.lastname: Plemenitaš
authors.affiliation: Institute of Biochemistry, Faculty of Medicine, University of Ljubljana, Vrazov Trg 2, SI-1000 Ljubljana, Slovenia Centre of Excellence for Integrated Approaches in Chemistry and Biology of Proteins (CIPKeBiP), Jamova 39, SI-1000 Ljubljana, Slovenia metka.lenassi@mf.uni-lj.si.
authors.forename: Metka
authors.initials: M
authors.lastname: Lenassi
mesh_terms: D000595:Amino Acid Sequence; D001203:Ascomycota / Q000201:enzymology / Q000235:genetics / Q000502:physiology*; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015966:Gene Expression Regulation, Fungal; D007093:Imidazoles / Q000494:pharmacology; D020928:Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D003904:Mitosporic Fungi / Q000201:enzymology* / Q000235:genetics / Q000502:physiology*; D009997:Osmotic Pressure; D010766:Phosphorylation; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D011725:Pyridines / Q000494:pharmacology; D011994:Recombinant Proteins / Q000302:isolation & purification; D012441:Saccharomyces cerevisiae / Q000235:genetics; D055049:Salt Tolerance / Q000235:genetics
chemical_list: D005656:Fungal Proteins; D007093:Imidazoles; D047428:Protein Kinase Inhibitors; D011725:Pyridines; D011994:Recombinant Proteins; D020928:Mitogen-Activated Protein Kinases; C093642:SB 203580
keywords: HOG pathway; Hortaea werneckii; MAP kinase; kinase activity; osmotolerance; recombinant Hog1