Stress-activated plant mitogen-activated protein (MAP) kinase pathways play roles in growth adaptation to the environment by modulating cell division through cytoskeletal regulation, but the mechanisms are poorly understood. We performed protein interaction and phosphorylation experiments with cytoskeletal proteins, mass spectrometric identification of MPK6 complexes and immunofluorescence analyses of the microtubular cytoskeleton of mitotic cells using wild-type, mpk6-2 mutant and plants overexpressing the MAP kinase-inactivating phosphatase, AP2C3. We showed that MPK6 interacted with γ-tubulin and co-sedimented with plant microtubules polymerized in vitro. It was the active form of MAP kinase that was enriched with microtubules and followed similar dynamics to γ-tubulin, moving from poles to midzone during the anaphase-to-telophase transition. We found a novel substrate for MPK6, the microtubule plus end protein, EB1c. The mpk6-2 mutant was sensitive to 3-nitro-l-tyrosine (NO2 -Tyr) treatment with respect to mitotic abnormalities, and root cells overexpressing AP2C3 showed defects in chromosome segregation and spindle orientation. Our data suggest that the active form of MAP kinase interacts with γ-tubulin on specific subsets of mitotic microtubules during late mitosis. MPK6 phosphorylates EB1c, but not EB1a, and has a role in maintaining regular planes of cell division under stress conditions.

Department of Medical Chemistry Molecular Biology and Pathobiochemistry Semmelweis University Tűzoltó u 37 47 H 1094 Budapest Hungary

Department of Medical Chemistry Molecular Biology and Pathobiochemistry Semmelweis University Tűzoltó u 37 47 H 1094 Budapest Hungary Technical Analytical Research Group of HAS Szent Gellért tér 4 H 1111 Budapest Hungary

Institute of Microbiology AS CR v v i Vídeňská 1083 142 20 Prague 4 Czech Republic

Max F Perutz Laboratories University of Vienna Vienna Austria Institute of Biotechnology University of Vilnius Vilnius Lithuania

School of Biological Sciences Royal Holloway University of London Egham Surrey TW20 0EX UK

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