Identifizierung von RNA Editing Faktoren in Pflanzen durch EMS-Mutanten
Pflanzenphysiologie
Zusammenfassung der Projektergebnisse
We have made progress in the identification of new components of the RNA editing complex in plant mitochondria and plastids. RNA editing process converts specific C nucleotides to U mostly in mRNAs of plant organelles. So far several RNA editing factors have been isolated including PLS-type PPR proteins and MORF proteins. PPR-type RNA editing factors consist of a N- terminal PPR domain, which has a sequence specific RNA binding activity and a C- terminal E domain. A half of them additionally contain a further C-terminal DYW domain, which is most likely to have cytidine deaminase activity. Therefore, it has been considered that additional components, which provide DYW domain to PLS-type PPR proteins with only E domains (E or E+ subclass) are required. To identify such additional factors, we employed two distinct approaches. The first approach, screening for EMS mutants showing RNA editing in multiple sites identified only canonical PLS-type PPR RNA editing factors, whereas in vivo co-immunoprecipitation with known RNA editing factors revealed several new types of candidate co-factors of E or E+ subclass PPR proteins. Among them, DYW2 and NUWA proteins were genetically and phenotypically analyzed in collaboration with other two research groups. The obtained results suggested that DYW2 is likely to confer its DYW domain to E+ subclass PPR protein, while NUWA appears to enhance the interaction between the E+ subclass PPR and DYW2. We also analyzed the function of MORF (multiple organellar RNA editing factor) proteins in RNA editing complexes. MORF proteins are a protein family involved in RNA editing process in seedling plants. Structural analysis of MORF1 and MORF9 revealed that MORFs form a dimer at the central part of the conserved MORF domain. MORF proteins were co-immunoprecipitated with DYW2 and NUWA and interact with various PPR-type RNA editing factors, suggesting they play pivotal roles for constructing RNA editosomes in seedling plants.
Projektbezogene Publikationen (Auswahl)
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(2015) MEF13 requires MORF3 and MORF8 for RNA editing at eight targets in mitochondrial mRNAs in Arabidopsis thaliana. Mol. Plant 8, 1466-77
Glass, F., Härtel, B., Zehrmann, A., Verbitskiy, D., Takenaka, M.
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(2015) Selective homo- and heteromer interactions between the multiple organellar RNA editing factor (MORF) proteins in Arabidopsis thaliana. J. Biol. Chem., 290, 6445–56
Zehrmann, A., Härtel, B., Glass, F., Bayer-Császár, E., Obata, T., Meyer, E., Brennicke, A. and Takenaka, M.
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(2016) A PPR protein in the PLS subfamily stabilizes the 5’-end of processed rpl16 mRNAs in maize chloroplasts. Nucleic Acids Res. 44, 4278-4288
Hammani, K. Takenaka M. Miranda, R. Barkan, A.
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(2017) Crystal structures of the Arabidopsis organellar RNA editing factors MORF1 and MORF9. Nucleic Acids Res. 45, 4915–4928
Haag, S., Schindler, M., Berndt, L., Brennicke, A., Takenaka, M., Weber, G.
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(2017) Two interacting PPR proteins are major Arabidopsis editing factors in plastid and mitochondria. Proc. Natl. Acad. Sci. USA. 114, 8877-8882
Guillaumot, D., Lopez-Obando, M., Baudry, K., Avon, A., Rigaill, G., Falcon de Longevialle, A., Broche, B., Takenaka, M., Berthomé, R., De Jaeger, G., Delannoy, E., Lurin, C.
