Leader of the research program: Jozef Šamaj and the respective department
We will continue in our effort to unravel developmental and environmental stress signalling by mitogen-activated protein kinases (MAPKs) in plants, as related to polarity, cytoskeleton, vesicular trafficking and hormonal control. An important part of this activity will be the adopting and application of innovative advanced microscopy techniques such as super-resolution and light-sheet microscopy for bioimaging of plant seedlings (mainly focused on MAPKs, cytoskeleton and endosomes). Seedling plants represent crucial developmental stage, which is extremely important for plant growth and production. They represent ideal model to study post-embryogenic plant adaptation to the environment. We would like to build up on our recent results and prepare new transgenic plant lines with selected genetically-altered (point mutations, deletions, constitutive active and dominant negative versions) and fluorescently-tagged MAPKs, cytoskeletal and endomembrane proteins. These transgenic plant lines will be characterized at molecular, subcellular, cellular, tissue, organ and whole plant levels. Research team will apply integrative approaches using molecular methods for gene expression (qPCR, promoter-GUS histochemistry, in situ hybridization), subcellular localization (GFP technology, split-YFP, immunolocalization on the level of light and electron microscopy as well as super-resolution, light-sheet, spinning disc and confocal laser scanning microscopies) in combination with genetic (T-DNA insertion mutants, RNAi technology, overexpression, point mutations and deletions of functional protein domains), (phospho-)proteomic (gel-based and gel-free approaches) and immunobiochemical (Phos-Tag technology, immunoblots, subcellular fractionation, co-immunoprecipitation) methods. The basic research will be performed on model plant Arabidopsis as well as on important crop species such as alfalfa and barley. It is expected that we will obtain unique information about molecular, cellular and developmental control of seedling development related to cytoskeleton, endocytosis and signalling by MAPKs. It is also expected that we will establish and employ the latest microscopical methods for plant bioimaging and provide more complex datasets on proteomes and phosphoproteomes of seedling roots, hypocotyls and cotyledons during development of wild type and mutant/transgenic plants (Arabidopsis, alfalfa and barley).