Synaptotagmin function in virus movement and plant development

Cornell University, Ithaca NY

Investigators

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Abstract

DESCRIPTION (provided by applicant): Plant virus movement proteins transport viral genomes across the cell wall by altering plasmodesmata, transwall pores that connect adjacent plant cells. Synaptotagmins, a multigene family thought to be exclusive to animals, are calcium (Ca2+) sensors that regulate synaptic vesicle exo/endocytosis. Our long term goal is to understand how synaptotagmins regulate virus movement and intercellular transport to control disease resistance, the spread of RNAi signals and cell fate. SYTA is 1 of 5 Arabidopsis synaptotagmins (SYT A-E). SYTA binds to the distinct movement proteins encoded by the Begomoviruses CaLCuV (MPCaLCuV) and SqLCV, and the Tobamovirus TMV (30K). We have shown that SYTA regulates both the formation of early endosomes and the cell-to-cell transport of MPCaLCuV and TMV 30K. We propose that SYTs regulate macromolecular trafficking to plasmodesmata via an endocytic recapture pathway, and potentially entry into and exit from the pore as well. We will use a combination of transgenic plant studies and cell-based expression assays to investigate if SYTs A, C and E are key regulators of plant virus movement and intercellular transport. Biochemical and infectivity studies, and a leaf-based cell trafficking assay will examine if SYTA alone regulates the cell-to-cell movement of most viruses, or whether SYTC and SYTE are also involved, either independent of or partnering with SYTA. The roles of SYTC and SYTE in vesicle trafficking will be defined by immune localization, and the use of cell- and leaf-based secretion assays. Multiphoton imaging of SYTA itself, or with MPCaLCuV and 30K, or SYTC and SYTE, will show how SYTA endosomes ferry movement proteins to plasmodesmata, and how SYTC or SYTE may cooperate in this process. This knowledge of SYT A, C and E action in plant cells will be connected to the regulation of development through promoter::GUS fusion studies to define their spatial and temporal expression, and analysis of transgenic lines in which the expression of each is inhibited by RNAi or T-DNA insertions. Our demonstration of a SYTA-regulated functional link between early endosome formation and intercellular transport provides novel insights into how plasmodesmal function is controlled. Beyond the agronomic impact of engineering disease-resistant plants, our studies are relevant to human health in that they can inform and accelerate studies of animal synaptotagmins in cell-cell communication and development, and of mechanisms by which retroviruses redirect the endosomal machinery for virus maturation and exit from an infected cell. This research investigates the regulation of what seems to be a common pathway for transport between plant and animal cells. As such, it can impact public health by improving nutrition, providing knowledge about growth defects, and identifying novel ways to limit the spread of HIV and other retroviruses.