title: Investigating plasmodesmata genetics with virus-induced gene silencing and an agrobacterium-mediated GFP movement assay.
abstract: Plasmodesmata (PD) are channels that connect the cytoplasm of adjacent plant cells, permitting intercellular transport and communication. PD function and formation are essential to plant growth and development, but we still know very little about the genetic pathways regulating PD transport. Here, we present a method for assaying changes in the rate of PD transport following genetic manipulation. Gene expression in leaves is modified by virus-induced gene silencing. Seven to ten days after infection with Tobacco rattle virus carrying a silencing trigger, the gene(s) of interest is silenced in newly arising leaves. In these new leaves, individual cells are then transformed with Agrobacterium to express GFP, and the rate of GFP diffusion via PD is measured. By measuring GFP diffusion both within the epidermis and between the epidermis and mesophyll, the assay can be used to study the effects of silencing a gene(s) on PD transport in general, or transport through secondary PD specifically. Plant biologists working in several fields will find this assay useful, since PD transport impacts plant physiology, development, and defense.
authors.affiliation: Department of Plant and Microbial Biology, University of California, 281A Koshland Hall, Berkeley, CA, 94720, USA.
authors.forename: Jacob O
authors.initials: JO
authors.lastname: Brunkard
authors.forename: Tessa M
authors.initials: TM
authors.lastname: Burch-Smith
authors.forename: Anne M
authors.initials: AM
authors.lastname: Runkel
authors.forename: Patricia
authors.initials: P
authors.lastname: Zambryski
mesh_terms: D060054:Agrobacterium / Q000235:genetics / Q000378:metabolism; D002450:Cell Communication; D018506:Gene Expression Regulation, Plant; D020868:Gene Silencing; D005818:Genetic Engineering; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D018515:Plant Leaves / Q000235:genetics* / Q000378:metabolism / Q000382:microbiology / Q000821:virology; D010940:Plant Proteins / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism; D010942:Plant Viruses / Q000235:genetics / Q000378:metabolism; D031425:Plasmodesmata / Q000235:genetics / Q000378:metabolism* / Q000382:microbiology / Q000821:virology; D021381:Protein Transport; D015398:Signal Transduction; D014026:Nicotiana / Q000235:genetics* / Q000378:metabolism / Q000382:microbiology / Q000821:virology
chemical_list: D010940:Plant Proteins; D049452:Green Fluorescent Proteins