title: Live cell imaging of interactions between replicase and capsid protein of Brome mosaic virus using Bimolecular Fluorescence Complementation: implications for replication and genome packaging.
abstract: In Brome mosaic virus, it was hypothesized that a physical interaction between viral replicase and capsid protein (CP) is obligatory to confer genome packaging specificity. Here we tested this hypothesis by employing Bimolecular Fluorescent Complementation (BiFC) as a tool for evaluating protein-protein interactions in living cells. The efficacy of BiFC was validated by a known interaction between replicase protein 1a (p1a) and protein 2a (p2a) at the endoplasmic reticulum (ER) site of viral replication. Additionally, co-expression in planta of a bona fide pair of interacting protein partners of p1a and p2a had resulted in the assembly of a functional replicase. Subsequent BiFC assays in conjunction with mCherry labeled ER as a fluorescent cellular marker revealed that CP physically interacts with p2a, but not p1a, and this CP:p2a interaction occurs at the cytoplasmic phase of the ER. The significance of the CP:p2a interaction in BMV replication and genome packaging is discussed.
authors.affiliation: Department of Plant Pathology & Microbiology, University of California, Riverside, CA 92521-0122, USA.
authors.forename: Sonali
authors.initials: S
authors.lastname: Chaturvedi
authors.affiliation: Department of Plant Pathology & Microbiology, University of California, Riverside, CA 92521-0122, USA. Electronic address: arao@ucr.edu.
authors.forename: A L N
authors.initials: ALN
authors.lastname: Rao
mesh_terms: D017795:Bromovirus / Q000737:chemistry / Q000201:enzymology / Q000235:genetics / Q000502:physiology*; D036022:Capsid Proteins / Q000235:genetics / Q000378:metabolism*; D012321:DNA-Directed RNA Polymerases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004721:Endoplasmic Reticulum / Q000737:chemistry / Q000821:virology; D016679:Genome, Viral; D057054:Molecular Imaging; D010935:Plant Diseases / Q000821:virology*; D011485:Protein Binding; D012324:RNA-Dependent RNA Polymerase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014026:Nicotiana / Q000737:chemistry / Q000821:virology*; D014764:Viral Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D019065:Virus Assembly; D014779:Virus Replication
chemical_list: D036022:Capsid Proteins; D014764:Viral Proteins; D012324:RNA-Dependent RNA Polymerase; D012321:DNA-Directed RNA Polymerases
keywords: BiFC; Bromovirus; Endoplasmic reticulum; Genome Packaging; Protein–Protein Interactions; Replication