Characterization of a novel barley protein, HCP1, that interacts with the Brome mosaic virus coat protein.
Brome mosaic virus (BMV) requires the coat protein (CP) not only for encapsidation but also for viral cell-to-cell and long-distance movement in barley plants. This suggests that BMV infection is controlled by interactions of CP with putative host factors as well as with viral components. To identify the host factors that interact with BMV CP, we screened a barley cDNA library containing 2.4 x 10(6) independent clones, using a yeast two-hybrid system. Using full-length and truncated BMV CPs as baits, four candidate cDNA clones were isolated. One of the candidate cDNAs encodes a unique oxidoreductase enzyme, designated HCP1. HCP1 was found predominantly in the soluble fractions after differential centrifugation of BMV-infected and mock-inoculated barley tissues. A two-hybrid binding assay using a series of truncated BMV CPs demonstrated that a C-terminal portion of CP is essential for its interaction with HCP1. Interestingly, experiments with CP mutants bearing single amino acid substitutions at the C-terminus revealed that the capacity for mutant CP-HCP1 binding correlates well with the infectivity of the corresponding mutant viruses in barley. These results indicate that CP-HCP1 binding controls BMV infection of barley, interacting directly with CP, probably in the cell cytoplasm.
Okinaka
Yasushi
Y
Laboratory of Plant Pathology, Graduate School of Agriculture, Kyoto University, Kyoto 606-8502, Japan.
Mise
Kazuyuki
K
Okuno
Tetsuro
T
Furusawa
Iwao
I
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D001483:Base Sequence; D001665:Binding Sites; D017795:Bromovirus / Q000502:physiology*; D036022:Capsid Proteins / Q000502:physiology*; D003001:Cloning, Molecular; D001467:Hordeum / Q000235:genetics / Q000821:virology*; D008969:Molecular Sequence Data; D016296:Mutagenesis; D010940:Plant Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D011991:Receptors, Virus / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism
D036022:Capsid Proteins; D010940:Plant Proteins; D011991:Receptors, Virus; D011994:Recombinant Proteins