Phosphorylation of tobacco mosaic virus cell-to-cell movement protein by a developmentally regulated plant cell wall-associated protein kinase.
In host plants, cell-to-cell spread of tobacco mosaic virus (TMV) presumably occurs through intercellular connections, the plasmodesmata. TMV movement is mediated by a specific virus-encoded single-strand nucleic acid-binding protein, P30. The mechanism by which P30 operates is largely unknown. Here, we demonstrate that P30 expressed in transgenic plants is a phosphoprotein. We have developed an assay for in vitro phosphorylation of purified P30 by plant cell wall fractions and have localized the phosphorylation sites to amino acid residues Ser-258, Thr-261, and Ser-265. Interestingly, the P30 phosphorylation sites do not correspond to any known consensus phosphorylation sites for protein kinases. While P30 binding to single-stranded DNA (ssDNA) was shown to involve Thr-261, phosphorylation of this residue does not appear to play a role in binding activity. The protein kinase activity contained in the cell wall fractions was developmentally regulated, expressed predominantly in leaves. Within a leaf, this protein kinase activity increased with leaf maturation and correlated with the reported development of secondary plasmodesmata, sites of P30 accumulation. We suggest that phosphorylation may represent a mechanism for the host plant to sequester P30 following its localization to cell walls.
Citovsky
V
V
Department of Plant Biology, University of California, Berkeley 94720.
McLean
B G
BG
Zupan
J R
JR
Zambryski
P
P
D000595:Amino Acid Sequence; D002473:Cell Wall / Q000201:enzymology; D004252:DNA Mutational Analysis; D004277:DNA, Single-Stranded / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli; D015967:Gene Expression Regulation, Viral; D005796:Genes / Q000502:physiology*; D005814:Genes, Viral / Q000502:physiology; D008969:Molecular Sequence Data; D010766:Phosphorylation; D053830:Plant Viral Movement Proteins; D030821:Plants, Genetically Modified; D010947:Plants, Toxic; D011494:Protein Kinases / Q000502:physiology*; D012313:RNA / Q000378:metabolism; D011994:Recombinant Proteins; D012694:Serine / Q000737:chemistry; D013912:Threonine / Q000737:chemistry; D014026:Nicotiana / Q000201:enzymology* / Q000254:growth & development / Q000648:ultrastructure; D014027:Tobacco Mosaic Virus / Q000378:metabolism*; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism*
D004277:DNA, Single-Stranded; D053830:Plant Viral Movement Proteins; D011994:Recombinant Proteins; D014764:Viral Proteins; D013912:Threonine; D012694:Serine; D012313:RNA; D011494:Protein Kinases