Cellular targets of functional and dysfunctional mutants of tobacco mosaic virus movement protein fused to green fluorescent protein.
Intercellular transport of tobacco mosaic virus (TMV) RNA involves the accumulation of virus-encoded movement protein (MP) in plasmodesmata (Pd), in endoplasmic reticulum (ER)-derived inclusion bodies, and on microtubules. The functional significance of these interactions in viral RNA (vRNA) movement was tested in planta and in protoplasts with TMV derivatives expressing N- and C-terminal deletion mutants of MP fused to the green fluorescent protein. Deletion of 55 amino acids from the C terminus of MP did not interfere with the vRNA transport function of MP:GFP but abolished its accumulation in inclusion bodies, indicating that accumulation of MP at these ER-derived sites is not a requirement for function in vRNA intercellular movement. Deletion of 66 amino acids from the C terminus of MP inactivated the protein, and viral infection occurred only upon complementation in plants transgenic for MP. The functional deficiency of the mutant protein correlated with its inability to associate with microtubules and, independently, with its absence from Pd at the leading edge of infection. Inactivation of MP by N-terminal deletions was correlated with the inability of the protein to target Pd throughout the infection site, whereas its associations with microtubules and inclusion bodies were unaffected. The observations support a role of MP-interacting microtubules in TMV RNA movement and indicate that MP targets microtubules and Pd by independent mechanisms. Moreover, accumulation of MP in Pd late in infection is insufficient to support viral movement, confirming that intercellular transport of vRNA relies on the presence of MP in Pd at the leading edge of infection.
Boyko
V
V
Friedrich Miescher Institute, CH-4058 Basel, Switzerland.
van der Laak
J
J
Ferralli
J
J
Suslova
E
E
Kwon
M O
MO
Heinlein
M
M
D001692:Biological Transport; D004721:Endoplasmic Reticulum / Q000378:metabolism / Q000648:ultrastructure; D049452:Green Fluorescent Proteins; D002479:Inclusion Bodies / Q000378:metabolism; D008164:Luminescent Proteins / Q000378:metabolism*; D008870:Microtubules / Q000502:physiology; D053830:Plant Viral Movement Proteins; D012367:RNA, Viral / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism*; D013329:Structure-Activity Relationship; D014027:Tobacco Mosaic Virus / Q000378:metabolism*; D014764:Viral Proteins / Q000378:metabolism*; D014779:Virus Replication
D008164:Luminescent Proteins; D053830:Plant Viral Movement Proteins; D012367:RNA, Viral; D011993:Recombinant Fusion Proteins; D014764:Viral Proteins; D049452:Green Fluorescent Proteins