Complementation of Virus Movement in Transgenic Tobacco Expressing the Cucumber Mosaic Virus 3a Gene

Abstract

Tobacco plants were transformed with the cucumber mosaic cucumovirus (CMV) 3a gene and the in planta-expressed 3a protein was detected immunologically. The 3a protein was predominantly localized in a subcellular fraction corresponding to the cytosol. Two frameshift and four deletion mutants were created within the 3a open reading frame of CMV RNA 3. Five of these mutants, containing an N-terminal, large central, or C-terminal 70-amino-acid deletion could not infect nontransformed tobacco plants, but could infect the 3a transgenic tobacco plants, and generally accumulated to wild-type levels. The sixth mutant, lacking the C-terminal 43 amino acids of the 3a protein, was able to infect nontransformed tobacco plants. A delay in accumulation of viral RNA in both the inoculated and the systemically infected leaves was demonstrated for one of the mutants. Thus, the CMV 3a protein is a virus movement protein, the functions of which can be complemented in a transgenic plant. The CMV 3a transgenic plants were able to complement the long-distance movement of a pseudorecombinant cucumovirus defective for this function in tobacco, as well as the cell-to-cell, but not the long-distance, movement of two other related viruses. However, these transgenic plants were unable to complement the long-distance movement of viruses from several other taxonomic groups that could move cell to cell but not long distance in tobacco.

Conversion in the Requirement of Coat Protein in Cell-to-Cell Movement Mediated by the Cucumber Mosaic Virus Movement Protein

“…This study demonstrates that the CMV MP truncated in its C terminus can promote viral cell-to-cell movement indepen- (19,48), (ii) increase plasmodesmal size exclusion limit (15,48), (iii) bind single-stranded nucleic acids in vitro (10,27), and (iv) complement their respective movement-deficient mutants (13,23). In contrast to these similarities, CMV MP cannot promote viral cell-to-cell movement without its cognate CP, although TMV MP can.…”

“…It is noteworthy that the function of CMV MP to mediate viral cell-to-cell movement was abolished when only three amino acids were deleted from its C terminus. A movement-incapable CMV variant with the C-terminal deletion of 43 amino acids corresponds to a CMV mutant that infects tobacco plants systemically (23). To test this, a CMV variant was created by the manner identical to that for Kaplan's mutant.…”

The C-terminal 33 amino acids of the cucumber mosaic virus 3a protein affect virus movement, RNA binding and inhibition of infection and translation

“…The cell-to-cell and long-distance movement of Cucumber mosaic virus (CMV) involve the participation of each of the five encoded proteins of CMV, expressed from the three genomic and two subgenomic RNAs (Ding et al, 1995a, b;Gal-On et al, 1994;Hellwald & Palukaitis, 1995;Kaplan et al, 1995;Suzuki et al, 1991). Nevertheless, the 3a protein, expressed from RNA 3 of CMV, is designated the virus movement protein (MP), since this protein is involved in a number of functions associated with movement (Canto & Palukaitis, 1999a, b;Ding et al, 1995a;Gal-On et al, 1995;Kaplan et al, 1995;Li & Palukaitis, 1996;Li et al, 2001).…”

“…Nevertheless, the 3a protein, expressed from RNA 3 of CMV, is designated the virus movement protein (MP), since this protein is involved in a number of functions associated with movement (Canto & Palukaitis, 1999a, b;Ding et al, 1995a;Gal-On et al, 1995;Kaplan et al, 1995;Li & Palukaitis, 1996;Li et al, 2001). The 3a MP binds ssRNA cooperatively (Li & Palukaitis, 1996), forming tightly packed nucleoprotein complexes (Nurkiyanova et al, 2001).…”

“…Different cell types can form barriers to the movement of RNA by various mutants of the 3a MP (Canto & Palukaitis, 1999a, b;Kaplan et al, 1997). The 3a MP expressed in transgenic plants can complement the movement of various, but not all, 3a protein mutants (Canto et al, 1997;Kaplan et al, 1995;Li et al, 2001). …”