Immunological detection of the 8K protein of potato virus X (PVX) in cell walls of PVX-infected tobacco and transgenic potato

Abstract

Open reading frame 4 (ORF4) of the potato virus X (PVX) genome encodes an 8K protein which is a part of the "triple gene block" and is known to play a role in the cell-to-cell movement of the virus in infected plants. To locate the 8K protein and further elucidate the mechanism of cell-to-cell transport of PVX, antibodies were raised against the 8K protein and used to localize this protein in PVX-infected tobacco and in transgenic potato plants expressing the 8K protein both by subcellular fractionation and by immunolabeling with colloidal gold. The results indicated that the 8K protein was localized to the cell wall.

Structural Properties of Carnation Mottle Virus p7 Movement Protein and Its RNA-binding Domain

“…Using synthetic peptides derived from the p7 sequence, we have identified three distinct putative domains within the protein. EMSA showed that the central region, from residue 17 to 35 (represented by peptide p7 [17][18][19][20][21][22][23][24][25][26][27][28][29][30][31][32][33][34][35] ), is responsible for the RNA binding properties of CarMV p7. This binding peptide populates a nascent ␣-helix in water solution that is further stabilized in the presence of either secondary structure inducers, such as trifluoroethanol and monomeric SDS, or RNA (which also changes its conformation upon binding to the peptide).…”

“…There are viruses that encode multiple MPs, which, in addition, do not fall in the 30K superfamily. The triple gene block of potexviruses encodes three proteins (open reading frames 2-4) that mediate cell-tocell transport, the first one with RNA-binding and NTPase/ helicase activities and the other two with putative membranespanning domains and, in the case of the third, demonstrated cell wall localization (22,23). Carmoviruses are among the smallest known plant viruses; their genome is a singlestranded RNA of ϳ4 kilobases encoding at least five proteins (24,25).…”

“…The concentration of all of the other peptides was obtained by quantitative amino acid analysis. The CD spectra of the p7 [17][18][19][20][21][22][23][24][25][26][27][28][29][30][31][32][33][34][35] -RNA complex were recorded at 5°C with 20 M of p7 [17][18][19][20][21][22][23][24][25][26][27][28][29][30][31][32][33][34][35] , and increasing concentrations of CarMV ssRNA transcribed in vitro from clone pCarM.D5 (Ref. 29; this RNA covers exactly the first 181 residues of the CarMV genome) were added (from 1 to 20 ng), until no change in the difference spectrum was observed.…”

“…Spectra were recorded in the far UV region in 5 mM MOPS/NaOH, pH 7.0, buffer. In peptide tritation experiments, increasing concentrations of p7 [17][18][19][20][21][22][23][24][25][26][27][28][29][30][31][32][33][34][35] were added (from 0 to 200 M) to a 1 mM RNA solution and left on ice for 30 min before the CD spectrum was recorded in the near UV region (250 -350 nm).…”

Arg-16 and Arg-21 in the N-terminal region of the triple-gene-block protein 1 of Bamboo mosaic virus are essential for virus movement

“…TGBp2 and TGBp3 both contain stretches of hydrophobic amino acids and are associated with cell wall and membrane fractions both in vivo and in vitro (Donald et al, 1993;Hefferon et al, 1997;Morozov et al, 1990Morozov et al, , 1991NiesbachKlosgen et al, 1990;Solovyev et al, 2000). Using a microinjection and transient expression strategies, TGBp2 was shown to assist the movement of the ribonucleoprotein complex of potexviruses (Lough et al, 1998) and to be capable of increasing the SEL of the plasmodesmata (Tamai & Meshi, 2001).…”