Maintenance of coat protein N-terminal net charge and not primary sequence is essential for zucchini yellow mosaic virus systemic infectivity.
Zucchini yellow mosaic virus (ZYMV) surface exposed coat protein (CP) N-terminal domain (Nt) is 43 aa long and contains an equal number of positively and negatively charged amino acid residues (CP-Nt net charge = 0). A ZYMV-AGII truncation mutant lacking the first 20 aa of its CP-Nt (AGII-CP Delta 20; CP-Nt net charge = +2) was found to be systemically non-infectious even though AGII mutants harbouring larger CP-Nt deletions were previously demonstrated to be fully infectious. Nevertheless, AGII-CP Delta 20 infectivity was restored by fusion to its CP-Nt two Asp residues or a negatively charged Myc peptide, both predicted to neutralize CP-Nt net positive charge. To evaluate further the significance of CP-Nt net charge for AGII infectivity, a series of CP-Nt net charge mutants was generated and analysed for systemic infectivity of squash plants. AGII-CP(KKK) harbouring a CP-Nt amino fusion of three Lys residues (CP-Nt net charge = +3) was not systemically infectious. Addition of up to four Asp residues to CP-Nt did not abolish virus infectivity, although certain mutants were genetically unstable and had delayed infectivity. Addition of five negatively charged residues abolished infectivity (AGII-CP(DDDDD); CP-Nt net charge = -5) even though a recombinant CP(DDDDD) could assemble into potyviral-like particle in bacteria. Neutralization of CP-Nt net charge by fusing Asp or Lys residues recovered infectivity of AGII-CP(KKK) and AGII-CP(DDDDD). GFP-tagging of these mutants has demonstrated that both viruses have defective cell-to-cell movement. Together, these findings suggest that maintenance of CP-Nt net charge and not primary sequence is essential for ZYMV infectivity.
Department of Ornamental Horticulture, Agricultural Research Organization, The Volcani Center, PO Box 6, Bet Dagan 50250, Israel.
Boaz
B
Kimalov
Department of Virology, Agricultural Research Organization, The Volcani Center, PO Box 6, Bet Dagan 50250, Israel.
Amit
A
Gal-On
Department of Ornamental Horticulture, Agricultural Research Organization, The Volcani Center, PO Box 6, Bet Dagan 50250, Israel.
Ran
R
Stav
Horticulture Institute, Agricultural Research Organization, The Volcani Center, PO Box 6, Bet Dagan 50250, Israel.
Eduard
E
Belausov
Department of Ornamental Horticulture, Agricultural Research Organization, The Volcani Center, PO Box 6, Bet Dagan 50250, Israel.
Tzahi
T
Arazi
D000595:Amino Acid Sequence; D036022:Capsid Proteins / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics; D028464:Cucurbita / Q000821:virology*; D008969:Molecular Sequence Data; D009154:Mutation; D010935:Plant Diseases / Q000821:virology*; D017800:Potyvirus / Q000737:chemistry* / Q000472:pathogenicity; D017434:Protein Structure, Tertiary; D016415:Sequence Alignment; D014774:Virulence
D036022:Capsid Proteins; C086081:coat protein, Zucchini yellow mosaic virus