The C-terminal region of the 2a protein and 2b protein of cucumber mosaic virus are involved in the induction of shoestring-like leaf blade in tomato.

Cucumber mosaic virus (CMV) has numerous strains with distinct pathological properties in nature. In this study, we focused on the distinct host-specificity of two isolates of CMV regarding induction of the shoestring-like leaf blade (SLB) in tomato (Solanum lycopersicum cv. Sekaiichi). During the initial infection stage, plants inoculated with CMV-D8 and CMV-Y developed green/yellow systemic mosaic and stunting. Late in infection, CMV-D8 caused severe systemic symptoms with SLB on the newly emerged leaves, whereas CMY-Y caused severe yellow mosaic with stunting. Accumulation of viral RNA of CMV-D8 during initial infection was higher than for CMV-Y, but their levels did not differ significantly at 5 weeks post inoculation. Pseudorecombination and recombination analyses between CMV-D8 and CMV-Y genomic RNAs showed that recombinant that contained the C-terminal region of 2a and the entire 2b protein of CMV-D8 (D2a-C/D2b) induced SLB. Changing isoleucine to valine at position 830 in the 2a ORF played an important role in formation of chronic SLB. We further elucidated that infection with CMV-D8 or the recombinant Y1Y2(D2a-C/D2b)D3, but not with CMV-Y, upregulated miRNAs and transcript levels of AGO1, which is involved in RNA silencing, and of HD-ZIP, TCP4, and PHAN, which are essential for leaf morphogenesis. The present results first demonstrated that the cooperative function of D2a-C/D2b is involved indispensably in SLB formation. In addition, we suggest that D2a-C/D2b region interferes with the miRNA pathway that is associated with RNA silencing and leaf morphogenesis, leading to the enhanced virulence of CMV-D8.

The 2b protein and C-terminal region of the 2a protein indispensably facilitate systemic movement of cucumber mosaic virus in radish with supplementary function by either the 3a or the coat protein.

BACKGROUND: In Raphanus sativus (Japanese radish), strain D8 of cucumber mosaic virus (CMV-D8) establishes a systemic infection and induces mild mosaic on upper, non-inoculated leaves, whereas strain Y of CMV (CMV-Y) causes only a local infection in the inoculated leaves. Here, we further analyzed the specific viral factor(s) of CMV-D8 that is (are) indispensable for systemic infection in Japanese radish. METHODS: To identify which genomic RNA(s) is (are) involved in systemic infection in radish, we carried out a pseudorecombination analysis between CMV-D8 and CMV-Y. With recombination analyses between CMV-D8 and CMV-Y using mutant/recombinant RNA2s, chimeric and point-mutated RNA3s, we identified viral factors that are indispensable for systemic infection. RESULTS: Viral RNA2 and RNA3 of CMV-D8 facilitated efficient virus spread into the upper, non-inoculated plant tissues of radish (cv. Tokinashi), but not those of CMV-Y. Recombinant RNA2s demonstrated that the 2b protein (2b) and the C-terminus of the 2a protein (2a) of CMV-D8 have a crucial role in systemic infection. In addition, we used chimeric and point-mutated RNA3s to that Pro17 and Pro129 in the coat protein (CP) of CMV-D8 are involved in efficient systemic infection and that Ser51 in the 3a protein (3a) of CMV-D8 has positive effects on systemic spread. The results suggested that these viral factors facilitate systemic infection of CMV-D8 in Japanese radish. CONCLUSION: The C-terminal region of 2a, the entire region of 2b, and supplementary function of either Ser51 in 3a or Pro17/Pro 129 in CP confer systemic infectivity on CMV-D8 in radish. These results further elucidate the complex interaction of viral proteins of CMV to complete systemic infection as a host-specific manner.