Infectivity of okra enation leaf curl virus and the role of its V2 protein in pathogenicity

Abstract

Cotton crop has been severely affected by multiple begomoviruses in Pakistan and India. In our previous study, we found okra enation leaf curl virus (OELCuV), cotton leaf curl Multan betasatellite (CLCuMuB) and cotton leaf curl Multan alphasatellite (CLCuMuA) infecting cotton in Pakistan. The current study was designed to investigate the infectivity of OELCuV and its ability to trans-replicate non-cognate CLCuMuB. Agro-infectious clones containing the partial tandem repeats of OELCuV and CLCuMuB were constructed and the infectivity assays were carried out through Agrobacterium mediated transformation in the model host species Nicotiana benthamiana under controlled conditions. The results showed that in the inoculated plants OELCuV alone can cause downward curling and yellowing of leaves with thickened veins. However, when co-inoculated with the non-cognate CLCuMuB it could functionally trans-replicate CLCuMuB resulting in a more severe phenotype. The expression of Pre-coat/V2 protein in the N. benthamiana plants through the potato virus X (PVX) system caused localized cell death after severe leaf curling in the infiltrated leaves. The tissue tropism of the virus was associated with the systemic development of a hypersensitive response (HR), which ultimately lead to the plant death. The results indicated the involvement of V2 protein in the pathogenicity of OELCuV and its ability to trigger the host defense machinery. This study also demonstrated the ability of OELCuV to trans-replicate CLCuMuB resulting in typical leaf curl disease symptoms in N. benthamiana.

Characterization of Pathogenicity-Associated V2 Protein of Tobacco Curly Shoot Virus

“…Some reports showed that V2 proteins encoded by some geminiviruses could substantially function in viral pathogenicity [43,55,[57][58][59]. To investigate the role of V2 encoded by TbCSV in virus infection, V2 coding sequence was inserted into and heteroexpressed by potato virus X.…”

Differences in the 3’ intergenic region and the V2 protein of two sequence variants of tomato curly stunt virus play an important role in disease pathology in Nicotiana benthamiana

“…Two hydrophobic domains (H1 and H2) and the putative protein kinase C (PKC) phosphorylation motif which have been characterised previously [ 31 , 34 , 35 ] were mapped to this variable N-terminal region ( Fig 1C ). The C-terminal half of V2 contained a CxC motif [ 31 , 33 ] conserved between the two V2 proteins whilst the V2 of V22 also contained three additional C-terminal residues (Gln-Lys-Pro) absent in V30 V2 ( Fig 1C ).…”

“…We observed that serine substitution of V2 Thr58 resulted in delayed symptom development. V2 aa positions 58 to 60 constitute the PKC motif which has been reported to have a consensus sequence of Thr-X-Arg, where X is any residue [ 31 , 33 , 35 ]. Alanine substitution of the conserved PKC motif residues previously showed that these residues are required for the subcellular localisation, pathogenicity-enhancing, and RNA silencing suppression activities of several geminivirus V2 proteins [ 34 , 35 , 89 ].…”

Artificial microRNA-mediated resistance against Oman strain of tomato yellow leaf curl virus

“…Once the optimal density (OD 600 ) of bacterium was achieved, the bacterial cells were propagated in 10 ml MgCl 2 (10 mM) and acetosyringone (100 mM) incubated at 25-28°C for 3-4 hours before inoculation. The inoculum was subverted into the leaves of N. benthamiana from the underside by using a 5/10 ml needleless injection syringe, as reported earlier ( Saeed et al., 2018 ; Shafiq et al., 2021 ). The inoculated plants were maintained in an insect-free glasshouse with 16/8 hours day/night at 28°C.…”