Tomato Brown Rugose Fruit Virus Evades Tm‐2 ² ‐Mediated Resistance by Avoiding the Induction of Tm‐2 ² Self‐Association

Tomato brown rugose fruit virus (ToBRFV) overcomes Tm-2²-mediated resistance, posing a serious threat to global tomato production. We previously identified that key residues in the movement protein (MP)-H67, N125, K129, A134, I147 and I168 are essential for ToBRFV evasion of Tm-2² resistance. However, the underlying mechanisms remain unclear. In Tm-2²-transgenic Nicotiana benthamiana plants at later infection stages, MP mutations H67C, N125A, K129Q and A134N abolished ToBRFV-GFP cell-to-cell and systemic movement, whereas I168N abolished ToBRFV-GFP systemic but not cell-to-cell movement, and I147M partially impaired both. Correspondingly, the MPH67C, MPN125A, MPK129Q and MPA134N mutants elicited strong Tm-2²-mediated hypersensitive response (HR), MPI168N elicited mild HR and MPI147M elicited none. Confocal microscopy revealed I147M reduced MP localisation at plasmodesmata in Tm-2²-transgenic N. benthamiana leaves. The interaction with Tm-2² was weak for wild-type MP and I147M (slightly enhanced), but moderately or strongly enhanced for I168N and the other four mutants. Wild-type MP and MPI147M failed to induce Tm-2² self-association, MPI168N only weakly induced it, whereas the other four markedly induced it. Collectively, these findings demonstrate that ToBRFV evades Tm-2²-mediated resistance by attenuating MP-Tm-2² interaction strength, thereby avoiding Tm-2² self-association and downstream immune activation.