Validation of microtubule-associated Tobacco mosaic virus RNA movement and involvement of microtubule-aligned particle trafficking

Functional studies of Tobacco mosaic virus (TMV) infection using virus derivatives expressing functional,dysfunctional, and temperature-sensitive movement protein (MP) mutants indicated that the cell-to-celltransport of TMV RNA is functionally correlated with the association of MP with microtubules. However, therole of microtubules in the movement process during early infection remains unclear, since MP accumulates onmicrotubules rather late in infection and treatment of plants with microtubule-disrupting agents fails tostrongly interfere with cell-to-cell movement of TMV RNA. To further test the role of microtubules in TMV cellto-cell movement, we investigated TMV strain Ni2519, which is temperature-sensitive for movement. Wedemonstrate that the temperature-sensitive defect in movement is correlated with temperature-sensitivechanges in the localization of MP to microtubules. Furthermore, we show that during early phases of recoveryfrom non-permissive conditions, the MP localizes to microtubule-associated particles. Similar particles arefound in cells at the leading front of spreading TMV infection sites. Initially mobile, the particles becomeimmobile when MP starts to accumulate along the length of the particle-associated microtubules. Ourobservations confirm a role for microtubules in the spread of TMV infection and associate this role withmicrotubule-associated trafficking of MP-containing particles in cells engaged in the cell-to-cell movement ofthe TMV genome.