Identification of domains of the Tomato spotted wilt virus NSm protein involved in tubule formation, movement and symptomatology

Deletion and alanine-substitution mutants of the Tomato spotted wilt virus NSm protein were generated to identify domains involved in tubule formation, movement and symptomatology using a heterologous Tobacco mosaic virus expression system. Two regions of NSm, G19-S159 and G209-V283, were required for both tubule formation in protoplasts and cell-to-cell movement in plants, indicating a correlation between these activities. Three amino acid groups, D154, EYKK205-208 and EEEEE284-288 were linked with long-distance movement in Nicotiana benthamiana. EEEEE284-288 was essential for NSm-mediated long-distance movement, whereas D154 was essential for tubule formation and cell-to-cell movement; indicating separate genetic controls for cell-to-cell and long-distance movement. The region I57-N100 was identified as the determinant of foliar necrosis in Nicotiana benthamiana, and mutagenesis of HH93-94 greatly reduced necrosis. These findings are likely applicable to other tospovirus species, especially those within the 'New World' group as NSm sequences are highly conserved.