Properties of satellite tobacco mosaic virus phenotypes expressed in the presence and absence of helper virus

•When co-expressed with helper virus in N. benthamiana but not in N. tabacum, STMV generates a truncated RNA.•A highly conserved 3′ stem-loop structure is obligatory for STMV translation.•STMV is competent to form empty VLPs without helper virus.•STMV assembly requires a 3′ tRNA-like structure.

In this study, we assembled an Agrobacterium-based transient expression system for the ectopic expression of Satellite tobacco mosaic virus (STMV) (+) or (−) transcripts and their biological activity was confirmed when Nicotiana benthamiana plants were co-expressed with helper Tobacco mosaic virus replicase. Characterization of STMV in the presence and absence of its HV revealed: (i) HV-dependent expression of STMV (+) in N. benthamiana, but not in N. tabacum, generated a replication-deficient but translation and encapsidation competent variant lacking the highly conserved 3′ 150 nucleotides (nt) (STMVΔ150); (ii) mutational analysis demonstrated that a conserved 3′ stem-loop structure in wild type and STMVΔ150 located between nt 874 and 897 is essential for translation of CP; (iii) helper virus-independent expression of CP from wt STMV was competent for the assembly of empty aberrant virion-like particles; whereas, CP translated from STMVΔ150 resulted in disorganized CP aggregates suggesting a role for the 3′tRNA-like structure in STMV assembly.