Small RNA profiles of wild-type and silencing suppressor-deficient tomato spotted wilt virus infected Nicotiana benthamiana

Comparison of small RNA profiles in Nicotiana benthamiana and Solanum lycopersicum infected by polygonum ringspot tospovirus reveals host-specific responses to viral infection

“…Leaf tissues from three different plants of each species, which did not show statistically significant difference in virus titer, were pooled and used for RNA extraction. Total RNA was extracted using "Directzol RNA prep" kit (Zymoresearch, Irvine, CA, USA), as previously described (Margaria et al, 2015a). RNA integrity and quantity were verified by 2% agarose gel electrophoresis, and using the NanoDrop 2000 Spectrophotometer (Thermoscientific, Waltham, MA, USA).…”

“…Host-specific profiles related to vsRNA size, preferential targeting of genomic segments and single ORFs, and vsRNA polarity were observed. Moreover, a comparison of PolRSV vsRNA profiles with the ones observed for tomato spotted wilt virus (TSWV) isolates infecting the same hosts (Mitter et al, 2013;Margaria et al, 2015a), showed significant differences between the two viral species.…”

Tospovirus : induction and suppression of RNA silencing

“…These observations altogether support the idea that during viral infections NSs likely may bind to stretches of dsRNA present within the IGR-encoded putative hairpin structure to prevent their recognition and cleavage by DCL and slicer (Ago) from the RISC complex. Deep sequencing analysis of small RNAs from TSWV infected plants showed that the low production levels of siRNAs from the IGR sequence is not only observed with the ambisense S RNA but also with the ambisense M RNA segment (Margaria et al, 2015a), and provides further support for the above idea. This interaction is also supported by previous work in which this 3'-UTR structure was shown to act as a functional equivalent of a poly(A)-tail in chimeric mRNA molecules and co-expression with NSs lead to translational enhancement of the corresponding gene (Geerts-Dimitriadou et al, 2012).…”

“…Although speculative, similar to hantavirus N (Panganiban and Mir, 2009), the tospovirus N protein might bind to the cap-structure of messenger RNAs and act as an eIF4F surrogate (Merrick, 2015). In further support for the idea that the 3'-UTR interacts with more proteins other than NSs only, is a recent observation that the 3'-UTR encoded-hairpin sequence is weakly targeted by RNA silencing even in silencing suppressor-defective isolates (NSs-defective) (Margaria et al, 2015a).…”

“…While currently more than 20 distinct tospovirus species have been defined, eight of which confirmed by the ICTV (Pappu et al, 2009;King et al, 2012), research on tospovirus silencing suppression is primarily limited to TSWV NSs. Silencing-suppressor defective (NSs-mutant) TSWV strains show a clear increase of viral siRNAs (vsiRNAs), specifically of the 21 nt class (Margaria et al, 2015a). Besides being able to suppress local RNA silencing, NSs TSWV has also been briefly reported to suppress systemic silencing (Takeda et al, 2002).…”