Deep sequencing analysis reveals a TMV mutant with a poly(A) tract reduces host defense responses in Nicotiana benthamiana

- TMV-43A is a TMV mutant in which the UPD region was substituted by a 43 nt poly(A) tract.
- TMV-43A generated less virus-derived small interfering RNA (vsiRNA) than that of TMV in N. benthamiana.
- vsiRNAs produced from the UPD region of TMV were predicted to target several host defense response genes.
- Expression of genes involved in small RNA biogenesis was lower in TMV-43-infected N. benthamiana.
- Expression of host response genes was significantly lower in TMV-43-infected N. benthamiana.

Tobacco mosaic virus (TMV) possesses an upstream pseudoknotted domain (UPD), which is important for replication. After substituting the UPD with an internal poly(A) tract (43 nt), a mutant TMV-43A was constructed. TMV-43A replicated slower than TMV and induced a non-lethal mosaic symptom in Nicotiana benthamiana. In this study, deep sequencing was performed to detect the differences of small RNA profiles between TMV- and TMV-43A-infected N. benthamiana. The results showed that TMV-43A produced lesser amount of virus-derived interfering RNAs (vsiRNAs) than that of TMV. However, the distributions of vsiRNAs generation hotspots between TMV and TMV-43A were similar. Expression of genes related to small RNA biogenesis in TMV-43A-infected N. benthamiana was significantly lower than that of TMV, which leads to generation of lesser vsiRNAs. The expressions of host defense response genes were up-regulated after TMV infection, as compared to TMV-43A-infected plants. Host defense response to TMV-43A infection was lower than that to TMV. The absence of UPD might contribute to the reduced host response to TMV-43A. Our study provides valuable information in the role of the UPD in eliciting host response genes after TMV infection in N. benthamiana. (187 words)