Temporal analysis of the regulatory activities of small RNAs during the cell-to-cell spread of tobamovirus infection

Project leader: PD Dr. Manfred Heinlein, Dr. Franck Vazquez

Project Team members: Dr. Khalid Amari

Plant viruses spread their genomes through infected tissues by cell-to-cell movement through plasmodesmata (PD) and replication in each infected cell. During spread the viruses encounter the efficient host RNA silencing antiviral defense response. As a counter-defence viruses generally protect their genomes by producing specialized RNA silencing suppressor proteins. Strikingly, several plant viruses, like Tobacco mosaic virus (TMV), encode proteins that enhance RNA silencing, rather than block it. This indicates that interaction of the virus with the host silencing machinery is more complex than previously thought. In the case of TMV, the silencing suppressor acts behind the spreading infection front whereas at the front the movement protein (MP) of the virus spreads small RNAs as well as the viral genome. This complex mechanism potentially manipulates host gene expression and enhances cell susceptibility before invasion.

To gain insight into this process, we use RNA deep sequencing and RNAseq combined with RNA degradome sequencing to profile small RNAs and corresponding mRNA targets during the spread of GFP-tagged TMV infection sites in Nicotiana benthamiana. Respective profiles of cells ahead, at, and behind the leading front of infection will be evaluated with the help of bioinformatics experts and verified by Northern blots, RACE analysis, and different reporter constructs. These data will provide important knowledge about the role of RNA silencing-mediated gene regulation during viral invasion of plants.