Molecular characterisation of integrated sequences of banana streak virus in the banana plant genome to intend plant vaccination

The genome of banana (Musa sp.) harbours multiple integrations of several species of Banana streak virus (BSV), certainly resulting from illegitimate recombination between host and viral DNA. Surprisingly, this badnavirus does not require integration for its replication. Some integrations, only existing in the Musa balbisiana genome (denoted B), are infectious by releasing a functional viral genome following stresses such as in vitro culture and interspecific crosses. To date, four widespread species of BSV (Goldfinger - BSGFV, Imové - BSImV, Mysore - BSMysV and Obino l'Ewai - BSOLV) have been reported as integrated into the B genome with three of them as infectious (eBSGFV, eBSImV and eBSOLV). In order to study BSV expression from such viral integrants, a characterisation of BSV integrants (eBSV) was undergone i) by studying a Musa BAC library obtained from the wild diploid M. balbisiana cv. Pisang Klutuk Wulung (PKW) containing the four eBSV species described above and known as eBSV carrier, ii) by using the cytological FISH technique and iii) an interspecific genetic cross using PKW as virus-free female parent. Very low copies of integrations were recorded for each BSV species. The full characterisation of eBSGFV was recently performed in our lab (Gayral et al., 2008). eBSGFV results from a single event of integration corresponding to an allelic insertion of at least one full-length viral genome extensively rearranged with several viral regions duplicated. Although the four BSV species present important differences with each other, the organisation of eBSOLV and eBSImV looks like eBSGFV. Indeed, each of them is more or less extensively rearranged in PKW and is present as allelic insertions at the same locus. In contrary, the non infectious eBSMysV presents two independent insertions sites. Interestingly, M. balbisiana diploid genotypes (BB) such as Pisang Klutuk Wulung (PKW) harbor infectious eBSVs in their genome but are nevertheless resistant to any multiplication of BSV. The mechanisms underlying such resistance are believed to be driven by epigenetic phenomena but no evidence has been obtained so far in banana plants. In collaboration with the group headed by M. Pooggin (Basel, Switzerland), we have obtained, for the first time, experimental evidence of virus-derived small RNA in PKW. Deep sequencing of the population of siRNAs using the Illumina ultra-high-throughput technology is on the way. This would help us to detect RNA molecules of very low bundance to single-nucleotide resolution and create a complete map of eBSV siRNAs. This deep-sequencing analysis is done by FASTERIS AG, a Swiss member of our COST action. (Texte intégral