Infection by tomato yellow leaf curl Sardinia virus (TYLCSV) alters the methylation profile of tomato genome.

~lanthsa ve developed several genetically regulated defense strategies against a wide range of biotic stresses (viruses, fungi, bacteria, nematodes). There is experimental evidence that DNA methylation and post-transcriptional gene silencing (PTGS) are strongly associated, and evolved as an important part of plant defense mechanisms against invasion of foreign nucleic acids, including vimses. In plants infected by the geminivirus Tornato yellow leaf curl Sardinia virus (TYLCSV), an important limiting factor for tomato production in the Mediterranean basin, recent studies have described the activation of PTGS, as evidenced by the presence of small interfering RNAs (siRNAs). Moreover, geminiviruses can interact with several host proteins, such as plant retinoblastoma-related protein (RBR), a fundamental ce11 cycle regulator, and proliferating nuclear ce11 antigen (PCNA), an essential component of the ce11 DNA replication machinery. In this respect, they have similarities to DNA tumor viruses infecting mammals. Since anima1 viruses can regulate host gene expression through genome methylation, we have investigated the methylation of tomato genome during infection by TYLCSV. A methylation-sensitive amplified polymorphism (MSAP) methodology was developed to compare the DNA methylation profiles of infected vs uninfected tomato plants (cv 'Moneymaker'). We have analysed the DNA extracted from leaves collected at different times (1, 7 and 14 days) following inoculation by the insect vector, the whitefly Bemisia tubaci. Ten polyrnorphic fragments were detected and sequenced following elution fiom gel and reamplification. Sequences obtained were analysed for similarity to known sequences in public databases available on the web: the Solanaceae Genomics Network (SGN) website, Solanaceae Genomics Database (SolGD) and the GenBank database (NCBI). Some of the differentially methylated genes appeared to be involved in the plant defence mechanism, i.e. cysteine protease, methionine adenosyltransferase, while others showed similarity to proteins containing the leucine-rich repeat (LRR) domain, a structural motif present in many resistance (R) proteins and associated with protein-protein interactions. The expression leve1 of these genes is currently being evaluated with Reverse Transcription-Quantitative PCR. This work confirms the potential of MSAP in identi@ing gene sequences possibly involved in the reaction of plants to virus infection.