Cap-snatching mechanism-mediated unveiling of the transcriptional initiation sites of two distinct begomoviruses

Significant economic losses are inflicted by plant viruses, which pose a risk to sustainable agriculture. The proliferation of novel viral diseases is predominantly attributable to factors such as climate change, international trade, and the rapid evolutionary capabilities of viruses. Begomoviruses are a major group of plant-infecting viruses that pose an imminent threat to global agriculture by causing devastating viral diseases in many crop species. The transcriptional start sites (TSSs) of many plant viruses are typically found in the intergenic region (IR), which is the non-coding (NC) area between the viral genes. The promoters play a crucial role in initiating the transcription process by aiding in the recruitment of cellular transcription machinery. The TSSs are precise nucleotide sequences where RNA polymerase initiates the transcription process. The primary objective of this study was to determine the total number of TSSs for two devastating begomoviruses, family: Geminiviridae, Cotton leaf curl Multan virus (CLCuMuV) and Ageratum yellow vein mosaic virus (AYVMV), using the cap-snatching method in conjunction with one heterologous plant virus. These two begomoviruses, along with their infectious clones, were intentionally infected with selected heterologous plant virus in N. benthamiana plants. The identification of the 5′ ends of heterologous viral mRNA was accomplished by employing high-throughput sequencing to assess the capped RNA leaders (CRLs). The determination of the 5′ termini of suspected begomoviral mRNAs was achieved by aligning the collected CRLs of heterologous virus with the genome of each begomovirus, taking into account only those that were a perfect match with the begomoviral genome. In this study, the TSSs of both begomoviruses were identified via complementary approach. The utilization of high-throughput sequencing for both begomoviruses has facilitated the acquisition of millions of sequences. Comprehending the TSSs and promoter components of begomoviruses is crucial for understanding their ability to cause disease, their interactions with host organisms, and for developing effective methods to manage and control the diseases, they inflict on economically significant crop plants.