Genome-wide identification and analysis of small RNAs originated from natural antisense transcripts in <i>Oryza sativa</i>

Natural antisense transcripts (NATs) have been shown to play important roles in post-transcriptional regulation through the RNA interference pathway. We have combined pyrophosphate-based high-throughput sequencing and computational analysis to identify and analyze, in genome scale, cis -NAT and trans -NAT small RNAs that are derived under normal conditions and in response to drought and salt stresses in the staple plant Oryza sativa . Computationally, we identified 344 cis -NATs and 7142 trans -NATs that are formed by protein-coding genes. From the deep sequencing data, we found 108 cis -NATs and 7141 trans -NATs that gave rise to small RNAs from their overlapping regions. Consistent with early findings, the majority of these 108 cis -NATs seem to be associated with specific conditions or developmental stages. Our analyses also revealed several interesting results. The overlapping regions of the cis -NATs and trans -NATs appear to be more enriched with small RNA loci than non-overlapping regions. The small RNAs generated from cis -NATs and trans -NATs have a length bias of 21 nt, even though their lengths spread over a large range. Furthermore, >40% of the small RNAs from cis -NATs and trans -NATs carry an A as their 5′-terminal nucleotides. A substantial portion of the transcripts are involved in both cis -NATs and trans -NATs, and many trans -NATs can form many-to-many relationships, indicating that NATs may form complex regulatory networks in O. sativa . This study is the first genome-wide investigation of NAT-derived small RNAs in O. sativa . It reveals the importance of NATs in biogenesis of small RNAs and broadens our understanding of the roles of NAT-derived small RNAs in gene regulation, particularly in response to environmental stimuli.