Submitted to: Proceedings of the National Academy of Sciences
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 7/5/2005
Publication Date: 5/13/2005
Citation: Williams, L., Carles, C.C., Osmont, K.S., Fletcher, J.C. 2005. A database analysis method identifies an endogenous trans-acting short-interfering RNA that targets the Arabidopsis ARF2, ARF3, and ARF4 genes. Proceedings of the National Academy of Sciences. 102(27):9703-9708. Interpretive Summary: Here we describe a computational method to uncover new plant small RNAs within a database of Arabidopsis sequences. We identified a novel non-coding small regulatory RNA we called tasi-ARF. We found three genes encoding tasi-ARF in the Arabidopsis genome, and showed that the sequences are conserved in rice and maize. We showed that tasi-ARF is complementary to three Auxin Response Factor (ARF) messenger RNAs, enabling the tasi-ARF to target the ARF mRNAs for destruction. Our method therefore proves to be useful for identifying additional small regulatory RNAs.
Technical Abstract: Two classes of small RNAs, microRNAs and short-interfering RNA (siRNAs), have been extensively studied in plants and animals. In Arabidopsis, the capacity to uncover previously uncharacterized small RNAs by means of conventional strategies seems to be reaching its limits. To discover new plant small RNAs, we developed a protocol to mine an Arabidopsis nonannotated, noncoding EST database. Using this approach, we identified an endogenous small RNA, trans-acting short-interfering RNA-auxin response factor (tasiR-ARF), that shares a 21- and 22-nt region of sequence similarity with members of the ARF gene family. tasiR-ARF has characteristics of both short-interfering RNA and microRNA, recently defined as tasiRNA. Accumulation of trans-acting siRNA depends on DICER-LIKE1 and RNA-DEPENDENT RNA POLYMERASE6 but not RNA-DEPENDENT RNA POLYMERASE2. We demonstrate that tasiR-ARF targets three ARF genes, ARF2, ARF3/ETT, and ARF4, and that both the tasiR-ARF precursor and its target genes are evolutionarily conserved. The identification of tasiRNA-ARF as a low-abundance, previously uncharacterized small RNA species proves our method to be a useful tool to uncover additional small regulatory RNAs.