A database analysis method identifies an endogenous trans-acting short-interfering RNA that targets the Arabidopsis ARF2, ARF3, and ARF4 genes.
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.
Williams
Leor
L
United States Department of Agriculture-Agricultural Research Service, Plant Gene Expression Center, 800 Buchanan Street, Albany, CA 94710, USA.
Carles
Cristel C
CC
Osmont
Karen S
KS
Fletcher
Jennifer C
JC
D017360:Arabidopsis / Q000235:genetics*; D029681:Arabidopsis Proteins / Q000235:genetics* / Q000378:metabolism; D001483:Base Sequence; D018797:Cell Cycle Proteins / Q000378:metabolism; D019295:Computational Biology / Q000379:methods*; D017931:DNA Primers; D004268:DNA-Binding Proteins / Q000235:genetics*; D030541:Databases, Genetic; D008969:Molecular Sequence Data; D005810:Multigene Family / Q000235:genetics*; D009687:Nuclear Proteins / Q000235:genetics*; D021141:Nucleic Acid Amplification Techniques; D034741:RNA, Small Interfering / Q000235:genetics*; D012324:RNA-Dependent RNA Polymerase / Q000378:metabolism; D034743:RNA-Induced Silencing Complex / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D043244:Ribonuclease III / Q000378:metabolism; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA
D029681:Arabidopsis Proteins; D018797:Cell Cycle Proteins; D017931:DNA Primers; D004268:DNA-Binding Proteins; C109765:ETT protein, Arabidopsis; D009687:Nuclear Proteins; D034741:RNA, Small Interfering; D034743:RNA-Induced Silencing Complex; D012324:RNA-Dependent RNA Polymerase; C402383:DCL1 protein, Arabidopsis; D043244:Ribonuclease III