An Arabidopsis RNase III-like protein, AtRTL2, cleaves double-stranded RNA in vitro

Class 1 ribonuclease III (RNase III), found in bacteria and yeast, is involved in processing functional RNA molecules such as ribosomal RNAs (rRNAs). However, in Arabidopsis thaliana, the lack of an obvious phenotype or quantitative change in mature rRNAs in class 1 RNase III (AtRTL2) mutants and overexpressing plants suggests that AtRTL2 is not involved in rRNA maturation. We characterized the in vitro activity of AtRTL2 to consider its in vivo function. AtRTL2 cleaved double-stranded RNA (dsRNA) specifically in vitro, yielding products of approximately 25 nt or longer in length, in contrast to 10-20 nt long products in bacteria and yeasts. Although dsRNA-binding activity was not detected, the dsRNA-binding domains in AtRTL2 were essential for its dsRNA-cleaving activity. Accumulation of small RNAs derived from transgene dsRNAs was increased when AtRTL2 was transiently expressed in Nicotiana benthamiana leaves by agroinfiltration. These results raise the possibility that AtRTL2 has functions distinct from those of other class 1 RNase IIIs in vivo.