Open AccessA Single dicer Gene Is Required for Efficient Gene Silencing Associated with Two Classes of Small Antisense RNAs in Mucor circinelloides
Author(s) -
Juan P. de Haro,
Silvia Calo,
María Cervantes,
Francisco E. Nicolás,
Santiago Torres-Martı́nez,
Rosa M. RuizVázquez
Publication year - 2009
Publication title -
eukaryotic cell
Language(s) - English
Resource type - Journals
eISSN - 1535-9778
pISSN - 1535-9786
DOI - 10.1128/ec.00191-09
Subject(s) - dicer , mucor circinelloides , biology , gene silencing , rna silencing , rna interference , trans acting sirna , rna induced silencing complex , small interfering rna , sense (electronics) , genetics , argonaute , gene , microbiology and biotechnology , rna , mucor , chemistry , aspergillus
RNA silencing in the zygomyceteMucor circinelloides exhibits uncommon features, such as induction by self-replicative sense transgenes and the accumulation of two size classes of antisense small interfering RNAs (siRNAs). To investigate whether this silencing phenomenon follows the rules of a canonical RNA-silencing mechanism, we used hairpin RNA (hpRNA)-producing constructs as silencing triggers and analyzed the efficiency and stability of silencing in different genetic backgrounds. We show here that the dsRNA-induced silencing mechanism is also associated with the accumulation of two sizes of antisense siRNAs and that this mechanism is not mediated by the previously knowndcl-1 (dicer -like) gene, which implies the existence of an additionaldicer gene. AnM. circinelloides dcl-2 gene was cloned and characterized, and the corresponding null mutant was generated by gene replacement. This mutant is severely impaired in the silencing mechanism induced by self-replicative sense or inverted-repeat transgenes, providing the first genetic evidence of a canonical silencing mechanism in this class of fungus and pointing to a role fordcl-2 in the mechanism. Moreover, a functionaldcl-2 gene is required for the normal accumulation of the two sizes of antisense RNAs, as deduced from the analysis ofdcl-2 − transformants containing hpRNA-expressing plasmids. In addition to its critical role in transgene-induced silencing, thedcl-2 gene seems to play a role in the control of vegetative development, since thedcl-2 null mutants showed a significant decrease in their production of asexual spores.