Abstract
The small interfering RNA (siRNA) pathway of Drosophila melanogaster, mainly characterized by the activity of the enzymes Dicer 2 (Dcr-2) and Argonaute 2 (Ago-2), has been described as the major antiviral immune response. Several lines of evidence demonstrated its pivotal role in conferring resistance against viral infections at cellular and systemic level. However, only few studies have addressed the regulation and induction of this system upon infection and knowledge on stability and turnover of the siRNA pathway core components transcripts and proteins remains scarce. In the current work, we explore whether the siRNA pathway is regulated following viral infection in D. melanogaster. After infecting different fly strains with two different viruses and modes of infection, we observed changes in Dcr-2 and Ago-2 protein concentrations that were not related with changes in gene expression. This response was observed either upon viral infection or upon stress-related experimental procedure, indicating a bivalent function of the siRNA system operating as a general gene regulation rather than a specific antiviral system.
Highlights
RNA interference (RNAi) is a defensive and gene regulatory process based in sequence homology among nucleic acids [1,2,3]
In the small interfering RNA (siRNA) pathway, the ribonuclease Dicer 2 (Dcr-2) recognizes and dices double stranded RNA molecules of exogenous or endogenous origin, producing 21-nucleotide length siRNA duplexes that are loaded into the protein Argonaute 2 (Ago-2) within the RNAinduced silencing complex (RISC)
To study the regulation of the siRNA pathway during viral infections we used different D. melanogaster strains, viruses and modes of infection in order to establish the commonality of the response
Summary
RNA interference (RNAi) is a defensive and gene regulatory process based in sequence homology among nucleic acids [1,2,3]. Small RNAs (sRNAs) are produced and used as guides to target complementary DNA or RNA sequences [4, 5]. Three main sRNA pathways are described to date, differing in the origin and biogenesis of the double-stranded sRNAs and their molecular function: the micro RNA pathway (miRNA), the small interfering RNA pathway (siRNA), and the Piwi-interacting RNA pathway (piRNA) [6]. In the siRNA pathway, the ribonuclease Dicer 2 (Dcr-2) recognizes and dices double stranded RNA molecules of exogenous (virus) or endogenous (cellular) origin, producing 21-nucleotide length siRNA duplexes that are loaded into the protein Argonaute 2 (Ago-2) within the RNAinduced silencing complex (RISC). The siRNA duplexes are unwound and only one RNA strand is used by Ago-2 to target and slice the complementary RNA. Two Dcr-2 cofactors are indispensable for siRNA production and correct loading into RISC: LOQS and R2D2 [4, 6]
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