FOXO regulates RNA interference in Drosophila and protects from RNA virus infection

Abstract

Small RNA pathways are important players in posttranscriptional regulation of gene expression. These pathways play important roles in all aspects of cellular physiology from development to fertility to innate immunity. However, almost nothing is known about the regulation of the central genes in these pathways. The forkhead box O (FOXO) family of transcription factors is a conserved family of DNAbinding proteins that responds to a diverse set of cellular signals. FOXOs are crucial regulators of cellular homeostasis that have a conserved role in modulating organismal aging and fitness. Here, we show that Drosophila FOXO (dFOXO) regulates the expression of core small RNA pathway genes. In addition, we find increased dFOXO activity results in an increase in RNA interference (RNAi) efficacy, establishing a direct link between cellular physiology and RNAi. Consistent with these findings, dFOXO activity is stimulated by viral infection and is required for effective innate immune response to RNA virus infection. Our study reveals an unanticipated connection among dFOXO, stress responses, and the efficacy of small RNA-mediated gene silencing and suggests that organisms can tune their gene silencing in response to environmental and metabolic conditions.O rganisms must be able to respond to changing conditions to survive. Radical changes in environment or metabolism can induce cellular stress signaling pathways meant to help the cell return to a normal range of function. Whereas there are dedicated transcriptional response pathways to deal with specific individual stresses such as heat shock (HSF) (1) or heavy metals (MTF-1) (2), the forkhead box O (FOXO) pathway has emerged as a general stress responsive transcription factor. Although initially characterized as downstream targets of insulin signaling, it is now clear that the FOXOs respond to multiple cellular stress signaling pathways including nutrient deprivation, oxidative stress, mitochondrial dysfunction, DNA damage, bacterial infection, and other cellular stress signals (3, 4).The FOXO family of transcription factors is conserved from Caenorhabditis elegans to humans. Invertebrates have a single FOXO gene: daf-16 in worms and dFOXO in Drosophila (5, 6). In mammals, the four FOXO genes (FOXO1, FOXO3, FOXO4, and FOXO6) have diversified, with some factors having a stressspecific response (7-10). Therefore, invertebrates offer a particularly attractive system for understanding FOXO function because the single isoform is responsible for all of the roles of the four mammalian FOXOs.Small noncoding RNA pathways are capable of regulating gene expression in a sequence-specific manner (11). Effector complexes called RNA-induced silencing complexes (RISC) containing small single-stranded RNAs bound to Argonaute proteins find target mRNAs and posttranscriptionally prevent their expression. Three major small RNA pathways have been identified in animal cells, the microRNA (miRNA) pathway, the small interfering RNA pathway (siRNA), and PIWI RNA (piRNA) pathways, each contain uni...

SAGA complex mediates the transcriptional up-regulation of antiviral RNA silencing

“…Their identification using the currently developed screen protocol is underway. In this regard, it is noteworthy that a basic leucine zipper-type and a forkhead box O-type transcription factor have been shown to regulate RNA silencing genes in plant and insect, respectively (29,46). It will also be of interest to further explore whether SAGA, conserved across eukaryotic organisms from yeast to humans, serves as an RNAi regulator in these organisms and what viral factors determine the virus-specific RNAi activation and requirement of AGL2 for RNAi induction.…”

“…Mechanisms of its transcriptional regulation remain largely unknown but appear to be regulated differently in different organisms (5,6,29,30). We have developed a genetic screen protocol using an infectious cDNA clone of a CHV1 mutant, Δp69, which lacks a viral suppressor of RNA silencing (VSR), p29, and induces the key RNAi genes excessively (Fig.…”

The impact of FOXO on dopamine and octopamine metabolism in Drosophila under normal and heat stress conditions

“…FOXO activity is significantly higher in D. melanogaster larvae with genetically ablated corpora allata , the organ that produces JH (Mirth et al, 2014). Overexpression of dFOXO increases oxidative stress tolerance (Giannakou et al, 2004; Hwangbo et al, 2004) while dFOXO-null mutants are more sensitive to oxidative stress (Jünger et al, 2003) and deficient in viral defence (Spellberg and Marr, 2015). …”

Defense Mechanisms against Viral Infection in <em>Drosophila</em>: RNAi versus Non-RNAi

“…The requirement for base-pairing prior to initiation of degradation provides great specificity and mechanisms for enhancement of efficiency have also evolved such as amplification via DNA forms and production of secondary siRNAs followed by systemic spread via exosomes [74,75]. On the other hand, efficiency of antiviral RNAi can be affected by physiological conditions such as nutritional status and stress [19,168]. Furthermore, during persistent viral infections, the RNAi pathway may be partially dismantled or may function in different ways that are not completely understood [206].…”

“…When energy levels are low and insulin/insulin-like growth factor signaling is reduced, the forkhead transcription factor dFOXO responds by translocation to the nucleus resulting in increased target gene expression [18]. It was observed that induction of dFOXO in transgenic flies results in increased expression of the RNAi machinery genes ago-1, ago-2 and dcr-2 and concomitant resistance to virus infection [19]. In dFOXO null flies, the greater susceptibility to RNA viruses can be rescued by over-expression of Dcr-2.…”