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

Abstract

Pathogen recognition and transcriptional activation of defense-related genes are crucial steps in cellular defense responses. RNA silencing (RNAi) functions as an antiviral defense in eukaryotic organisms. Several RNAi-related genes are known to be transcriptionally up-regulated upon virus infection in some host organisms, but little is known about their induction mechanism. A phytopathogenic ascomycete, <i>Cryphonectria parasitica</i> (chestnut blight fungus), provides a particularly advantageous system to study RNAi activation, because its infection by certain RNA viruses induces the transcription of dicer-like 2 (<i>dcl2</i>) and argonaute-like 2 (<i>agl2</i>), two major RNAi players. To identify cellular factors governing activation of antiviral RNAi in <i>C. parasitica</i>, we developed a screening protocol entailing multiple transformations of the fungus with cDNA of a hypovirus mutant lacking the RNAi suppressor (CHV1-Δp69), a reporter construct with a GFP gene driven by the <i>dcl2</i> promoter, and a random mutagenic construct. Screening for GFP-negative colonies allowed the identification of <i>sgf73</i>, a component of the SAGA (Spt-Ada-Gcn5 acetyltransferase) complex, a well-known transcriptional coactivator. Knockout of other SAGA components showed that the histone acetyltransferase module regulates transcriptional induction of <i>dcl2</i> and <i>agl2</i>, whereas histone deubiquitinase mediates regulation of <i>agl2</i> but not <i>dcl2</i> Interestingly, full-scale induction of <i>agl2</i> and <i>dcl2</i> by CHV1-Δp69 required both DCL2 and AGL2, whereas that by another RNA virus, mycoreovirus 1, required only DCL2, uncovering additional roles for DCL2 and AGL2 in viral recognition and/or RNAi activation. Overall, these results provide insight into the mechanism of RNAi activation.