Inhibition of RNA Interference and Modulation of Transposable Element Expression by Cell Death in Drosophila

Abstract

RNA interference (RNAi) regulates gene expression by sequence-specific destruction of RNA. It acts as a defense mechanism against viruses and represses the expression of transposable elements (TEs) and some endogenous genes. We report that mutations and transgene constructs that condition cell death suppress RNA interference in adjacent cells in Drosophila melanogaster. The reversal of RNAi is effective for both the white (w) eye color gene and green fluorescent protein (GFP), indicating the generality of the inhibition. Antiapoptotic transgenes that reverse cell death will also reverse the inhibition of RNAi. Using GFP and a low level of cell death produced by a heat shock-head involution defective (hs-hid) transgene, the inhibition appears to occur by blocking the conversion of double-stranded RNA (dsRNA) to short interfering RNA (siRNA). We also demonstrate that the mus308 gene and endogenous transposable elements, which are both regularly silenced by RNAi, are increased in expression and accompanied by a reduced level of siRNA, when cell death occurs. The finding that chronic ectopic cell death affects RNAi is critical for an understanding of the application of the technique in basic and applied studies. These results also suggest that developmental perturbations, disease states, or environmental insults that cause ectopic cell death would alter transposon and gene expression patterns in the organism by the inhibition of small RNA silencing processes.

Rapid selection against arbovirus-induced apoptosis during infection of a mosquito vector

“…Thus, induction of apoptosis in large numbers of cells, both infected and uninfected, likely weakened the physical midgut barrier. Immune defenses of the mosquitoes may have also been disrupted; for example, one of the main immune responses against arbovirus infection, RNA interference, was previously shown to be inhibited by apoptosis in Drosophila (47). This combination of factors likely was responsible for the increase in SINV infection observed when widespread apoptosis was stimulated by Aeiap1 knockdown before infection (33).…”

Reflections on the inhibition of RNAi by cell death signaling

“…Neither was there any evidence for an effect on the inhibition of RNAi nor for an increased expression of ADAR in the hs-hid strain. 1 Thus, these tests proved negative for an involvement of ADAR in the RNAi inhibition. If there were a modification of the dsRNA that prevents it from entering into the RNAi pathway, the basis of such modification would need to be further investigated.…”

“…1 This situation was first realized by combining an RNAi construct for the white eye color gene with the Bar eye mutation. A strip of red color was present in the eye behind the anterior region of the Bar eye.…”

“…Reversing the action of the cell death transgenes also reversed the inhibition of RNAi. 1 The effects could be extended to RNAi for Green Fluorescent Protein (GFP) illustrating the generality of the effects. Nevertheless, in the GFP RNAi strain we found no reason to suspect that programmed cell death as part of normal development had any impact on RNAi.…”

“…1 This construct would cause specks of red in the white eye color RNAi flies and a generalized return of the expression of GFP in larvae and flies that otherwise had RNAi operating on GFP. This stock permitted an analysis on the molecular level.…”