Temperature-responsive miRNAs in Drosophila orchestrate adaptation to different ambient temperatures

Abstract

The majority of genes are expressed in a temperature-dependent manner, but the way in which small RNAs may contribute to this effect is completely unknown as we currently lack an idea of how small RNA transcriptomes change as a function of temperature. Applying high-throughput sequencing techniques complemented by quantitative real-time PCR experiments, we demonstrate that altered ambient temperature induces drastic but reversible changes in sequence composition and total abundance of both miRNA and piRNA populations. Further, mRNA sequencing reveals that the expression of miRNAs and their predicted target transcripts correlates inversely, suggesting that temperature-responsive miRNAs drive adaptation to different ambient temperatures on the transcriptome level. Finally, we demonstrate that shifts in temperature affect both primary and secondary piRNA pools, and the observed aberrations are consistent with altered expression levels of the involved Piwi-pathway factors. We further reason that enhanced ping-pong processing at 29°C is driven by dissolved RNA secondary structures at higher temperatures, uncovering target sites that are not accessible at low temperatures. Together, our results show that small RNAs are an important part of epigenetic regulatory mechanisms that ensure homeostasis and adaptation under fluctuating environmental conditions.

Environmentally-induced epigenetic conversion of a piRNA cluster

“…We have tested if such stresses were able to convert BX2 OFF into BX2 ON in one generation but neither heat-shock nor osmotic stress induces BX2 conversion (TABLE S9), suggesting that BX2 activation does not depend on dATF-2. In a more recent paper, Fast et al (34) found less piRNAs at 29°C than at 18°C. However, RNAseq analyses of differentially expressed genes involved in the piRNA pathway were not conclusive, as some piRNA genes were more expressed at 29°C (ago3, aub, zuc, armi) while others were less expressed (shu, hsp83, Yb) (34).…”

“…In a more recent paper, Fast et al (34) found less piRNAs at 29°C than at 18°C. However, RNAseq analyses of differentially expressed genes involved in the piRNA pathway were not conclusive, as some piRNA genes were more expressed at 29°C (ago3, aub, zuc, armi) while others were less expressed (shu, hsp83, Yb) (34). Overall, the enhancement of the piRNA pingpong amplification loop observed at 29°C was attributed to RNA secondary structures, because of a lack of specificity for any particular class of TE (34).…”

“…However, RNAseq analyses of differentially expressed genes involved in the piRNA pathway were not conclusive, as some piRNA genes were more expressed at 29°C (ago3, aub, zuc, armi) while others were less expressed (shu, hsp83, Yb) (34). Overall, the enhancement of the piRNA pingpong amplification loop observed at 29°C was attributed to RNA secondary structures, because of a lack of specificity for any particular class of TE (34). Furthermore, in contrast to our RT-qPCR results obtained at 25°C and 29°C (FIG 4D), AGO1 was not differentially expressed between 18°C and 29°C (34).…”

“…Overall, the enhancement of the piRNA pingpong amplification loop observed at 29°C was attributed to RNA secondary structures, because of a lack of specificity for any particular class of TE (34). Furthermore, in contrast to our RT-qPCR results obtained at 25°C and 29°C (FIG 4D), AGO1 was not differentially expressed between 18°C and 29°C (34). This difference can be explained, because, in our experiments, we checked for specific spliced transcripts of AGO1 that originate upstream the BX2 insertion point (FIG 4C).…”

Temperature-dependent small RNA expression in Drosophila melanogaster

“…Based on the observation that temperature-responsive genes were enriched for miRNA target sites it was further assumed that miRNAs could play a critical role in temperature-dependent gene regulation. To test this hypothesis, we have recently sequenced and analyzed ovary expressed small RNAs and mRNA from Drosophila cohorts kept at 18 C and 29 C. We further subjected each cohort to a temperature shift from 18-to-29 C or 29 C-to-18 C, respectively, to investigate the dynamics of putative expression changes [4]. We were able to verify profound and reversible changes in miRNA expression patterns and additionally showed that the expression levels of temperature-responsive miRNAs and their predicted target genes correlate inversely ( Fig.…”