New Levels of Transcriptome Complexity at Upper Thermal Limits in WildDrosophilaRevealed by Exon Expression Analysis

Abstract

While the cellular heat-shock response has been a paradigm for studying the impact of thermal stress on RNA metabolism and gene expression, the genome-wide response to thermal stress and its connection to physiological stress resistance remain largely unexplored. Here, we address this issue using an array-based exon expression analysis to interrogate the transcriptome in recently established Drosophila melanogaster stocks during severe thermal stress and recovery. We first demonstrated the efficacy of exon-level analyses to reveal a level of thermally induced transcriptome complexity extending well beyond gene-level analyses. Next, we showed that the upper range of both the cellular and physiological thermal stress response profoundly affected message expression and processing in D. melanogaster, limiting expression to a small subset of transcripts, many that share features of known rapidly responding stress genes. As predicted from cellular heat-shock research, constitutive splicing was blocked in a set of novel genes; we did not detect changes to alternative splicing during heat stress, but rather induction of intronless isoforms of known heatresponsive genes. We observed transcriptome plasticity in the form of differential isoform expression during recovery from heat shock, mediated by multiple mechanisms including alternative transcription and alternative splicing. This affected genes involved in DNA regulation, immune response, and thermotolerance. These patterns highlight the complex nature of innate transcriptome responses under stress and potential for adaptive shifts through plasticity and evolved genetic responses at different hierarchical levels.T HE distribution of ectotherms including Drosophila species can often be linked to their physiological thermal tolerances (Addo-Bediako et al. 2000;Mitchell and Hoffmann 2010;Kellermann et al. 2012). Terrestrial Drosophila from a range of environments may exist close to their maximal range and be constrained to increase upper tolerance limits, posing a threat to persistence under climate warming (Kellermann et al. 2012). Elucidating the factors delimiting upper thermal limits depends on understanding how physiological responses link with the underlying molecular processes in an integrative framework. Limited progress toward this end has been made so far, despite the cellular reaction to heat stress being the most ubiquitous and well-characterized molecular stress response.Seminal work exploiting the tightly controlled conditions of homogeneous cell lines has led to fine-scale molecular dissections of the heat-shock response in wide-ranging taxa including yeast, Drosophila, and humans. At the cellular level, heat shock triggers a dramatic reprogramming of gene expression to favor the rapid turnover of a class of molecular chaperones known as the Heat-shock proteins (Hsps) (Lindquist and Craig 1988;Yost et al. 1990). Apart from the selective activation of a subset of genes predominantly harboring heat-shock factor (HSF) sequence-binding elements, trans...

Biological invasions, climate change and genomics

“…These sources of variation for climate adaptation are only just starting to be explored (Telonis‐Scott et al. ).…”

“…For instance, in D. melanogaster, the transcriptome can be modified qualitatively in a variety of ways, such as through alternative splicing and expression of alternative isoforms, that may ultimately have more impact on plastic responses than quantitative changes in gene expression. These sources of variation for climate adaptation are only just starting to be explored (Telonis-Scott et al 2013).…”

Drosophila Small Heat Shock Proteins: An Update on Their Features and Functions

“…Ten out of the 12 sHsps are heat-inducible; only CG14207 is not and no data is available on CG43851 (Landis et al 2012 ;Telonis-Scott et al 2013 ;Vos 2009 ). The set of sHsps induced by the heat shock response is stage-dependent; while heatshocked embryos express hsp22 , hsp26 , hsp27 and hsp67Bc (Leemans et al 2000 ), 10 days-old fl ies additionally express hsp67Ba , CG7409 and l ( 2 ) efl (Landis et al 2012 ).…”

“…The set of sHsps induced by the heat shock response is stage-dependent; while heatshocked embryos express hsp22 , hsp26 , hsp27 and hsp67Bc (Leemans et al 2000 ), 10 days-old fl ies additionally express hsp67Ba , CG7409 and l ( 2 ) efl (Landis et al 2012 ). CG4461 , hsp23 , hsp26 , hsp27 and hsp67Bc are part of the early heatresponding genes in fl ies and are also the more abundant shsp transcripts following heat-shock in S2 cells (Telonis-Scott et al 2013 ;Vos 2009 ). The heat-shock response is driven by the heat shock factor (Hsf), which has been shown to bind tightly to hsp22 , hsp23 , hsp26 , hsp27 , hsp67Ba and hsp67Bc promoter while the CG4461 promoter was apparently not bound (Birch-Machin et al 2005 ).…”

Spatial analysis of gene regulation reveals new insights into the molecular basis of upper thermal limits

“…Telonis‐Scott et al . (), however, demonstrated that stv transcript isoforms are modulated in markedly different ways in response to heat stress, whereby the shorter isoforms underpinned the strong transcriptional response following heat shock. Interestingly, at least a subset of the shorter isoforms showed weak evidence of RNA processing during heat shock which is unusual given that hyperthermia largely inhibits pre‐mRNA splicing, a process bypassed in the majority of intron‐lacking Hsps (Yost & Lindquist ; Bond ; Lindquist & Craig ).…”