Dynamic response of RNA editing to temperature in Drosophila

Leila E Rieder 1 Yiannis A Savva 1 Matthew A Reyna 0 Yao-Jen Chang 1 Jacquelyn S Dorsky 1 Ali Rezaei 1 Robert A Reenan 1 0 Department of Mathematical Sciences, Rensselaer Polytechnic Institute , Troy, NY 12180 , USA 1 Department of Molecular Biology, Cellular Biology, and Biochemistry, Brown University , Providence, RI 02912 , USA - of RNA editing to Drosophila Rieder et al. Open Access Dynamic response of RNA editing to temperature in Drosophila Background: Adenosine-to-inosine RNA editing is a highly conserved process that post-transcriptionally modifies mRNA, generating proteomic diversity, particularly within the nervous system of metazoans. Transcripts encoding proteins involved in neurotransmission predominate as targets of such modifications. Previous reports suggest that RNA editing is responsive to environmental inputs in the form of temperature alterations. However, the molecular determinants underlying temperature-dependent RNA editing responses are not well understood. Results: Using the poikilotherm Drosophila, we show that acute temperature alterations within a normal physiological range result in substantial changes in RNA editing levels. Our examination of particular sites reveals diversity in the patterns with which editing responds to temperature, and these patterns are conserved across five species of Drosophilidae representing over 10 million years of divergence. In addition, we show that expression of the editing enzyme, ADAR (adenosine deaminase acting on RNA), is dramatically decreased at elevated temperatures, partially, but not fully, explaining some target responses to temperature. Interestingly, this reduction in editing enzyme levels at elevated temperature is only partially reversed by a return to lower temperatures. Lastly, we show that engineered structural variants of the most temperature-sensitive editing site, in a sodium channel transcript, perturb thermal responsiveness in RNA editing profile for a particular RNA structure. Conclusions: Our results suggest that the RNA editing process responds to temperature alterations via two distinct molecular mechanisms: through intrinsic thermo-sensitivity of the RNA structures that direct editing, and due to temperature sensitive expression or stability of the RNA editing enzyme. Environmental cues, in this case temperature, rapidly reprogram the Drosophila transcriptome through RNA editing, presumably resulting in altered proteomic ratios of edited and unedited proteins. Background Natural DNAs are usually limited to double-stranded helical shapes; however, RNA is different the repertoire of possible RNA secondary and tertiary structures appears limitless. RNA secondary structure is strongly correlated with function, and both the structure and corresponding thermodynamic stability of an RNA molecule contribute to functional regulation [1]. Dynamic RNA structures are acutely responsive to input in the form of molecular and environmental factors; it is this * Correspondence: 1Department of Molecular Biology, Cellular Biology, and Biochemistry, Brown University, Providence, RI 02912, USA Full list of author information is available at the end of the article mutability of RNA structure that allows RNA to act as a sensor and elicit rapid cellular responses [2]. RNA structure is fundamentally sensitive to abiotic factors, such as temperature and metal ion concentration. Bacterial RNA thermometers, riboswitches sensitive to temperature, are responsive regulatory elements that control translation of heat-shock, cold-shock, and virulence genes [3,4]. Yet, there is no direct evidence of translational RNA thermometers in eukaryotes. With the addition of large expanses of intronic sequence, eukaryotic RNA thermometers could be considerably less conserved than those found in bacteria, confounding detection. Indeed, the regulation of alternative splicing by the eukaryotic thymidine pyrophosphate riboswitch depends on complex long-distance nucleotide interactions [5]. 2015 Reenan et al.; licensee BioMed Central. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly credited. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. Therefore, temperature-sensitive structures found in eukaryotic mRNA could, in theory, act anywhere in the transcript to alter processing, translation, transport, degradation or protein binding. Adenosine-to-inosine (A-to-I) RNA editing is a posttranscriptional modification known to be directed by secondary [6] and tertiary RNA structures, including those that act over a distance of up to several thousand nucleotides [7,8]. We reasoned that there might be a class of eukaryotic RNA thermometer-like structures that, instead of controlling translation, rather exert their effects on RNA editing levels. A-to-I RNA editing involves the hydrolytic deamination of adenosine into inosine, which is read as guanosine by the protein synthesis machinery. Editing, therefore, has the ability to recode the transcriptome at select sites [9] and diversify the proteome. ADARs (adenosine deaminases acting on RNA), the highly conserved proteins responsible for A-to-I editing in all metazoa, are found localized to both cytoplasm and nucleus. While there are multiple ADAR proteins in mammals, the mammalian ADAR2 and single Drosophila ADAR (dADAR) appear to function primarily in the neuronal nucleus [10]. This is consistent with the observation that ADARs target transcripts encode proteins involved in chemical and electrical neurotransmission. Phenotypes in model organisms with editing deficiencies range from embryonic lethality [11] and seizures [12] (mouse) to defects in motor control [13] (Drosophila) and chemotaxis [14] (Caenorhabditis elegans). Such phenotypic consequences of ADAR deficiency indicate that editing plays an integral role in organismal behavior and viability. Some evidence suggests that environmental factors, specifically temperature, affect editing of select transcripts [15-17], and that RNA structure may regulate this relationship [18,19]. For example, Garrett and Rosenthal showed that editing in an octopus delayed rectifier potassium channel transcript correlates with ambient water temperature for different species, and even suggest that editing differences contribute to octopus adaptation [20]. Additionally, auto-editing of the Drosophila adar (dadar) transcript decreases as temperature increases. Given that the ratio of edited to unedited dADAR isoforms fine-tunes the global editosome [21] as well as complex organismal behavior, these data point toward an intriguing role of thermal control in global RNA editing [9]. However, little (...truncated)