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Comparative Ribosome Profiling Uncovers a Dominant Role for Translational Control in Toxoplasma Gondii

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Comparative Ribosome Profiling Uncovers a Dominant Role for Translational Control in Toxoplasma Gondii Comparative ribosome profiling uncovers a dominant role for translational control in Toxoplasma gondii The MIT Faculty has made this article openly available. Please share how this access benefits you. Your story matters. Citation Hassan, Musa A. et al. "Comparative ribosome profiling uncovers a dominant role for translational control in Toxoplasma gondii." BMC Genomics 18 (December 2017): 961 © 2017 The Author(s) As Published http://dx.doi.org/10.1186/s12864-017-4362-6 Publisher BioMed Central Version Final published version Citable link http://hdl.handle.net/1721.1/114431 Terms of Use Creative Commons Attribution Detailed Terms http://creativecommons.org/licenses/by/4.0/ Hassan et al. BMC Genomics (2017) 18:961 DOI 10.1186/s12864-017-4362-6 RESEARCH ARTICLE Open Access Comparative ribosome profiling uncovers a dominant role for translational control in Toxoplasma gondii Musa A. Hassan1,2*, Juan J. Vasquez3,8, Chew Guo-Liang4, Markus Meissner5,6 and T. Nicolai Siegel3,6,7 Abstract Background: The lytic cycle of the protozoan parasite Toxoplasma gondii, which involves a brief sojourn in the extracellular space, is characterized by defined transcriptional profiles. For an obligate intracellular parasite that is shielded from the cytosolic host immune factors by a parasitophorous vacuole, the brief entry into the extracellular space is likely to exert enormous stress. Due to its role in cellular stress response, we hypothesize that translational control plays an important role in regulating gene expression in Toxoplasma during the lytic cycle. Unlike transcriptional profiles, insights into genome-wide translational profiles of Toxoplasma gondii are lacking. Methods: We have performed genome-wide ribosome profiling, coupled with high throughput RNA sequencing, in intracellular and extracellular Toxoplasma gondii parasites to investigate translational control during the lytic cycle. Results: Although differences in transcript abundance were mostly mirrored at the translational level, we observed significant differences in the abundance of ribosome footprints between the two parasite stages. Furthermore, our data suggest that mRNA translation in the parasite is potentially regulated by mRNA secondary structure and upstream open reading frames. Conclusion: We show that most of the Toxoplasma genes that are dysregulated during the lytic cycle are translationally regulated. Keywords: Ribosome profiling, RNA-sequencing, Translation efficiency, Toxoplasma gondii, Apicomplexan Background translation, and protein degradation [5]. Consequently, All living organisms are constantly exposed to a variety of transcript abundance rarely mirrors cellular protein levels biological stress, which may include limiting nutrient [6]. Although the relative contribution of each of these availability, oxidative stress, temperature shock, DNA steps in the gene-expression pathway is equivocal, mRNA damage and infection. Consequently, organisms show re- translation is known to play a significant role in modulat- markable regulatory plasticity that allows them to thrive ing cellular protein levels [7, 8]. Indeed, translational con- under different, sometimes harsh, environmental condi- trol of gene expression is known to provide opportunities tions [1, 2]. Historically, due to the relative ease of obtain- for controlling spatial and temporal protein distribution ing global transcript abundance estimates, most studies [9]. Furthermore, because most eukaryotic mRNAs can be quantify fluctuations in mRNA abundance to gain insights detected in cells at least 2 h after expression [10], com- into organismal response to stress [3, 4]. However, the pared to de novo transcription, regulating the translation catalogue of expressed genes and proteins is modulated at of the available mRNAs provides a mechanism to rapidly various steps, including mRNA splicing, export, stability, adjust cellular protein levels in response to drastic changes in the environment or developmental stages [11, 12]. In * Correspondence: [email protected] fact, most translationally regulated mRNAs are known to 1Division of Infection and Immunity, The Roslin Institute, University of encode proteins that regulate important cellular processes Edinburgh, Edinburgh, UK 2The Centre for Tropical Livestock Genetics and Health, The Roslin Institute, such as stress response, development, and synaptic trans- University of Edinburgh, Edinburgh, UK mission [7]. Full list of author information is available at the end of the article © The Author(s). 2017 Open Access This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. 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. Hassan et al. BMC Genomics (2017) 18:961 Page 2 of 12 Toxoplasma gondii is an obligate intracellular apicom- regulatory serine (serine-71) are reported to exhibit de- plexan that infects virtually all warm-blooded verte- creased extracellular viability [26]. Lower expression brates. In the definitive feline host, Toxoplasma levels of eIF4, another translation initiation factor, has undergoes sexual recombination, but reverts to asexual been observed in bradyzoites and attenuated Toxo- reproduction in the intermediate host, which includes plasma strains [2, 23, 24, 26]. Finally, the endoplasmic humans. Asexual reproduction in Toxoplasma is charac- reticulum (ER) stress response in Toxoplasma is charac- terized by the rapidly dividing tachyzoite stage. However, terized by preferential translation of a subset of genes, in response to host-derived stress factors, such as im- including the transcriptional regulator AP2 [23, 27]. This mune response, the rapidly dividing tachyzoites convert is particularly important since the integrity of the para- to the semi-dormant encysted bradyzoites and establish site ER is pivotal for the proper folding of essential pro- lifelong chronic infections in the central nervous system teins required for parasite invasion, immune evasion and and muscle tissues of the host [13, 14]. Establishment of the establishment of chronic infection [24]. Thus, it is chronic infection is important for the re-entry of the plausible that the viability, pathogenesis, and transmission parasite into the definitive host, and for horizontal trans- of Toxoplasma are dependent on its ability to recognize mission within intermediate hosts, through the preda- and translationally respond to host-derived stress. tion and consumption of food products from chronically Genome-wide insights on translational control and the infected hosts, respectively [15]. The tachyzoite-to- underlying molecular factors that regulate mRNA trans- bradyzoite conversion reportedly mirrors a stress re- lation in Toxoplasma are largely unknown. Here, we ac- sponse [2] and does not only involve significant changes cess the translational landscape in Toxoplasma gondii in the parasite physiology and morphology, but also is and determine its impact on intercellular parasite trans- accompanied by altered gene expression profiles [16]. mission. To do this, we have used ribosome profiling to During the lytic cycle the parasite invades a host cell, capture genome-wide translational profiles of intracellu- replicates, and then lyses out of the host cell before in- lar and extracellular Toxoplasma parasites infecting hu- fecting a new host cell. This process temporarily exposes man foreskin fibroblasts. Our data reveal a putative role the parasite to the extracellular milieu. The extracellular for translational control in regulating parasite gene viability of the parasite is reported to decrease dramatic- expression during the lytic cycle. Additionally, our data ally between 6 and 12 h after egress [17], indicating the revealed variable translational efficiency of several dys- level of biological stress induced on the parasite by host regulated Toxoplasma mRNAs, such as the mRNAs en- factors. Indeed, transcriptional data on most Toxoplasma coding dense granules, which are known to be spatially strains have revealed stage-specific expression of several secreted during the lytic cycle. Finally, our data suggest genes, such as surface antigens, stress response genes, that that mRNA secondary structure, putatively affect virulence genes, and metabolic enzymes [13, 18, 19]. mRNA translation in Toxoplasma. These results not Consequently, regulating transcript abundance, and by only provide greater insights into Toxoplasma gene extension their protein products, is key in regulating regulation, but also provide a resource and template for Toxoplasma developmental stages and intercellular elucidating the function of translational control in transmission. Toxoplasma biology. Finally, the ribosome footprints, Translational regulation of gene expression has will provide an additional resource for annotating Toxo- emerged as a key factor in the biology of apicomplexan plasma transcript features. parasites [20–23]. In Plasmodium, translational regula- tion is reported to modulate stage conversion and host- Results parasite interactions [20, 21]. For example, while Pb2 Generation of mRNA profiles and ribosome footprints
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