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1. Quality checking and expression analysis of high-throughput small RNA sequencing data

Author

Beckers, Matthew

Subjects
572.8
Abstract

The advent of high-throughput RNA sequencing (RNA-seq) methods have made it possible to sequence transcriptomes for the cell-wide identi�cation of small non-coding RNAs (sRNAs) and to assess their regulation using di�erential expression analysis by comparing two or more di�erent conditions. During an analysis of a typical set of sRNA sequencing (sRNA-seq) libraries, a large variety of tools and methods are used on the dataset in order to understand the data's quality, content, and to summarise the knowledge gained from the entire analysis. Many of the tools available to do this were created for mRNA sequencing (mRNA-seq) datasets. In this thesis, we present and implement a processing pipeline that can be used to assess the quality and the di�erential expression of sRNA-seq datasets over two or more di�erent conditions. We then utilise aspects of this pipeline in various sRNA-seq experiments. Firstly, we combine our pipeline with current tools for miRNA identi�cation to assess the regulation of miRNAs during larval caste di�erentiation in a novel genome; the European bumblebee (Bombus terrestris). Secondly, we explore the di�erential expression during cell stress of all classes of sRNAs using two cell lines in humans. We also �nd that a speci�c protein, Ro60, is required for the expression of mRNA-derived sRNAs during stress, similar to the way in which sRNAs derived from Y RNAs are regulated. Finally, we utilise our understanding of sRNA mapping patterns, alongside current tools for miRNA identi�cation, to search for functional miRNAs and other sRNAs in the novel genomes of two diatoms. The lack of canonical miRNA predictions in this study has repercussions for the evolutionary theory behind miRNAs. The implementation of our pipeline for sRNA-seq data provides an interactive and quality controlled work ow that can be used to process a dataset from raw sequences to the results of several di�erential expression experiments for all identi�ed sRNA classes within a sequenced transcriptome.

Published
2015

2. The genomes of two key bumblebee species with primitive eusocial organization

Author

Sadd, Ben M., Barribeau, Seth M., Bloch, Guy, de Graaf, Dirk C., Dearden, Peter, Elsik, Christine G., Gadau, Jürgen, Grimmelikhuijzen, Cornelis J. P., Hasselmann, Martin, Lozier, Jeffrey D., Robertson, Hugh M., Smagghe, Guy, Stolle, Eckart, Van Vaerenbergh, Matthias, Waterhouse, Robert M., Bornberg-Bauer, Erich, Klasberg, Steffen, Bennett, Anna K., Câmara, Francisco, Guigó, Roderic, Hoff, Katharina, Mariotti, Marco, Munoz-Torres, Monica, Murphy, Terence, Santesmasses, Didac, Amdam, Gro V., Beckers, Matthew, Beye, Martin, Biewer, Matthias, Bitondi, Márcia M. G., Blaxter, Mark L., Bourke, Andrew F. G., Brown, Mark J. F., Buechel, Severine D., Cameron, Rossanah, Cappelle, Kaat, Carolan, James C., Christiaens, Olivier, Ciborowski, Kate L., Clarke, David F., Colgan, Thomas J., Collins, David H., Cridge, Andrew G., Dalmay, Tamas, Dreier, Stephanie, du Plessis, Louis, Duncan, Elizabeth, Erler, Silvio, Evans, Jay, Falcon, Tiago, Flores, Kevin, Freitas, Flávia C. P., Fuchikawa, Taro, Gempe, Tanja, Hartfelder, Klaus, Hauser, Frank, Helbing, Sophie, Humann, Fernanda C., Irvine, Frano, Jermiin, Lars S., Johnson, Claire E., Johnson, Reed M., Jones, Andrew K., Kadowaki, Tatsuhiko, Kidner, Jonathan H., Koch, Vasco, Köhler, Arian, Kraus, F. Bernhard, Lattorff, H. Michael G., Leask, Megan, Lockett, Gabrielle A., Mallon, Eamonn B., Antonio, David S. Marco, Marxer, Monika, Meeus, Ivan, Moritz, Robin F. A., Nair, Ajay, Näpflin, Kathrin, Nissen, Inga, Niu, Jinzhi, Nunes, Francis M. F., Oakeshott, John G., Osborne, Amy, Otte, Marianne, Pinheiro, Daniel G., Rossié, Nina, Rueppell, Olav, Santos, Carolina G., Schmid-Hempel, Regula, Schmitt, Björn D., Schulte, Christina, Simões, Zilá L. P., Soares, Michelle P. M., Swevers, Luc, Winnebeck, Eva C., Wolschin, Florian, Yu, Na, Zdobnov, Evgeny M., Aqrawi, Peshtewani K., Blankenburg, Kerstin P., Coyle, Marcus, Francisco, Liezl, Hernandez, Alvaro G., Holder, Michael, Hudson, Matthew E., Jackson, LaRonda, Jayaseelan, Joy, Joshi, Vandita, Kovar, Christie, Lee, Sandra L., Mata, Robert, Mathew, Tittu, Newsham, Irene F., Ngo, Robin, Okwuonu, Geoffrey, Pham, Christopher, Pu, Ling-Ling, Saada, Nehad, Santibanez, Jireh, Simmons, DeNard, Thornton, Rebecca, Venkat, Aarti, Walden, Kimberly Ko, Wu, Yuan-Qing, Debyser, Griet, Devreese, Bart, Asher, Claire, Blommaert, Julie, Chipman, Ariel D., Chittka, Lars, Fouks, Bertrand, Liu, Jisheng, O'Neill, Meaghan P., Sumner, Seirian, Puiu, Daniela, Qu, Jiaxin, Salzberg, Steven L., Scherer, Steven E., Muzny, Donna M., Richards, Stephen, Robinson, Gene E., Gibbs, Richard A., Schmid-Hempel, Paul, Worley, Kim C., Sadd, Ben M., Barribeau, Seth M., Bloch, Guy, de Graaf, Dirk C., Dearden, Peter, Elsik, Christine G., Gadau, Jürgen, Grimmelikhuijzen, Cornelis J. P., Hasselmann, Martin, Lozier, Jeffrey D., Robertson, Hugh M., Smagghe, Guy, Stolle, Eckart, Van Vaerenbergh, Matthias, Waterhouse, Robert M., Bornberg-Bauer, Erich, Klasberg, Steffen, Bennett, Anna K., Câmara, Francisco, Guigó, Roderic, Hoff, Katharina, Mariotti, Marco, Munoz-Torres, Monica, Murphy, Terence, Santesmasses, Didac, Amdam, Gro V., Beckers, Matthew, Beye, Martin, Biewer, Matthias, Bitondi, Márcia M. G., Blaxter, Mark L., Bourke, Andrew F. G., Brown, Mark J. F., Buechel, Severine D., Cameron, Rossanah, Cappelle, Kaat, Carolan, James C., Christiaens, Olivier, Ciborowski, Kate L., Clarke, David F., Colgan, Thomas J., Collins, David H., Cridge, Andrew G., Dalmay, Tamas, Dreier, Stephanie, du Plessis, Louis, Duncan, Elizabeth, Erler, Silvio, Evans, Jay, Falcon, Tiago, Flores, Kevin, Freitas, Flávia C. P., Fuchikawa, Taro, Gempe, Tanja, Hartfelder, Klaus, Hauser, Frank, Helbing, Sophie, Humann, Fernanda C., Irvine, Frano, Jermiin, Lars S., Johnson, Claire E., Johnson, Reed M., Jones, Andrew K., Kadowaki, Tatsuhiko, Kidner, Jonathan H., Koch, Vasco, Köhler, Arian, Kraus, F. Bernhard, Lattorff, H. Michael G., Leask, Megan, Lockett, Gabrielle A., Mallon, Eamonn B., Antonio, David S. Marco, Marxer, Monika, Meeus, Ivan, Moritz, Robin F. A., Nair, Ajay, Näpflin, Kathrin, Nissen, Inga, Niu, Jinzhi, Nunes, Francis M. F., Oakeshott, John G., Osborne, Amy, Otte, Marianne, Pinheiro, Daniel G., Rossié, Nina, Rueppell, Olav, Santos, Carolina G., Schmid-Hempel, Regula, Schmitt, Björn D., Schulte, Christina, Simões, Zilá L. P., Soares, Michelle P. M., Swevers, Luc, Winnebeck, Eva C., Wolschin, Florian, Yu, Na, Zdobnov, Evgeny M., Aqrawi, Peshtewani K., Blankenburg, Kerstin P., Coyle, Marcus, Francisco, Liezl, Hernandez, Alvaro G., Holder, Michael, Hudson, Matthew E., Jackson, LaRonda, Jayaseelan, Joy, Joshi, Vandita, Kovar, Christie, Lee, Sandra L., Mata, Robert, Mathew, Tittu, Newsham, Irene F., Ngo, Robin, Okwuonu, Geoffrey, Pham, Christopher, Pu, Ling-Ling, Saada, Nehad, Santibanez, Jireh, Simmons, DeNard, Thornton, Rebecca, Venkat, Aarti, Walden, Kimberly Ko, Wu, Yuan-Qing, Debyser, Griet, Devreese, Bart, Asher, Claire, Blommaert, Julie, Chipman, Ariel D., Chittka, Lars, Fouks, Bertrand, Liu, Jisheng, O'Neill, Meaghan P., Sumner, Seirian, Puiu, Daniela, Qu, Jiaxin, Salzberg, Steven L., Scherer, Steven E., Muzny, Donna M., Richards, Stephen, Robinson, Gene E., Gibbs, Richard A., Schmid-Hempel, Paul, and Worley, Kim C.

Abstract

BACKGROUND: The shift from solitary to social behavior is one of the major evolutionary transitions. Primitively eusocial bumblebees are uniquely placed to illuminate the evolution of highly eusocial insect societies. Bumblebees are also invaluable natural and agricultural pollinators, and there is widespread concern over recent population declines in some species. High-quality genomic data will inform key aspects of bumblebee biology, including susceptibility to implicated population viability threats. RESULTS: We report the high quality draft genome sequences of Bombus terrestris and Bombus impatiens, two ecologically dominant bumblebees and widely utilized study species. Comparing these new genomes to those of the highly eusocial honeybee Apis mellifera and other Hymenoptera, we identify deeply conserved similarities, as well as novelties key to the biology of these organisms. Some honeybee genome features thought to underpin advanced eusociality are also present in bumblebees, indicating an earlier evolution in the bee lineage. Xenobiotic detoxification and immune genes are similarly depauperate in bumblebees and honeybees, and multiple categories of genes linked to social organization, including development and behavior, show high conservation. Key differences identified include a bias in bumblebee chemoreception towards gustation from olfaction, and striking differences in microRNAs, potentially responsible for gene regulation underlying social and other traits. CONCLUSIONS: These two bumblebee genomes provide a foundation for post-genomic research on these key pollinators and insect societies. Overall, gene repertoires suggest that the route to advanced eusociality in bees was mediated by many small changes in many genes and processes, and not by notable expansion or depauperation.

Published
2015
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3. Global discovery and characterization of small non-coding RNAs in marine microalgae

Author

Biotechnology and Biological Sciences Research Council (UK), Leverhulme Trust, Ibercaja Obra Social, Ministerio de Ciencia e Innovación (España), López-Gomollón, Sara, Beckers, Matthew, Rathjen, Tina, Moxon, Simon, Maumus, Florian, Mohorianu, Irina, Moulton, Vincent, Dalmay, Tamas, Mock, Thomas, Biotechnology and Biological Sciences Research Council (UK), Leverhulme Trust, Ibercaja Obra Social, Ministerio de Ciencia e Innovación (España), López-Gomollón, Sara, Beckers, Matthew, Rathjen, Tina, Moxon, Simon, Maumus, Florian, Mohorianu, Irina, Moulton, Vincent, Dalmay, Tamas, and Mock, Thomas

Abstract

[Background] Marine phytoplankton are responsible for 50% of the CO2 that is fixed annually worldwide and contribute massively to other biogeochemical cycles in the oceans. Diatoms and coccolithophores play a significant role as the base of the marine food web and they sequester carbon due to their ability to form blooms and to biomineralise. To discover the presence and regulation of short non-coding RNAs (sRNAs) in these two important phytoplankton groups, we sequenced short RNA transcriptomes of two diatom species (Thalassiosira pseudonana, Fragilariopsis cylindrus) and validated them by Northern blots along with the coccolithophore Emiliania huxleyi., [Results] Despite an exhaustive search, we did not find canonical miRNAs in diatoms. The most prominent classes of sRNAs in diatoms were repeat-associated sRNAs and tRNA-derived sRNAs. The latter were also present in E. huxleyi. tRNA-derived sRNAs in diatoms were induced under important environmental stress conditions (iron and silicate limitation, oxidative stress, alkaline pH), and they were very abundant especially in the polar diatom F. cylindrus (20.7% of all sRNAs) even under optimal growth conditions., [Conclusions] This study provides first experimental evidence for the existence of short non-coding RNAs in marine microalgae. Our data suggest that canonical miRNAs are absent from diatoms. However, the group of tRNA-derived sRNAs seems to be very prominent in diatoms and coccolithophores and maybe used for acclimation to environmental conditions.

Published
2014

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