Keeling
Patrick J.
Keeling
Patrick J.
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ArticleThe Marine Microbial Eukaryote Transcriptome Sequencing Project (MMETSP) : illuminating the functional diversity of eukaryotic life in the oceans through transcriptome sequencing(Public Library of Science, 2014-06-24) Keeling, Patrick J. ; Burki, Fabien ; Wilcox, Heather M. ; Allam, Bassem ; Allen, Eric E. ; Amaral-Zettler, Linda A. ; Armbrust, E. Virginia ; Archibald, John M. ; Bharti, Arvind K. ; Bell, Callum J. ; Beszteri, Bank ; Bidle, Kay D. ; Cameron, Connor T. ; Campbell, Lisa ; Caron, David A. ; Cattolico, Rose Ann ; Collier, Jackie L. ; Coyne, Kathryn J. ; Davy, Simon K. ; Deschamps, Phillipe ; Dyhrman, Sonya T. ; Edvardsen, Bente ; Gates, Ruth D. ; Gobler, Christopher J. ; Greenwood, Spencer J. ; Guida, Stephanie M. ; Jacobi, Jennifer L. ; Jakobsen, Kjetill S. ; James, Erick R. ; Jenkins, Bethany D. ; John, Uwe ; Johnson, Matthew D. ; Juhl, Andrew R. ; Kamp, Anja ; Katz, Laura A. ; Kiene, Ronald P. ; Kudryavtsev, Alexander N. ; Leander, Brian S. ; Lin, Senjie ; Lovejoy, Connie ; Lynn, Denis ; Marchetti, Adrian ; McManus, George ; Nedelcu, Aurora M. ; Menden-Deuer, Susanne ; Miceli, Cristina ; Mock, Thomas ; Montresor, Marina ; Moran, Mary Ann ; Murray, Shauna A. ; Nadathur, Govind ; Nagai, Satoshi ; Ngam, Peter B. ; Palenik, Brian ; Pawlowski, Jan ; Petroni, Giulio ; Piganeau, Gwenael ; Posewitz, Matthew C. ; Rengefors, Karin ; Romano, Giovanna ; Rumpho, Mary E. ; Rynearson, Tatiana A. ; Schilling, Kelly B. ; Schroeder, Declan C. ; Simpson, Alastair G. B. ; Slamovits, Claudio H. ; Smith, David R. ; Smith, G. Jason ; Smith, Sarah R. ; Sosik, Heidi M. ; Stief, Peter ; Theriot, Edward ; Twary, Scott N. ; Umale, Pooja E. ; Vaulot, Daniel ; Wawrik, Boris ; Wheeler, Glen L. ; Wilson, William H. ; Xu, Yan ; Zingone, Adriana ; Worden, Alexandra Z.Microbial ecology is plagued by problems of an abstract nature. Cell sizes are so small and population sizes so large that both are virtually incomprehensible. Niches are so far from our everyday experience as to make their very definition elusive. Organisms that may be abundant and critical to our survival are little understood, seldom described and/or cultured, and sometimes yet to be even seen. One way to confront these problems is to use data of an even more abstract nature: molecular sequence data. Massive environmental nucleic acid sequencing, such as metagenomics or metatranscriptomics, promises functional analysis of microbial communities as a whole, without prior knowledge of which organisms are in the environment or exactly how they are interacting. But sequence-based ecological studies nearly always use a comparative approach, and that requires relevant reference sequences, which are an extremely limited resource when it comes to microbial eukaryotes. In practice, this means sequence databases need to be populated with enormous quantities of data for which we have some certainties about the source. Most important is the taxonomic identity of the organism from which a sequence is derived and as much functional identification of the encoded proteins as possible. In an ideal world, such information would be available as a large set of complete, well-curated, and annotated genomes for all the major organisms from the environment in question. Reality substantially diverges from this ideal, but at least for bacterial molecular ecology, there is a database consisting of thousands of complete genomes from a wide range of taxa, supplemented by a phylogeny-driven approach to diversifying genomics. For eukaryotes, the number of available genomes is far, far fewer, and we have relied much more heavily on random growth of sequence databases, raising the question as to whether this is fit for purpose.
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ArticleA kleptoplastidic dinoflagellate and the tipping point between transient and fully integrated plastid endosymbiosis(National Academy of Sciences, 2019-09-03) Hehenberger, Elisabeth ; Gast, Rebecca J. ; Keeling, Patrick J.Plastid endosymbiosis has been a major force in the evolution of eukaryotic cellular complexity, but how endosymbionts are integrated is still poorly understood at a mechanistic level. Dinoflagellates, an ecologically important protist lineage, represent a unique model to study this process because dinoflagellate plastids have repeatedly been reduced, lost, and replaced by new plastids, leading to a spectrum of ages and integration levels. Here we describe deep-transcriptomic analyses of the Antarctic Ross Sea dinoflagellate (RSD), which harbors long-term but temporary kleptoplasts stolen from haptophyte prey, and is closely related to dinoflagellates with fully integrated plastids derived from different haptophytes. In some members of this lineage, called the Kareniaceae, their tertiary haptophyte plastids have crossed a tipping point to stable integration, but RSD has not, and may therefore reveal the order of events leading up to endosymbiotic integration. We show that RSD has retained its ancestral secondary plastid and has partitioned functions between this plastid and the kleptoplast. It has also obtained genes for kleptoplast-targeted proteins via horizontal gene transfer (HGT) that are not derived from the kleptoplast lineage. Importantly, many of these HGTs are also found in the related species with fully integrated plastids, which provides direct evidence that genetic integration preceded organelle fixation. Finally, we find that expression of kleptoplast-targeted genes is unaffected by environmental parameters, unlike prey-encoded homologs, suggesting that kleptoplast-targeted HGTs have adapted to posttranscriptional regulation mechanisms of the host.
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ArticleThe reduced genome of the parasitic microsporidian Enterocytozoon bieneusi lacks genes for core carbon metabolism(Oxford University Press, 2010-05-13) Keeling, Patrick J. ; Corradi, Nicolas ; Morrison, Hilary G. ; Haag, Karen L. ; Ebert, Dieter ; Weiss, Louis M. ; Akiyoshi, Donna E. ; Tzipori, SaulReduction of various biological processes is a hallmark of the parasitic lifestyle. Generally, the more intimate the association between parasites and hosts the stronger the parasite relies on its host's physiology for survival and reproduction. However, some systems have been held to be indispensable, for example, the core pathways of carbon metabolism that produce energy from sugars. Even the most hardened anaerobes that lack oxidative phosphorylation and the tricarboxylic acid cycle have retained glycolysis and some downstream means to generate ATP. Here we describe the deep-coverage genome resequencing of the pathogenic microsporidiian, Enterocytozoon bieneusi, which shows that this parasite has crossed this line and abandoned complete pathways for the most basic carbon metabolism. Comparing two genome sequence surveys of E. bieneusi to genomic data from four other microsporidia reveals a normal complement of 353 genes representing 30 functional pathways in E. bieneusi, except that only 2 out of 21 genes collectively involved in glycolysis, pentose phosphate, and trehalose metabolism are present. Similarly, no genes encoding proteins involved in the processing of spliceosomal introns were found. Altogether, E. bieneusi appears to have no fully functional pathway to generate ATP from glucose. Therefore, this intracellular parasite relies on transporters to import ATP from its host.
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ArticleGenetic tool development in marine protists: emerging model organisms for experimental cell biology(Nature Research, 2020-04-06) Faktorová, Drahomíra ; Nisbet, R. Ellen R. ; Fernández Robledo, José A. ; Casacuberta, Elena ; Sudek, Lisa ; Allen, Andrew E. ; Ares, Manuel, Jr. ; Aresté, Cristina ; Balestreri, Cecilia ; Barbrook, Adrian C. ; Beardslee, Patrick ; Bender, Sara J. ; Booth, David S. ; Bouget, François-Yves ; Bowler, Chris ; Breglia, Susana A. ; Brownlee, Colin ; Burger, Gertraud ; Cerutti, Heriberto ; Cesaroni, Rachele ; Chiurillo, Miguel A. ; Clemente, Thomas ; Coles, Duncan B. ; Collier, Jackie L. ; Cooney, Elizabeth C. ; Coyne, Kathryn J. ; Docampo, Roberto ; Dupont, Christopher L. ; Edgcomb, Virginia P. ; Einarsso, Elin ; Elustondo, Pía A. ; Federici, Fernan ; Freire-Beneitez, Veronica ; Freyria, Nastasia J. ; Fukuda, Kodai ; García, Paulo A. ; Girguis, Peter R. ; Gomaa, Fatma ; Gornik, Sebastian G. ; Guo, Jian ; Hampl, Vladimír ; Hanawa, Yutaka ; Haro-Contreras, Esteban R. ; Hehenberger, Elisabeth ; Highfield, Andrea ; Hirakawa, Yoshihisa ; Hopes, Amanda ; Howe, Christopher J. ; Hu, Ian ; Ibañez, Jorge ; Irwin, Nicholas A. T. ; Ishii, Yuu ; Janowicz, Natalia Ewa ; Jones, Adam C. ; Kachale, Ambar ; Fujimura-Kamada, Konomi ; Kaur, Binnypreet ; Kaye, Jonathan Z. ; Kazana, Eleanna ; Keeling, Patrick J. ; King, Nicole ; Klobutcher, Lawrence A. ; Lander, Noelia ; Lassadi, Imen ; Li, Zhuhong ; Lin, Senjie ; Lozano, Jean-Claude ; Luan, Fulei ; Maruyama, Shinichiro ; Matute, Tamara ; Miceli, Cristina ; Minagawa, Jun ; Moosburner, Mark ; Najle, Sebastián R. ; Nanjappa, Deepak ; Nimmo, Isabel C. ; Noble, Luke ; Novák Vanclová, Anna M. G. ; Nowacki, Mariusz ; Nuñez, Isaac ; Pain, Arnab ; Piersanti, Angela ; Pucciarelli, Sandra ; Pyrih, Jan ; Rest, Joshua S. ; Rius, Mariana ; Robertson, Deborah ; Ruaud, Albane ; Ruiz-Trillo, Iñaki ; Sigg, Monika A. ; Silver, Pamela A. ; Slamovits, Claudio H. ; Smith, G. Jason ; Sprecher, Brittany N. ; Stern, Rowena ; Swart, Estienne C. ; Tsaousis, Anastasios D. ; Tsypin, Lev ; Turkewitz, Aaron ; Turnšek, Jernej ; Valach, Matus ; Vergé, Valérie ; von Dassow, Peter ; von der Haar, Tobias ; Waller, Ross F. ; Wang, Lu ; Wen, Xiaoxue ; Wheeler, Glen L. ; Woods, April ; Zhang, Huan ; Mock, Thomas ; Worden, Alexandra Z. ; Lukes, JuliusDiverse microbial ecosystems underpin life in the sea. Among these microbes are many unicellular eukaryotes that span the diversity of the eukaryotic tree of life. However, genetic tractability has been limited to a few species, which do not represent eukaryotic diversity or environmentally relevant taxa. Here, we report on the development of genetic tools in a range of protists primarily from marine environments. We present evidence for foreign DNA delivery and expression in 13 species never before transformed and for advancement of tools for eight other species, as well as potential reasons for why transformation of yet another 17 species tested was not achieved. Our resource in genetic manipulation will provide insights into the ancestral eukaryotic lifeforms, general eukaryote cell biology, protein diversification and the evolution of cellular pathways.
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ArticleGenomic survey of the non-cultivatable opportunistic human pathogen, Enterocytozoon bieneusi(Public Library of Science, 2009-01-09) Akiyoshi, Donna E. ; Morrison, Hilary G. ; Lei, Shi ; Feng, Xiaochuan ; Zhang, Quanshun ; Corradi, Nicolas ; Mayanja, Harriet ; Tumwine, James K. ; Keeling, Patrick J. ; Weiss, Louis M. ; Tzipori, SaulEnterocytozoon bieneusi is the most common microsporidian associated with human disease, particularly in the immunocompromised population. In the setting of HIV infection, it is associated with diarrhea and wasting syndrome. Like all microsporidia, E. bieneusi is an obligate, intracellular parasite, but unlike others, it is in direct contact with the host cell cytoplasm. Studies of E. bieneusi have been greatly limited due to the absence of genomic data and lack of a robust cultivation system. Here, we present the first large-scale genomic dataset for E. bieneusi. Approximately 3.86 Mb of unique sequence was generated by paired end Sanger sequencing, representing about 64% of the estimated 6 Mb genome. A total of 3,804 genes were identified in E. bieneusi, of which 1,702 encode proteins with assigned functions. Of these, 653 are homologs of Encephalitozoon cuniculi proteins. Only one E. bieneusi protein with assigned function had no E. cuniculi homolog. The shared proteins were, in general, evenly distributed among the functional categories, with the exception of a dearth of genes encoding proteins associated with pathways for fatty acid and core carbon metabolism. Short intergenic regions, high gene density, and shortened protein-coding sequences were observed in the E. bieneusi genome, all traits consistent with genomic compaction. Our findings suggest that E. bieneusi is a likely model for extreme genome reduction and host dependence.