Jump to ContentJump to Main Navigation
Show Summary Details

Gene discovery from a pilot study of the transcriptomes from three diverse microbial eukaryotes: Corallomyxa tenera, Chilodonella uncinata, and Subulatomonas tetraspora

Jessica R. Grant
  • Department of Biological Sciences, Smith College, 44 College Lane, Northampton, Massachusetts 01063, USA
/ Daniel J.G. Lahr
  • Department of Biological Sciences, Smith College, 44 College Lane, Northampton, Massachusetts 01063, USA
  • Program in Organismic and Evolutionary Biology, University of Massachusetts, 611 North Pleasant Street, Amherst, Massachusetts 01003, USA
  • Department of Zoology, Institute of Biosciences, University of Sao Paulo, Brazil
/ Federico E. Rey
  • Center for Genome Science & Systems Biology, Washington University School of Medicine, St Louis, MO 63108 USA
/ J. Gordon Burleigh
  • Department of Biology, University of Florida, PO Box 118526, Gainseville, FL 32611, USA
/ Jeffrey I. Gordon
  • Center for Genome Science & Systems Biology, Washington University School of Medicine, St Louis, MO 63108 USA
/ Rob Knight
  • Department of Chemistry & Biochemistry, University of Colorado, Boulder, CO 80309 USA
/ Robert E. Molestina
  • American Type Culture Collection, Protistology Collection, 10801 University Blvd., Manassas, Virginia 20110, USA
/ Laura A. Katz
  • Department of Biological Sciences, Smith College, 44 College Lane, Northampton, Massachusetts 01063, USA
  • Program in Organismic and Evolutionary Biology, University of Massachusetts, 611 North Pleasant Street, Amherst, Massachusetts 01003, USA
  • Email:
Published Online: 2012-12-28 | DOI: https://doi.org/10.2478/prge-2012-0002

Abstract

Background:

Characterizing genome-scale data from diverse eukaryotes is essential for gene discovery and for inferring major transitions across the eukaryotic tree of life. Yet, the bulk of eukaryotic diversity remains undersampled, particularly for free-living microbial lineages. Analysis of transcriptome data generated from high throughput (e.g. 454) sequencing of mRNAs provides an efficient way to characterize genes from diverse eukaryotes.

Results:

Here we report analyses of RNA-Seq data from the rhizarian net-like amoeba Corallomyxa tenera, the ciliate Chilodonella uncinata and a recently-described genus representing a novel major clade of eukaryotes, Subulatomonas tetraspora. We generated 16,983, 11,529 and 10,630 contigs plus single reads for these taxa respectively. Given that these organisms cannot be cultured axenically, we developed custom scripts to remove bacterial contaminants through an iterative BLAST based protocol and we then identified expressed genes using BLAST2GO [1;2]. This approach yielded a large number of genes with eukaryotic homologs, as well as numerous novel genes. To assess our approach and to explore the resulting sequences, we searched for genes involved in anaerobic metabolism, RNAi and meiosis. Further, we report the results of a preliminary phylogenomic analysis including these organisms.

Conclusions:

We characterized the transcriptomes of three phylogenetically diverse eukaryotes. After applying several filters to ensure the retention of only high-quality, non-contaminant data, we identified numerous sequences that can be used for gene discovery and phylogenomics. We found candidate genes involved in RNAi, meiosis, and anaerobic metabolism, and generated phylogenies that place the target taxa in positions predicted by previous analyses. This work supports the use of high throughput approaches for assessing features of non-model organisms, even in instances when species cannot be cultured axenically or grown to large numbers.

This article offers supplementary material which is provided at the end of the article.

Keywords: Transcriptome; 454 sequencing; Nonmodel organisms; Phylogenetics; Chilodonella uncinata; Corallomyxa tenera; Subulatomonas tetraspora

  • Gotz, S., Garcia-Gomez, J.M., Terol, J., Williams, T.D., Nagaraj, S.H., Nueda, M.J., et al., High-throughput functional annotation and data mining with the Blast2GO suite, Nucleic Acids Res., 2008, 36, 3420-3435

  • Conesa, A., Gotz, S., Garcia-Gomez, J.M., Terol, J., Talon, M., Robles, M., Blast2GO: a universal tool for annotation, visualization and analysis in functional genomics research, Bioinformatics, 2005, 21, 3674-3676

  • Katz, L.A., Origin and diversification of eukaryotes, Annu. Rev. Microbiol., 2012, 66, 411-427

  • Roger, A.J., Hug, L.A., The origin and diversification of eukaryotes: problems with molecular phylogenetics and molecular clock estimation, Philos. Trans. R. Soc. B-Biol. Sci., 2006, 361, 1039-1054

  • Tekle, Y.I., Parfrey, L.W., Katz, L.A., Molecular data are transforming hypotheses on the origin and diversification of eukaryotes, Bioscience, 2009, 59, 471-481

  • Hampl, V., Hug, L., Leigh, J.W., Dacks, J.B., Lang, B.F., Simpson, A.G.B., et al., Phylogenomic analyses support the monophyly of Excavata and resolve relationships among eukaryotic “supergroups”, Proc. Natl. Acad. Sci. U. S. A., 2009, 106, 3859-3864 [Crossref]

  • Andersson, J.O., Sjogren, A.M., Horner, D.S., Murphy, C.A., Dyal, P.L., Svard, S.G., et al., A genomic survey of the fish parasite Spironucleus salmonicida indicates genomic plasticity among diplomonads and significant lateral gene transfer in eukaryote genome evolution, BMC Genomics, 2007, 8, 25

  • Yang, I., John, U., Beszteri, S., Glockner, G., Krock, B., Goesmann, A., et al., Comparative gene expression in toxic versus non-toxic strains of the marine dinoflagellate Alexandrium minutum, BMC Genomics, 2010, 11, 18

  • Joseph, S.J., Fernandez-Robledo, J.A., Gardner, M.J., El- Sayed, N.M., Kuo, C.H., Schott, E.J., et al., The Alveolate Perkinsus marinus: Biological Insights from EST Gene Discovery, BMC Genomics, 2010, 11, 21

  • Lanier, W., Moustafa, A., Bhattacharya, D., Comeron, J.M., EST Analysis of Ostreococcus lucimarinus, the Most Compact Eukaryotic Genome, Shows an Excess of Introns in Highly Expressed Genes, PLoS One, 2008, 3, 7

  • Canovas, F.G., Mota, C.F., Serrao, E.A., Pearson, G.A., Driving south: a multi-gene phylogeny of the brown algal family Fucaceae reveals relationships and recent drivers of a marine radiation, BMC Evol. Biol., 2011, 11, [Crossref]

  • Burki, F., Okamoto, N., Pombert, J.F., Keeling, P.J., The evolutionary history of haptophytes and cryptophytes: phylogenomic evidence for separate origins, Proceedings. Biological sciences / The Royal Society, 2012, 279, 2246- 2254

  • Wall, P.K., Leebens-Mack, J., Chanderbali, A.S., Barakat, A., Wolcott, E., Liang, H.Y., et al., Comparison of next generation sequencing technologies for transcriptome characterization, BMC Genomics, 2009, 10, - [PubMed] [Crossref]

  • Hert, D.G., Fredlake, C.P., Barron, A.E., Advantages and limitations of next-generation sequencing technologies: A comparison of electrophoresis and non-electrophoresis methods, Electrophoresis, 2008, 29, 4618-4626 [PubMed] [Crossref]

  • Hittinger, C.T., Johnston, M., Tossberg, J.T., Rokas, A., Leveraging skewed transcript abundance by RNA-Seq to increase the genomic depth of the tree of life, Proc. Natl. Acad. Sci. U. S. A., 2010, 107, 1476-1481 [Crossref]

  • Zhang, F.J., Guo, H.Y., Zheng, H.J., Zhou, T., Zhou, Y.J., Wang, S.Y., et al., Massively parallel pyrosequencingbased transcriptome analyses of small brown planthopper (Laodelphax striatellus), a vector insect transmitting rice stripe virus (RSV), BMC Genomics, 2010, 11, - [PubMed] [Crossref]

  • Zagrobelny, M., Scheibye-Alsing, K., Jensen, N.B., Moller, B.L., Gorodkin, J., Bak, S., 454 pyrosequencing based transcriptome analysis of Zygaena filipendulae with focus on genes involved in biosynthesis of cyanogenic glucosides, BMC Genomics, 2009, 10, - [Crossref]

  • Vera, J.C., Wheat, C.W., Fescemyer, H.W., Frilander, M.J., Crawford, D.L., Hanski, I., et al., Rapid transcriptome characterization for a nonmodel organism using 454 pyrosequencing, Mol. Ecol., 2008, 17, 1636-1647 [Crossref]

  • Parchman, T.L., Geist, K.S., Grahnen, J.A., Benkman, C.W., Buerkle, C.A., Transcriptome sequencing in an ecologically important tree species: assembly, annotation, and marker discovery, BMC Genomics, 2010, 11, - [Crossref]

  • Rismani-Yazdi, H., Haznedaroglu, B.Z., Bibby, K., Peccia, J., Transcriptome sequencing and annotation of the microalgae Dunaliella tertiolecta: Pathway description and gene discovery for production of next-generation biofuels, BMC Genomics, 2011, 12, - [Crossref]

  • McGrath, C., Zufall, R.A., Katz, L.A., Genome evolution in ciliates, In: LA Katz, D Bhattacharya, editors. Genomics and Evolution of Eukaryotic Microbes, Oxford University Press. 2006

  • Riley, J.L., Katz, L.A., Widespread distribution of extensive genome fragmentation in ciliates, Mol. Biol. Evol., 2001, 18, 1372-1377 [Crossref]

  • Aury, J.M., Jaillon, O., Duret, L., Noel, B., Jubin, C., Porcel, B.M., et al., Global trends of whole-genome duplications revealed by the ciliate Paramecium tetraurelia, Nature, 2006, 444, 171-178

  • Eisen, J.A., Coyne, R.S., Wu, M., Wu, D., Thiagarajan, M., Wortman, J.R., et al., Macronuclear genome sequence of the ciliate Tetrahymena thermophila, a model eukaryote, PLoS Biol, 2006, 4, e286 [Crossref]

  • Doak, T.G., Cavalcanti, A.R.O., Stover, N.A., Dunn, D.M., Weiss, R., Herrick, G., et al., Sequencing the Oxytricha trifallax macronuclear genome: a pilot project, Trends Genet., 2003, 19, 603-607 [Crossref]

  • Katz, L.A., Kovner, A.M., Alternative processing of scrambled genes generates protein diversity in the ciliate Chilodonella uncinata, J. Exper. Zool. B, 2010, 314, 480-488

  • Zufall, R.A., McGrath, C., Muse, S.V., Katz, L.A., Genome architecture drives protein evolution in ciliates., Mol. Biol. Evol., 2006, 23, 1681-1687 [Crossref] [PubMed]

  • Tekle, Y.I., Grant, J., Cole, J.C., Nerad, T.A., Anderson, O.R., Patterson, D.J., et al., A multigene analysis of Corallomyxa tenera sp. nov. suggests its membership in a clade that includes Gromia, Haplosporidia and Foraminifera, Protist, 2007, 158, 457-472

  • Habura, A., Hou, Y.B., Reilly, A.A., Bowser, S.S., Highthroughput sequencing of Astrammina rara: Sampling the giant genome of a giant foraminiferan protist, BMC Genomics, 2011, 12, 11

  • Burki, F., Kudryavtsev, A., Matz, M.V., Aglyamova, G.V., Bulman, S., Fiers, M., et al., Evolution of Rhizaria: new insights from phylogenomic analysis of uncultivated protists, BMC Evol. Biol., 2010, 10, 18

  • Katz, L.A., Grant, J.R., Parfrey, L.W., Gant, A., O’Kelly, C.J., Anderson, O.R., et al., Subulatomonas tetraspora nov. gen. nov. sp. is a member of a previously unrecognized major clade of eukaryotes, Protist, 2011, 162, 762-773

  • Minge, M.A., Silberman, J.D., Orr, R.J.S., Cavalier-Smith, T., Shalchian-Tabrizi, K., Burki, F., et al., Evolutionary position of breviate amoebae and the primary eukaryote divergence, Proc R. Soc Lond B Biol Sci, 2009, 276, 597-604

  • Dessen, P., Zagulski, M., Gromadka, R., Plattner, H., Kissmehl, R., Meyer, E., et al., Paramecium genome survey: a pilot project, Trends Genet., 2001, 17, 306-308 [PubMed] [Crossref]

  • Zufall, R.A., Katz, L.A., Micronuclear and macronuclear forms of beta-tubulin genes in the ciliate Chilodonella uncinata reveal insights into genome processing and protein evolution, J. Eukaryot. Microbiol., 2007, 54, 275-282 [Crossref]

  • Katz, L.A., DeBerardinis, J., Hall, M.S., Kovner, A.M., Dunthorn, M., Muse, S.V., Heterogeneous Rates of Molecular Evolution Among Cryptic Species of the Ciliate Morphospecies Chilodonella uncinata, J. Mol. Evol., 2011, 73, 266-272

  • Andersson, J.O., Hirt, R.P., Foster, P.G., Roger, A.J., Evolution of four gene families with patchy phylogenetic distributions: influx of genes into protist genomes, BMC Evol. Biol., 2006, 6, 18

  • Ginger, M.L., Fritz-Laylin, L.K., Fulton, C., Cande, W.Z., Dawson, S.C., Intermediary Metabolism in Protists: a Sequence-based View of Facultative Anaerobic Metabolism in Evolutionarily Diverse Eukaryotes, Protist, 2010, 161, 642- 671

  • Hug, L.A., Stechmann, A., Roger, A.J., Phylogenetic distributions and histories of proteins involved in anaerobic pyruvate metabolism in eukaryotes, Mol. Biol. Evol., 2010, 27, 311-324 [PubMed] [Crossref]

  • Parfrey, L.W., Katz, L.A., Dynamic genomes of eukaryotes and the maintenance of genomic integrity, Microbe 2010, 5, 156-164

  • Parfrey, L.W., Lahr, D.J.G., Katz, L.A., The dynamic nature of eukaryotic genomes, Mol. Biol. Evol., 2008, 25, 787-794 [PubMed] [Crossref]

  • Cerutti, H., Casas-Mollano, J.A., On the oridin and functions of RNA-mediated silencing: from protists to man, Curr. Genet., 2006, 50, 81-99

  • Shabalina, S.A., Koonin, E.V., Origins and evolution of eukaryotic RNA interference, Trends Ecol Evol, 2008, 23, 578-587 [PubMed] [Crossref]

  • Costa, F.F., Non-coding RNAs: Meet thy masters, Bioessays, 2010, 32, 599-608 [Crossref] [PubMed]

  • Aravin, A.A., Hannon, G.J., Brennecke, J., The Piwi-piRNA pathway provides an adaptive defense in the transposon arms race, Science, 2007, 318, 761-764

  • Alie, A., Leclere, L., Jager, M., Dayraud, C., Chang, P.R., Le Guyader, H., et al., Somatic stem cells express Piwi and Vasa genes in an adult ctenophore: Ancient association of “germline genes” with stemness, Dev. Biol., 2011, 350, 183-197

  • Mochizuki, K., Gorovsky, M.A., Small RNAs in genome rearrangement in Tetrahymena, Curr. Opin. Genet. Dev., 2004, 14, 181 [Crossref] [PubMed]

  • Burt, A., Sex, recombination, and the efficacy of natural selection — was Weisman right?, Evolution, 2000, 54, 337- 351

  • Felsenstein, J., The evolutionary advantage of recombination, Genetics, 1974, 78, 737-756 [PubMed]

  • Lahr, D.J., Parfrey, L.W., Mitchell, E.A., Katz, L.A., Lara, E., The chastity of amoebae: re-evaluating evidence for sex in amoeboid organisms, Proc Biol Sci, 2011, 278, 2081-2090

  • Dunthorn, M., Katz, L.A., Secretive ciliates and putative asexuality in microbial eukaryotes, Trends Microbiol., 2010, 18, 183-188 [Crossref] [PubMed]

  • Schurko, A.M., Logsdon, J.M., Using a meiosis detection toolkit to investigate ancient asexual ldquoscandalsrdquo and the evolution of sex, Bioessays, 2008, 30, 579-589 [Crossref] [PubMed]

  • Parfrey, L.W., Grant, J., Tekle, Y.I., Lasek-Nesselquist, E., Morrison, H.G., Sogin, M.L., et al., Broadly sampled multigene analyses yield a well-resolved eukaryotic tree of life, Syst Biol, 2010, 59, 518-533

  • Parfrey, L.W., Lahr, D.J.G., Knoll, A.H., Katz, L.A., Estimating the timing of early eukaryotic diversification with multigene molecular clocks, Proc. Natl. Acad. Sci. U. S. A., 2011, 108, 13624-13629 [Crossref]

  • Zhao, S., Burki, F., Brate, J., Keeling, P.J., Klaveness, D., Shalchian-Tabrizi, K., Collodictyon--an ancient lineage in the tree of eukaryotes, Mol. Biol. Evol., 2012, 29, 1557-1568 [PubMed] [Crossref]

  • Wehe, A., Bansal, M.S., Burleigh, J.G., Eulenstein, O., DupTree: a program for large-scale phylogenetic analyses using gene tree parsimony, Bioinformatics, 2008, 24, 1540- 1541 [PubMed] [Crossref]

  • Bansal, M.S., Burleigh, J.G., Eulenstein, O., Efficient genomescale phylogenetic analysis under the duplication-loss and deep coalescence cost models, BMC Bioinformatics, 2010, 11 Suppl 1, S42 [Crossref]

  • Burleigh, J.G., Driskell, A.C., Sanderson, M.J., Supertree bootstrapping methods for assessing phylogenetic variation among genes in genome-scale data sets, Syst Biol, 2006, 55, 426-440 [Crossref] [PubMed]

  • Philippe, H., Snell, E.A., Bapteste, E., Lopez, P., Holland, P.W., Casane, D., Phylogenomics of eukaryotes: impact of missing data on large alignments, Mol. Biol. Evol., 2004, 21, 1740-1752 [PubMed] [Crossref]

  • Cavalier-Smith, T., Chao, E.E.Y., Oates, B., Molecular phylogeny of Amoebozoa and the evolutionary significance of the unikont Phalansterium, Eur J Protistol, 2004, 40, 21-48 [Crossref]

  • Shalchian-Tabrizi, K., Eikrem, W., Klaveness, D., Vaulot, D., Minge, M.A., Le Gall, F., et al., Telonemia, a new protist phylum with affinity to chromist lineages, Proceedings. Biological sciences / The Royal Society, 2006, 273, 1833- 1842

  • Walker, G., Dacks, J.B., Embley, T.M., Ultrastructural description of Breviata anathema, n. Gen., n. Sp., the organism previously studied as “Mastigamoeba invertens”. J. Eukaryot. Microbiol., 2006, 53, 65-78

  • Simpson, A.G.B., Cytoskeletal organization, phylogenetic affinities and systematics in the contentious taxon Excavata (Eukaryota), Int J Syst Evol Micr, 2003, 53, 1759-1777

  • Simpson, A.G.B., Inagaki, Y., Roger, A.J., Comprehensive multigene phylogenies of excavate protists reveal the evolutionary positions of “primitive” eukaryotes, Mol. Biol. Evol., 2006, 23, 615-625 [PubMed]

  • Giardine, B., Riemer, C., Hardison, R.C., Burhans, R., Elnitski, L., Shah, P., et al., Galaxy: A platform for interactive largescale genome analysis, Genome Res., 2005, 15, 1451-1455 [Crossref]

  • Chen, F., Mackey, A.J., Stoeckert, C.J., Roos, D.S., OrthoMCL-DB: querying a comprehensive multi-species collection of ortholog groups, Nucleic Acids Res., 2006, 34, D363-D368 [Crossref]

  • Li, L., Stoeckert, C.J., Jr., Roos, D.S., OrthoMCL: Identification of Ortholog Groups for Eukaryotic Genomes, Genome Res. 10.1101/gr.1224503, 2003, 13, 2178-2189

  • Penn, O., Privman, E., Ashkenazy, H., Landan, G., Graur, D., Pupko, T., GUIDANCE: a web server for assessing alignment confidence scores, Nucleic Acids Res., 2010, 38, W23-W28 [Crossref]

  • Ott, M., Zola, J., Aluru, S., Stamatakis, A., Large-scale maximum likelihood-based phylogenetic analysis on the IBM BlueGene/L. Proceedings of ACM/IEEE Supercomputing conference, New York, NY, ACM. 2007

  • Stamatakis, A., RAxML-VI-HPC: Maximum likelihood-based phylogenetic analyses with thousands of taxa and mixed models, Bioinformatics, 2006, 22, 2688-2690 [Crossref] [PubMed]

  • Stamatakis, A., Hoover, P., Rougemont, J., A rapid bootstrap algorithm for the RAxML web-servers, Syst Biol, 2008, 57, 758-771

  • Stamatakis, A., Aberer, A.J., Goll, C., Smith, S.A., Berger, S.A., Izquierdo-Carrasco, F., RAxML-Light: a tool for computing terabyte phylogenies, Bioinformatics, 2012, 28, 2064-2066 [Crossref]

  • Chaudhary, R., Bansal, M.S., Wehe, A., Fernandez-Baca, D., Eulenstein, O., iGTP: A software package for large-scale gene tree parsimony analysis, BMC Bioinformatics, 2010, 11, [Crossref] [PubMed]

  • Junier, T., Zdobnov, E.M., The Newick utilities: highthroughput phylogenetic tree processing in the Unix shell, Bioinformatics, 2010, 26, 1669-1670 [Crossref]

  • Parfrey, L.W., Grant, J., Tekle, Y.I., Lasek-Nesselquist, E., Morrison, H.G., Sogin, M.L., et al., Broadly Sampled Multigene Analyses Yield a Well-Resolved Eukaryotic Tree of Life, Syst Biol, 2010, 59, 518-533

  • Miller, M.A., Pfeiffer, W., Schwartz, T., Creating the CIPRES Science Gateway for inference of large phylogenetic trees. Proceedings of the Gateway Computing Environments Workshop (GCE), New Orleans, LA p. 1-8, 2010

  • Stamatakis, A., Ott, M., Ludwig, T., RAxML-OMP: An efficient program for phylogenetic inference on SMPs, Parallel Computing Technologies, 2005, 3606, 288-302

  • Darriba, D., Taboada, G.L., Doallo, R., Posada, D., ProtTest 3: fast selection of best-fit models of protein evolution, Bioinformatics, 2011, 27, 1164-1165 [PubMed] [Crossref]

  • Aberer, A.J., Krompass, D., Stamatakis, A., RogueNaRok: an Efficient and Exact Algorithm for Rogue Taxon Identification, Heidelberg Institute for Theoretical Studies, 2011,

  • Schmidt, H.A., Strimmer, K., Vingron, M., von Haeseler, A., TREE-PUZZLE: maximum likelihood phylogenetic analysis using quartets and parallel computing, Bioinformatics, 2002, 18, 502-504 [PubMed] [Crossref]

About the article


Received: 2012-09-13

Accepted: 2012-12-19

Published Online: 2012-12-28



Citation Information: Protist Genomics, ISSN (Online) 2299-100X, DOI: https://doi.org/10.2478/prge-2012-0002. Export Citation

©2012 Versita Sp. z o.o.. This content is open access.

Supplementary Article Materials

Citing Articles

Here you can find all Crossref-listed publications in which this article is cited. If you would like to receive automatic email messages as soon as this article is cited in other publications, simply activate the “Citation Alert” on the top of this page.

[1]
Xiao Chen, Xiaolu Zhao, Xiaohui Liu, Alan Warren, Fangqing Zhao, and Miao Miao
Protein & Cell, 2015, Volume 6, Number 5, Page 373
[2]
Daniel J. G. Lahr, Jessica Grant, Robert Molestina, Laura A. Katz, and O. Roger Anderson
Journal of Eukaryotic Microbiology, 2015, Page n/a

Comments (0)

Please log in or register to comment.
Log in