Dataset Information

Transcriptome analysis of the model protozoan, Tetrahymena thermophila, using Deep RNA sequencing.

ABSTRACT: BACKGROUND: The ciliated protozoan Tetrahymena thermophila is a well-studied single-celled eukaryote model organism for cellular and molecular biology. However, the lack of extensive T. thermophila cDNA libraries or a large expressed sequence tag (EST) database limited the quality of the original genome annotation. METHODOLOGY/PRINCIPAL FINDINGS: This RNA-seq study describes the first deep sequencing analysis of the T. thermophila transcriptome during the three major stages of the life cycle: growth, starvation and conjugation. Uniquely mapped reads covered more than 96% of the 24,725 predicted gene models in the somatic genome. More than 1,000 new transcribed regions were identified. The great dynamic range of RNA-seq allowed detection of a nearly six order-of-magnitude range of measurable gene expression orchestrated by this cell. RNA-seq also allowed the first prediction of transcript untranslated regions (UTRs) and an updated (larger) size estimate of the T. thermophila transcriptome: 57 Mb, or about 55% of the somatic genome. Our study identified nearly 1,500 alternative splicing (AS) events distributed over 5.2% of T. thermophila genes. This percentage represents a two order-of-magnitude increase over previous EST-based estimates in Tetrahymena. Evidence of stage-specific regulation of alternative splicing was also obtained. Finally, our study allowed us to completely confirm about 26.8% of the genes originally predicted by the gene finder, to correct coding sequence boundaries and intron-exon junctions for about a third, and to reassign microarray probes and correct earlier microarray data. CONCLUSIONS/SIGNIFICANCE: RNA-seq data significantly improve the genome annotation and provide a fully comprehensive view of the global transcriptome of T. thermophila. To our knowledge, 5.2% of T. thermophila genes with AS is the highest percentage of genes showing AS reported in a unicellular eukaryote. Tetrahymena thus becomes an excellent unicellular model eukaryote in which to investigate mechanisms of alternative splicing.

SUBMITTER: Xiong J

PROVIDER: S-EPMC3274533 | biostudies-literature | 2012

REPOSITORIES: biostudies-literature

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Transcriptome analysis of the model protozoan, Tetrahymena thermophila, using Deep RNA sequencing.

Xiong Jie J Lu Xingyi X Zhou Zhemin Z Chang Yue Y Yuan Dongxia D Tian Miao M Zhou Zhigang Z Wang Lei L Fu Chengjie C Orias Eduardo E Miao Wei W

PloS one 20120207 2

<h4>Background</h4>The ciliated protozoan Tetrahymena thermophila is a well-studied single-celled eukaryote model organism for cellular and molecular biology. However, the lack of extensive T. thermophila cDNA libraries or a large expressed sequence tag (EST) database limited the quality of the original genome annotation.<h4>Methodology/principal findings</h4>This RNA-seq study describes the first deep sequencing analysis of the T. thermophila transcriptome during the three major stages of the l ...[more]

PMID: 22347391

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Project description:BackgroundCadmium (Cd) is a nonessential heavy metal with potentially deleterious effects on different organisms. The organisms have evolved sophisticated defense system to alleviate heavy metal toxicity. Hydrogen sulfide (H2S) effectively alleviates heavy metal toxicity in plants and reduces oxidative stress in mammals. However, the function of H2S for alleviating heavy metal toxicity in aquatic organisms remains less clear. Tetrahymena thermophila is an important model organism to evaluate toxic contaminants in an aquatic environment. In this study, the molecular roles of exogenously H2S application were explored by RNA sequencing under Cd stress in T. thermophila.ResultsThe exposure of 30 μM Cd resulted in T. thermophila growth inhibition, cell nigrescence, and malondialdehyde (MDA) content considerably increase. However, exogenous NaHS (donor of H2S, 70 μM) significantly alleviated the Cd-induced toxicity by inhibiting Cd absorbtion, promoting CdS nanoparticles formation and improving antioxidant system. Comparative transcriptome analysis showed that the expression levels of 9152 genes changed under Cd stress (4658 upregulated and 4494 downregulated). However, only 1359 genes were differentially expressed with NaHS treatment under Cd stress (1087 upregulated and 272 downregulated). The functional categories of the differentially expressed genes (DEGs) by gene ontology (GO) revealed that the transcripts involved in the oxidation-reduction process, oxidoreductase activity, glutathione peroxidase activity, and cell redox homeostasis were the considerable enrichments between Cd stress and NaHS treatment under Cd stress. Kyoto Encyclopedia of Genes and Genomes (KEGG) indicated that the carbon metabolism, glutathione metabolism, metabolism of xenobiotics by cytochrome P450, and ABC transporters were significantly differentially expressed components between Cd stress and NaHS treatment under Cd stress in T. thermophila. The relative expression levels of six DEGs were further confirmed through quantitative real-time polymerase chain reaction (qRT-PCR).ConclusionNaHS alleviated Cd stress mainly through inhibiting Cd absorbtion, promoting CdS nanoparticles formation, increasing oxidation resistance, and regulation of transport in free-living unicellular T. thermophila. These findings will expand our understanding for H2S functions in the freshwater protozoa.

Project description:The ciliate Tetrahymena thermophila is a model organism for molecular and cellular biology. Like other ciliates, this species has separate germline and soma functions that are embodied by distinct nuclei within a single cell. The germline-like micronucleus (MIC) has its genome held in reserve for sexual reproduction. The soma-like macronucleus (MAC), which possesses a genome processed from that of the MIC, is the center of gene expression and does not directly contribute DNA to sexual progeny. We report here the shotgun sequencing, assembly, and analysis of the MAC genome of T. thermophila, which is approximately 104 Mb in length and composed of approximately 225 chromosomes. Overall, the gene set is robust, with more than 27,000 predicted protein-coding genes, 15,000 of which have strong matches to genes in other organisms. The functional diversity encoded by these genes is substantial and reflects the complexity of processes required for a free-living, predatory, single-celled organism. This is highlighted by the abundance of lineage-specific duplications of genes with predicted roles in sensing and responding to environmental conditions (e.g., kinases), using diverse resources (e.g., proteases and transporters), and generating structural complexity (e.g., kinesins and dyneins). In contrast to the other lineages of alveolates (apicomplexans and dinoflagellates), no compelling evidence could be found for plastid-derived genes in the genome. UGA, the only T. thermophila stop codon, is used in some genes to encode selenocysteine, thus making this organism the first known with the potential to translate all 64 codons in nuclear genes into amino acids. We present genomic evidence supporting the hypothesis that the excision of DNA from the MIC to generate the MAC specifically targets foreign DNA as a form of genome self-defense. The combination of the genome sequence, the functional diversity encoded therein, and the presence of some pathways missing from other model organisms makes T. thermophila an ideal model for functional genomic studies to address biological, biomedical, and biotechnological questions of fundamental importance.

Dataset Information

Transcriptome analysis of the model protozoan, Tetrahymena thermophila, using Deep RNA sequencing.

Publications

Transcriptome analysis of the model protozoan, Tetrahymena thermophila, using Deep RNA sequencing.

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