Project description:We present a comprehensive transcriptome of ciliate T. thermophila using the Illumina RNA-seq platform. The data was generated from the six mRNA samples of growth, starvation and conjugation of Tetrahymena. Despite an AT rich genome, there are about 124.6 million reads mapped to T. thermophila genome. Using these mapped reads, we have significantly improved the previous genome annotation and investigated the gene expression. Besides, our result also provided a comprehensive understanding of the alternative splicing in T. thermophila, and suggested the existence of the regulated unproductive splicing and translation (RUST) in the single-celled eukaryote. RNA-seq for six samples of Tetrahymena growth, starvation and conjugation.
Project description:We present a comprehensive transcriptome of ciliate T. thermophila using the Illumina RNA-seq platform. The data was generated from the six mRNA samples of growth, starvation and conjugation of Tetrahymena. Despite an AT rich genome, there are about 124.6 million reads mapped to T. thermophila genome. Using these mapped reads, we have significantly improved the previous genome annotation and investigated the gene expression. Besides, our result also provided a comprehensive understanding of the alternative splicing in T. thermophila, and suggested the existence of the regulated unproductive splicing and translation (RUST) in the single-celled eukaryote.
Project description:Eleven genome wide microarrays containing the predicted coding sequences (putative genes) for the ciliated protozoan Tetrahymena thermophila used to study gene expression in conjugation cells (C-0, C-15m, C-2, C-4, C-6, C-8, C-10, C-12, C-14, C-16, C-18). Combined these eleven microarrays with 50 microarrays described in Miao et al (2009) and other 6 microarrays, we constructed the Tetrahymena gene network (TGN) using three methods: the Pearson correlation coefficient, the Spearman correlation coefficient and the context likelihood of relatedness (CLR) algorithm. The accuracy and coverage of the three networks were evaluated using four conserved protein complexes in yeast, and the CLR network was found to be the best network, with a Z-score threshold 3.49. Then the TGN was partitioned, and 55 modules were found. In addition, analysis for the arbitrarily determined 1200 hubs showed that these hubs could be sorted into six groups according to expression profiles. We also investigated human disease orthologs in Tetrahymena that are missing in yeast and found evidence indicating that some of these were involved in the same process in Tetrahymena as in human. For conjugation, B2086 and CU428 cells that had been starved for 18 hours were resuspended in 10 mM Tris (pH 7.5) at 200,000 cells/ml, mixed, and samples were collected at 15 min, 2, 4, 6, 8, 10, 12, 14, 16, 18 hours after the mixture (referred to as C-0, C-15m, C-2, C-4, C-6, C-8, C-10, C-12, C-14, C-16 and C-18).
Project description:Somatic macronucleus (MAC) and germline micronucleus (MIC) of Tetrahymena thermophila are different in chromosome numbers, sizes, functions and cohesin complex locations. Loss of cohesin complex resulted in genome-wide disappearance of topologically associating domains (TADs) and chromatin loops in mammalian cells. However, the higher-level chromatin organization in Tetrahymena thermophila which contains both cohesin free MAC and cohesin located MIC are largely unknown. Here, using the Hi-C and HiChIP methods, we reveal that, these two nuclei possess distinct three-dimensional genome structures. In the MAC, each chromosome occupies its own territory and there are no chromatin compartmentalization or chromatin domains. The chromatin loops in MAC are mainly related to chromatin structures rather than transcriptional regulation. The MIC also without chromatin compartmentalization, but with chromatin domains and the domain boundaries are consistent with chromatin breakage sites (CBSs) which indicates that each MIC chromatin domain developed to one MAC chromosome during conjugation. Besides, we found the MIC exhibits unique intra-arm and inter-chromosome interactions at the crescent stage of conjugation, when the MIC undergoes meiotic recombination.
Project description:Meiotic recombination is carried out through a specialized pathway for the formation and repair of DNA double-strand breaks (DSBs) made by the Spo11 protein. The present study shed light on the functional role of Cyclin, CYC2, in Tetrahymena thermophila which has transcriptionally high expression level during meiosis process. Knocking out the CYC2 gene results in arrest of meiotic conjugation process at 2.5â??3.5 h after conjugation initiation, before the meiosis division starts, and in company with the absence of DSBs. To investigate the underlying mechanism of this phenomenon, a complete transcriptome profile was performed between wild-type strain and CYC2 knock-out strain. Functional analysis of RNA-Seq results identifies related differentially expressed genes (DEGs) including SPO11 and these DEGs are enriched in DNA repair/mismatch repair (MMR) terms in homologous recombination (HR), which indicates that CYC2 could play a crucial role in meiosis by regulating SPO11 and participating in HR. We performed mRNA profiling on both wild-type strains and CYC2-knocking out strains at four different stages during meiosis prohase of Tetrahymena thermophila.