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Genome-wide identification and characterization of transcription start sites and promoters in the tunicate Ciona intestinalis.

ABSTRACT: The tunicate Ciona intestinalis, an invertebrate chordate, has recently emerged as a powerful model organism for gene regulation analysis. However, few studies have been conducted to identify and characterize its transcription start sites (TSSs) and promoters at the genome-wide level. Here, using TSS-seq, we identified TSSs at the genome-wide scale and characterized promoters in C. intestinalis. Specifically, we identified TSS clusters (TSCs), high-density regions of TSS-seq tags, each of which appears to originate from an identical promoter. TSCs were found not only at known TSSs but also in other regions, suggesting the existence of many unknown transcription units in the genome. We also identified candidate promoters of 79 ribosomal protein (RP) genes, each of which had the major TSS in a polypyrimidine tract and showed a sharp TSS distribution like human RP gene promoters. Ciona RP gene promoters, however, did not appear to have typical TATA boxes, unlike human RP gene promoters. In Ciona non-RP promoters, two pyrimidine-purine dinucleotides, CA and TA, were frequently used as TSSs. Despite the absence of CpG islands, Ciona TATA-less promoters showed low expression specificity like CpG-associated human TATA-less promoters. By using TSS-seq, we also predicted trans-spliced gene TSSs and found that their downstream regions had higher G+T content than those of non-trans-spliced gene TSSs. Furthermore, we identified many putative alternative promoters, some of which were regulated in a tissue-specific manner. Our results provide valuable information about TSSs and promoter characteristics in C. intestinalis and will be helpful in future analysis of transcriptional regulation in chordates.

SUBMITTER: Yokomori R

PROVIDER: S-EPMC4691747 | biostudies-literature | 2016 Jan

REPOSITORIES: biostudies-literature

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Genome-wide identification and characterization of transcription start sites and promoters in the tunicate Ciona intestinalis.

Yokomori Rui R Shimai Kotaro K Nishitsuji Koki K Suzuki Yutaka Y Kusakabe Takehiro G TG Nakai Kenta K

Genome research 20151214 1

The tunicate Ciona intestinalis, an invertebrate chordate, has recently emerged as a powerful model organism for gene regulation analysis. However, few studies have been conducted to identify and characterize its transcription start sites (TSSs) and promoters at the genome-wide level. Here, using TSS-seq, we identified TSSs at the genome-wide scale and characterized promoters in C. intestinalis. Specifically, we identified TSS clusters (TSCs), high-density regions of TSS-seq tags, each of which ...[more]

PMID: 26668163

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Project description:Despite almost 40 years of molecular genetics research in Escherichia coli a major fraction of its Transcription Start Sites (TSSs) are still unknown, limiting therefore our understanding of the regulatory circuits that control gene expression in this model organism. RegulonDB (http://regulondb.ccg.unam.mx/) is aimed at integrating the genetic regulatory network of E. coli K12 as an entirely bioinformatic project up till now. In this work, we extended its aims by generating experimental data at a genome scale on TSSs, promoters and regulatory regions. We implemented a modified 5' RACE protocol and an unbiased High Throughput Pyrosequencing Strategy (HTPS) that allowed us to map more than 1700 TSSs with high precision. From this collection, about 230 corresponded to previously reported TSSs, which helped us to benchmark both our methodologies and the accuracy of the previous mapping experiments. The other ca 1500 TSSs mapped belong to about 1000 different genes, many of them with no assigned function. We identified promoter sequences and type of sigma factors that control the expression of about 80% of these genes. As expected, the housekeeping sigma(70) was the most common type of promoter, followed by sigma(38). The majority of the putative TSSs were located between 20 to 40 nucleotides from the translational start site. Putative regulatory binding sites for transcription factors were detected upstream of many TSSs. For a few transcripts, riboswitches and small RNAs were found. Several genes also had additional TSSs within the coding region. Unexpectedly, the HTPS experiments revealed extensive antisense transcription, probably for regulatory functions. The new information in RegulonDB, now with more than 2400 experimentally determined TSSs, strengthens the accuracy of promoter prediction, operon structure, and regulatory networks and provides valuable new information that will facilitate the understanding from a global perspective the complex and intricate regulatory network that operates in E. coli.

Project description:Eight different types of sample (three samples each) were used in this study of cardiac cell migration in the tunicate Ciona intestinalis. The B7.5 lineage cells were isolated by Fluorescent Activated Cell Sorting (FACS) at the tailbud stage, when the cardiac precursors (anterior B7.5 lineage cells) are migrating into the trunk, while their sister cells (posterior 7.5 lineage cells) remain in the tail, where they form muscle cells. Cells were sorted at the late gastrula stage, prior to migration induction and after manipulation of the FGF-MAPK-Ets signaling pathway (targeted expression of dominant negative FGF receptor and Ets:VP16 fusion protein, targeted expression was achieved using the Mesp cis-regulatory DNA), FoxF function (a direct target of FGF-MAPK-Ets signaling, required primarily for cell migration) by targeted expression of FoxF:VP16 (constitutive activator) or FoxF:WRPW (constitutive repressor, dominant negative), Mesp function (over-expression of Mesp:VP16 inhibits primarily cell migration). Finally, the "whole embryo" samples were collected from dissociated whole tailbud embryos, without FACS. in summary the eight conditions are: "wt (files LC-wt-1,-2,-3)": wild_type B7.5 lineage cells, expressing GFP, sorted at the tailbud stage "Ets:VP16 (files LC-EV-1,-2,-3)": B7.5 lineage cells, expressing GFP and Ets:VP16, sorted at the tailbud stage "DnFGFR (files LC-dnFGFR-1,-2,-3)": B7.5 lineage cells, expressing GFP and dominant-negative FGF receptor, sorted at the tailbud stage. "FoxF:VP16 (files LC-FV-1,-2,-3)": B7.5 lineage cells, expressing GFP and FoxF:VP16, sorted at the tailbud stage. "FoxF:WRPW (files LC-FW-1,-2,-3)": B7.5 lineage cells, expressing GFP and FoxF:WRPW, sorted at the tailbud stage". "Mesp:VP16 (files LC-MV-1,-2,-3)": B7.5 lineage cells, expressing GFP and Mesp:VP16, sorted at the tailbud stage. "Late Gastrula (files LC-LG-1,-2,-3)": wild_type B7.5 lineage cells, expressing GFP, sorted at the late gastrula stage "whole embryo (files LC-whole-1,-2,-3": whole embryos, dissociated at the tailbud stage, not sorted.

Dataset Information

Genome-wide identification and characterization of transcription start sites and promoters in the tunicate Ciona intestinalis.

Publications

Genome-wide identification and characterization of transcription start sites and promoters in the tunicate Ciona intestinalis.

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