Addition-elimination mechanism activated nucleotide transition sequencing for RNA dynamics profiling
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ABSTRACT:
PROVIDER: PRJNA707546 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR13893142_1.fastq.gz | Fastqsanger.gz | |||
| SRR13893142_2.fastq.gz | Fastqsanger.gz | |||
| SRR13893143_1.fastq.gz | Fastqsanger.gz | |||
| SRR13893143_2.fastq.gz | Fastqsanger.gz | |||
| SRR13893144_1.fastq.gz | Fastqsanger.gz |
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Project description:The temporal information of intracellular transcripts is essential to understand their functional roles, yet routine RNA-seq methods only measure RNA species at a steady state. Here we develop addition-elimination mechanism activated nucleotide transition sequencing (AENT-seq) for transcriptome-wide profiling of RNA dynamics. In AENT-seq, intracellular transcripts are metabolically labeled with 4-thiouridine (4sU), and then isolated total RNA are subjected to 4sU-to-cytosine (C) transition reactions based on a mild addition-elimination chemistry. N2H4•H2O is demonstrated to convert 4sU to 4-hydrazino cytosine (C*) in RNA-friendly aqueous conditions. The C* is recorded as natural C during extension reactions with a high efficiency of ~95%. This 4sU-to-C transition marks 4sU-containing nascent transcripts, and it enables subsequent sequencing analysis of RNA dynamics with single-nucleotide resolution. We applies our AENT-seq to simply investigate transcript dynamic information of several genes involved in cancer progression and metastasis. This method avoids harsh chemical conditions and harmful reagents involved in previous approaches, allowing rapid dissemination and extensive applications of the technique.
2021-03-09 | GSE168485 | GEO
Project description:Epigenetic inheritance of acquired traits is widespread among eukaryotes but how and to what extent such information is trans-generationally inherited is still unclear. The patterns of programmed DNA elimination in ciliates are epigenetically and trans-generationally inherited, and it has been proposed that small RNAs, which shuttle between the germline and the soma, regulate this epigenetic inheritance. In this study, we test the existence of such small RNA-mediated communication by epigenetically disturbing the pattern of DNA elimination in Tetrahymena. We show that the pattern of DNA elimination is indeed determined by the selective turnover of small RNAs, which is induced by the interaction between germline-derived small RNAs and the somatic genome. In addition, we show that DNA elimination of an element is regulated by small RNA-mediated communication with other eliminated elements. By contrast, no evidence obtained thus far supports the notion that transfer of epigenetic information from the soma to the germline, if any, regulates DNA elimination. Our results indicate that small RNA-mediated trans-nuclear and trans-element communication, in addition to unknown information in the germline genome, contributes to determining the pattern of DNA elimination.
2018-05-15 | GSE111525 | GEO
Project description:Programmed DNA elimination is a developmentally regulated process leading to the reproducible loss of specific genomic sequences. DNA elimination occurs in unicellular ciliates and a variety of metazoans, including invertebrates and vertebrates. In metazoa, DNA elimination typically occurs in somatic cells during early development, leaving the germline genome intact. Reference genomes for metazoa that undergo DNA elimination are not available. Here, we generated germline and somatic reference genome sequences of the DNA eliminating pig parasitic nematode Ascaris suum and the horse parasite Parascaris univalens. In addition, we carried out in-depth analyses of DNA elimination in the parasitic nematode of humans, Ascaris lumbricoides, and the parasitic nematode of dogs, Toxocara canis. Our analysis of nematode DNA elimination reveals that in all species, repetitive sequences (that differ among the genera) and germline-expressed genes (approximately 1000-2000 or 5%-10% of the genes) are eliminated. Thirty-five percent of these eliminated genes are conserved among these nematodes, defining a core set of eliminated genes that are preferentially expressed during spermatogenesis. Our analysis supports the view that DNA elimination in nematodes silences germline-expressed genes. Over half of the chromosome break sites are conserved between Ascaris and Parascaris, whereas only 10% are conserved in the more divergent T. canis. Analysis of the chromosomal breakage regions suggests a sequence-independent mechanism for DNA breakage followed by telomere healing, with the formation of more accessible chromatin in the break regions prior to DNA elimination. Our genome assemblies and annotations also provide comprehensive resources for analysis of DNA elimination, parasitology research, and comparative nematode genome and epigenome studies.
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Project description:Programmed DNA elimination is a developmentally regulated process leading to the reproducible loss of specific genomic sequences. DNA elimination occurs in unicellular ciliates and a variety of metazoans, including invertebrates and vertebrates. In metazoa, DNA elimination typically occurs in somatic cells during early development, leaving the germline genome intact. Reference genomes for metazoa that undergo DNA elimination are not available. Here, we generated germline and somatic reference genome sequences of the DNA eliminating pig parasitic nematode Ascaris suum and the horse parasite Parascaris univalens. In addition, we carried out in-depth analyses of DNA elimination in the parasitic nematode of humans, Ascaris lumbricoides, and the parasitic nematode of dogs, Toxocara canis. Our analysis of nematode DNA elimination reveals that in all species, repetitive sequences (that differ among the genera) and germline-expressed genes (approximately 1000-2000 or 5%-10% of the genes) are eliminated. Thirty-five percent of these eliminated genes are conserved among these nematodes, defining a core set of eliminated genes that are preferentially expressed during spermatogenesis. Our analysis supports the view that DNA elimination in nematodes silences germline-expressed genes. Over half of the chromosome break sites are conserved between Ascaris and Parascaris, whereas only 10% are conserved in the more divergent T. canis. Analysis of the chromosomal breakage regions suggests a sequence-independent mechanism for DNA breakage followed by telomere healing, with the formation of more accessible chromatin in the break regions prior to DNA elimination. Our genome assemblies and annotations also provide comprehensive resources for analysis of DNA elimination, parasitology research, and comparative nematode genome and epigenome studies.
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Project description:Organisms invest significant effort into maintaining genome stability. However, in diverse groups of eukaryotes, portions or entire chromosomes are lost in early development or during sex determination, a process known as programmed DNA elimination. Little is known about how different segments of the genome are reproducibly retained and discarded during programmed DNA elimination. We tested the hypothesis that selective retention is mediated by regulation of centromere-mediated association of chromosome segments with the mitotic spindle. We report that on the holocentric chromosomes of the nematode Ascaris, the core centromeric histone CENP-A is localized differently in cells undergoing DNA elimination from those undergoing germline mitosis. Prior to DNA elimination, CENP-A is significantly reduced in chromosome regions that will be lost. CENP-A reduction in eliminated genomic regions leads to the absence of kinetochores and microtubule attachment sites necessary for chromosome segregation, and thus the loss of these DNA regions during Ascaris programmed DNA elimination. Our results show that holocentric chromosome organization in Ascaris is regulated and that changes in CENP-A deposition specify which portions of chromosomes will be eliminated during programmed DNA elimination. A total of 62 samples are analyzed. These include: (1). CENP-A ChIP-seq on 12 developmental stages with input and replicates (12 x 2 x 2 samples); (2). CENP-C ChIP-seq on 3 developmental stages with input and replicates (total 8 samples); and (3). Histone marks ChIP-seq with 6 samples
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2023-07-21 | PXD041832 | JPOST Repository
Project description:Analysis of genome-wide IES elimination of Late-scnRNA accumulation-defective cells inducates that Early-scnRNAs are sufficient to induce DNA elimination for a majority of IESs, whereas Late-scnRNAs are important for DNA elimination of some, mainly Type-B, IESs.
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Project description:Competitive cell-interactions play a crucial role in quality control during development and homeostasis. Here we show that cancer cells use such interactions to actively eliminate wild-type intestine cells in enteroid monolayers and organoids. This apoptosis-dependent process boosts proliferation of intestinal cancer cells. The remaining wild-type population activates markers of primitive epithelia and transits to a fetal-like state. Prevention of this cell fate transition avoids elimination of wild-type cells and, importantly, limits the proliferation of cancer cells. JNK signaling is activated in competing cells and is required for cell fate change and elimination of wild-type cells. Thus, cell competition drives growth of cancer cells by active out-competition of wild-type cells through forced cell death and cell fate change in a JNK dependent manner.
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