Cell cycle dependent regulation and function of Argonaute 1 in plants
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA397459 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR5907650.fastq.gz | Fastqsanger.gz | |||
| SRR5907660.fastq.gz | Fastqsanger.gz | |||
| SRR5907670.fastq.gz | Fastqsanger.gz | |||
| SRR5907680.fastq.gz | Fastqsanger.gz | |||
| SRR5907900.fastq.gz | Fastqsanger.gz |
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Project description:Cell cycle dependent regulation and function of Argonaute 1 in plants [RNA-Seq]
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Project description:The RNA silencing (RNAi) is a biological process, which plays a key role in a wide range of eukaryotes, such as gene expression regulation, transposable elements control and antiviral defense. In human, several studies showed that miRNAs and AGO2 are involved in the regulation of the cell cycle. Indeed, many cell cycle regulators such as cyclins, CDKs, CDK inhibitors, RBR proteins or E2F transcription factors have been identified as miRNA targets. Transcriptional data suggest that these cell cycle miRNAs are differentially accumulated during cell cycle progression and that misregulation of some could lead to proliferative diseases. However, in plants, almost nothing is known about the cell cycle regulation through miRNAs. Unpublished datas from the lab suggest that the Arabidopsis AGO1, which is the most important Argonaute for miRNA-mediated gene regulation, is essential for cell proliferation. In order to investigate the role of AGO1 during cell cycle, we set up a system based on cell cycle synchronization of tobacco BY-2 cell suspensions expressing a GFP-fused AGO1. We sequenced small RNAs from S, G2, M and G1 phases of synchronized cells from total RNA extract and GFP-AGO1 co-purified small RNAs, and investigate whether putative cell cycle specific miRNAs can be highlighted.
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Project description:The RNA silencing (RNAi) is a biological process, which plays a key role in a wide range of eukaryotes, such as gene expression regulation, transposable elements control and antiviral defense. In human, several studies showed that miRNAs and AGO2 are involved in the regulation of the cell cycle. Indeed, many cell cycle regulators such as cyclins, CDKs, CDK inhibitors, RBR proteins or E2F transcription factors have been identified as miRNA targets. Transcriptional data suggest that these cell cycle miRNAs are differentially accumulated during cell cycle progression and that misregulation of some could lead to proliferative diseases. However, in plants, almost nothing is known about the cell cycle regulation through miRNAs. Unpublished datas from the lab suggest that the Arabidopsis AGO1, which is the most important Argonaute for miRNA-mediated gene regulation, is essential for cell proliferation. In order to investigate the role of AGO1 during cell cycle, we set up a system based on cell cycle synchronization of tobacco BY-2 cell suspensions expressing a GFP-fused AGO1. We sequenced small RNAs from S, G2, M and G1 phases of synchronized cells from total RNA extract and GFP-AGO1 co-purified small RNAs, and investigate whether putative cell cycle specific miRNAs can be highlighted.
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Project description:The RNA silencing (RNAi) is a biological process, which plays a key role in a wide range of eukaryotes, such as gene expression regulation, transposable elements control and antiviral defense. In human, several studies showed that miRNAs and AGO2 are involved in the regulation of the cell cycle. Indeed, many cell cycle regulators such as cyclins, CDKs, CDK inhibitors, RBR proteins or E2F transcription factors have been identified as miRNA targets. Transcriptional data suggest that these cell cycle miRNAs are differentially accumulated during cell cycle progression and that misregulation of some could lead to proliferative diseases. However, in plants, almost nothing is known about the cell cycle regulation through miRNAs. Unpublished datas from the lab suggest that the Arabidopsis AGO1, which is the most important Argonaute for miRNA-mediated gene regulation, is essential for cell proliferation. In order to investigate the role of AGO1 during cell cycle, we set up a system based on cell cycle synchronization of tobacco BY-2 cell suspensions expressing a GFP-fused AGO1. We sequenced small RNAs from S, G2, M and G1 phases of synchronized cells from total RNA extract and GFP-AGO1 co-purified small RNAs, and investigate whether putative cell cycle specific miRNAs can be highlighted.
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Project description:An Argonaute phosphorylation cycle promotes microRNA-mediated silencing
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Project description:MicroRNAs (miRNAs) perform critical functions in normal physiology and disease by associating with Argonaute proteins and downregulating partially complementary messenger RNAs (mRNAs). To identify new regulators of the miRNA pathway, we employed CRISPR-Cas9 genome-wide loss-of-function screening coupled with a fluorescent reporter of miRNA activity. Iterative rounds of screening revealed a novel mechanism whereby target engagement by Argonaute 2 (AGO2) triggers its hierarchical, multi-site phosphorylation by CSNK1A1 on a set of highly conserved residues (S824-S834), followed by rapid dephosphorylation by the ANKRD52-PPP6C phosphatase complex. Although genetic and biochemical studies demonstrated that AGO2 phosphorylation on these residues inhibits target mRNA binding, inactivation of this phosphorylation cycle globally impairs miRNA-mediated silencing. Analysis of the transcriptome-wide binding profile of non-phosphorylatable AGO2 revealed a dramatic expansion of the target repertoire bound at steady-state, effectively reducing the active pool of AGO2 on a per target basis. These findings support a model in which an AGO2 phosphorylation cycle stimulated by target engagement regulates miRNA:target interactions to maintain the global efficiency of miRNA-mediated silencing.
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Project description:MicroRNAs (miRNAs) perform critical functions in normal physiology and disease by associating with Argonaute proteins and downregulating partially complementary messenger RNAs (mRNAs). To identify new regulators of the miRNA pathway, we employed CRISPR-Cas9 genome-wide loss-of-function screening coupled with a fluorescent reporter of miRNA activity. Iterative rounds of screening revealed a novel mechanism whereby target engagement by Argonaute 2 (AGO2) triggers its hierarchical, multi-site phosphorylation by CSNK1A1 on a set of highly conserved residues (S824-S834), followed by rapid dephosphorylation by the ANKRD52-PPP6C phosphatase complex. Although genetic and biochemical studies demonstrated that AGO2 phosphorylation on these residues inhibits target mRNA binding, inactivation of this phosphorylation cycle globally impairs miRNA-mediated silencing. Analysis of the transcriptome-wide binding profile of non-phosphorylatable AGO2 revealed a dramatic expansion of the target repertoire bound at steady-state, effectively reducing the active pool of AGO2 on a per target basis. These findings support a model in which an AGO2 phosphorylation cycle stimulated by target engagement regulates miRNA:target interactions to maintain the global efficiency of miRNA-mediated silencing.
2017-01-17 | GSE89943 | GEO
Project description:MicroRNAs (miRNAs) perform critical functions in normal physiology and disease by associating with Argonaute proteins and downregulating partially complementary messenger RNAs (mRNAs). To identify new regulators of the miRNA pathway, we employed CRISPR-Cas9 genome-wide loss-of-function screening coupled with a fluorescent reporter of miRNA activity. Iterative rounds of screening revealed a novel mechanism whereby target engagement by Argonaute 2 (AGO2) triggers its hierarchical, multi-site phosphorylation by CSNK1A1 on a set of highly conserved residues (S824-S834), followed by rapid dephosphorylation by the ANKRD52-PPP6C phosphatase complex. Although genetic and biochemical studies demonstrated that AGO2 phosphorylation on these residues inhibits target mRNA binding, inactivation of this phosphorylation cycle globally impairs miRNA-mediated silencing. Analysis of the transcriptome-wide binding profile of non-phosphorylatable AGO2 revealed a dramatic expansion of the target repertoire bound at steady-state, effectively reducing the active pool of AGO2 on a per target basis. These findings support a model in which an AGO2 phosphorylation cycle stimulated by target engagement regulates miRNA:target interactions to maintain the global efficiency of miRNA-mediated silencing.
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Project description:MicroRNAs (miRNAs) perform critical functions in normal physiology and disease by associating with Argonaute proteins and downregulating partially complementary messenger RNAs (mRNAs). To identify new regulators of the miRNA pathway, we employed CRISPR-Cas9 genome-wide loss-of-function screening coupled with a fluorescent reporter of miRNA activity. Iterative rounds of screening revealed a novel mechanism whereby target engagement by Argonaute 2 (AGO2) triggers its hierarchical, multi-site phosphorylation by CSNK1A1 on a set of highly conserved residues (S824-S834), followed by rapid dephosphorylation by the ANKRD52-PPP6C phosphatase complex. Although genetic and biochemical studies demonstrated that AGO2 phosphorylation on these residues inhibits target mRNA binding, inactivation of this phosphorylation cycle globally impairs miRNA-mediated silencing. Analysis of the transcriptome-wide binding profile of non-phosphorylatable AGO2 revealed a dramatic expansion of the target repertoire bound at steady-state, effectively reducing the active pool of AGO2 on a per target basis. These findings support a model in which an AGO2 phosphorylation cycle stimulated by target engagement regulates miRNA:target interactions to maintain the global efficiency of miRNA-mediated silencing.
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