Transcriptomic analysis of the Xrn1 depletion by Auxin-degron shutoff
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PROVIDER: PRJNA508323 | ENA |
REPOSITORIES: ENA
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Project description:The goal of the project was to study the effects on transcription and mRNA stability of the Xrn1 sudden depletion. We analyzed the effect of Xrn1 depletion caused by protein degradation of an Auxin-degron fusion on the transcription rates, mRNA stabilities and mRNA levels by doing Genomic Run-On (GRO) experiments at 30 min after Auxin addition with a control at 0 min.
2019-02-22 | GSE123326 | GEO
Project description:To study the role of the exonuclease Xrn1 in translational control, we performed ribosome profiling and RNA-seq in Xrn1-depleted cells. By using an auxin-inducible degron, we were able to study immediate effects of Xrn1 depletion in translational control. Therefore, we could overcome experimental limitations associated to stable deletion mutants.
2019-02-08 | GSE109734 | GEO
Project description:To study the role of the exonuclease Xrn1 in gene expression dynamics under osmotic stress conditions, we performed RNA-seq in Xrn1-depleted cells. By using an auxin-inducible degron, we were able to study immediate effects of Xrn1 depletion in gene expression dynamics. Therefore, we could overcome experimental limitations associated to stable deletion mutants.
2023-01-28 | GSE144440 | GEO
Project description:Cullin RING-type E3 ubiquitin ligase SCFTIR1/AFB1-5 and their ubiquitylation targets, AUX/IAAs, sense auxin concentrations in the nucleus. TIR1 binds a surface- exposed degron in AUX/IAAs promoting their ubiquitylation and rapid auxin-regulated proteasomal degradation. Here, we resolved TIR1·auxin·IAA7 and TIR1·auxin·IAA12 complex topology, and show that flexible intrinsically disordered regions (IDRs) cooperatively position AUX/IAAs on TIR1. The AUX/IAA PB1 interaction domain also assists in non-native contacts, affecting AUX/IAA dynamic interaction states. Our results establish a role for IDRs in modulating auxin receptor assemblies. By securing AUX/IAAs on two opposite surfaces of TIR1, IDR diversity supports locally tailored positioning for targeted ubiquitylation, and might provide conformational flexibility for adopting a multiplicity of functional states. We postulate IDRs in distinct members of the AUX/IAA family to be an adaptive signature for protein interaction and initiation region for proteasome recruitment.
2020-05-27 | PXD015392 | Pride
Project description:Auxin-degron system identifies immediate mechanisms of Oct4
| PRJNA708329 | ENA
Project description:Auxin-inducible degron (AID) technology is powerful for chemogenetic control of proteolysis. However, generation of human cell lines to deplete endogenous proteins with AID remains challenging. Typically, homozygous degron-tagging efficiency is low and overexpression of an auxin receptor requires additional engineering steps. Here, we establish a one-step genome editing procedure with high-efficiency homozygous tagging and auxin receptor expression. We demonstrate its application in 5 human cell lines, including embryonic stem (ES) cells. The method allowed isolation of AID single-cell clones in 10 days for 11 target proteins with >80% average homozygous degron-tagging efficiency in A431 cells, and >50% efficiency for 5 targets in H9 ES cells. The tagged endogenous proteins were inducibly degraded in all cell lines, including ES cells and ES-cell derived neurons, with robust expected functional readouts. This method facilitates the application of AID for studying endogenous protein functions in human cells, especially in stem cells.
2024-01-31 | GSE243637 | GEO
Project description:We obtained RNA polymerase II occupancy profiles across the genome of S.cerevisiae strains: Abf1 anchor-away cells after addition of rapamycin for different time points (including no addition of rapamycin) and Rap1-AID auxin-degron cells after addition of auxin for different time points (including no addition of auxin) . This allowed us to compare polymerase occupancy profiles when these proteins are depleted from the nucleus or degraded.
2018-12-20 | GSE114589 | GEO
Project description:We examined 3D chromatin structure in the absence of cohesin (Scc1-AID auxin-inducible degron) and a control line (E14-Tir1) and found that PRC1 core promoter component RING1B was one of the most enriched proteins. Hence, we performed calibrated ChIP-seq experiments on the control (E14-Tir1) and the Scc1-AID mESC lines with a spike in of HEK293 human cells to further study this relationship.
2020-01-23 | E-MTAB-7817 | biostudies-arrayexpress
Project description:The effect of Sfp1 depeltion at transcriptional level was investigated by construting a AID-Sfp1 strain (Auxin sensitive degron) that allowed to shutoff the Sfp1 expression and analyze the primary effects on synthesis rates (SR) and mRNA levels (RA) genome-wide. mRNA stabilities can be calculated as RA /SR for each gene in all samples.
2023-09-21 | GSE202748 | GEO
Project description:Background: CTCF is a well-established chromatin architectural protein that also plays various roles in transcriptional regulation. While CTCF biology has been extensively studied, how the domains of CTCF function to regulate transcription remains unknown. Additionally, the original auxin-inducible degron 1 (AID1) system has limitations to investigate the function of CTCF. Results: We employ an improved auxin-inducible degron technology, AID2, to facilitate the study of acute depletion of CTCF while overcoming the limitations of the previous AID system. As previously observed through the AID1 system and steady-state RNA analysis, the new AID2 system combined with SLAM-seq confirms that CTCF depletion leads to modest nascent and steady-state transcripts changes. A CTCF domain sgRNA library screening identifies the zinc finger (ZF) domain as the region within CTCF with the most functional relevance, including ZFs 1 and 10. Removal of ZFs 1 and 10 reveals genomic regions that independently require these ZFs for DNA binding and transcriptional regulation. Notably, loci regulated by either ZF1 or ZF10 exhibit unique CTCF binding motifs specific to each ZF. Conclusions: By extensively comparing the AID1 and AID2 systems for CTCF degradation in SEM cells, we confirm that AID2 degradation is superior for achieving miniAID tagged protein degradation without the limitations of the AID1 system. The model we create that combines AID2 depletion of CTCF with exogenous overexpression of CTCF mutants allows us to demonstrate how peripheral ZFs intricately orchestrate transcriptional regulation in a cellular context for the first time.
2022-12-09 | GSE201911 | GEO