The kinetic landscape of an RNA binding protein in cells
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PROVIDER: PRJNA631519 | ENA |
REPOSITORIES: ENA
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Project description:The binding and dissociation of RNA Binding Proteins (RBPs) to their cognate sites on cellular RNAs are critical for the regulation of gene expression. Yet, association and dissociation kinetics of RBPs at individual cellular binding sites have not been experimentally accessible. Inaccessibility of binding and dissociation kinetics of RBPs in cells severely limits or even precludes the establishment of quantitative connections between RBP-RNA interactions and cellular RBP function. Here, we describe an approach to measure binding and dissociation kinetics of the RBP Dazl at thousands of individual binding sites in cells. We then show how these kinetic parameters inform a quantitative understanding of the cellular function of Dazl.
2020-12-16 | GSE150214 | GEO
Project description:Gene expression in higher eukaryotic cells orchestrates interactions between thousands of RNA-binding proteins (RBPs) and tens of thousands of RNAs1. The kinetics by which RBPs bind to and dissociate from their RNA sites are critical for the coordination of cellular RNA-protein interactions2. However, these kinetic parameters have not been experimentally measured in cells. Here we show that time-resolved RNA-protein cross-linking with a pulsed femtosecond ultraviolet laser, followed by immunoprecipitation and high-throughput sequencing, allows the determination of binding and dissociation kinetics of the RBP DAZL for thousands of individual RNA-binding sites in cells. This kinetic cross-linking and immunoprecipitation (KIN-CLIP) approach reveals that DAZL resides at individual binding sites for time periods of only seconds or shorter, whereas the binding sites remain DAZL-free for markedly longer. The data also indicate that DAZL binds to many RNAs in clusters of multiple proximal sites. The effect of DAZL on mRNA levels and ribosome association correlates with the cumulative probability of DAZL binding in these clusters. Integrating kinetic data with mRNA features quantitatively connects DAZL-RNA binding to DAZL function. Our results show how kinetic parameters for RNA-protein interactions can be measured in cells, and how these data link RBP-RNA binding to the cellular function of RBPs.
| S-EPMC8299502 | biostudies-literature
Project description:Adaptation of iCLIP to plants determines the binding landscape of the clock-regulated RNA-binding protein AtGRP7
| PRJNA388894 | ENA
Project description:1. evaluation of diagnostic importance of insulin like growth factor binding protein3 in patient with recently diagnosed as Colorectal cancer
2. correlation between the diagnostic efficacy of insulin like growth factor binding protein 3 with routine marker carcinoembryonic antigen.
| 59449 | ecrin-mdr-crc
Project description:RNA-binding proteins play a key role in shaping gene expression profiles during stress, however, little is known about the dynamic nature of these interactions and how this influences the kinetics of gene expression. To address this, we developed kinetic χCRAC, a UV cross-linking method that enabled us to quantitatively measure the dynamics of protein-RNA interactions in vivo on a minute time-scale. Here, using kinetic χCRAC we measure the global RNA-binding dynamics of the yeast transcription termination factor Nab3 in response to glucose starvation. These measurement reveal rapid changes in protein-RNA interactions within one minute following stress imposition. Changes in Nab3 binding are largely independent of alterations in transcription rate during the early stages of stress response, indicating orthogonal transcriptional control mechanisms. We also uncover a function for Nab3 in dampening expression of stress-responsive genes. Kinetic χCRAC has the potential to greatly enhance our understanding of in vivo dynamics of protein-RNA interactions.
2017-04-11 | GSE85545 | GEO
Project description:iCLIP of the RNA-binding protein ARPP21
| PRJEB21898 | ENA
Project description:RNA-binding protein immunoprecipitation (RIP)and RNA sequencing
| PRJNA555809 | ENA
Project description:RFX7 DNA binding landscape in U2OS
| PRJNA680774 | ENA
Project description:RFX7 DNA binding landscape in HCT116
| PRJNA680775 | ENA
Project description:The vast landscape of RNA-protein interactions at the heart of post-transcriptional regulation remains largely unexplored. Indeed it is likely that, even in yeast, a substantial fraction of the regulatory RNA-binding proteins (RBPs) remain to be discovered. Systematic experimental methods can play a key role in discovering these RBPs - most of the known yeast RBPs lack RNA-binding domains that might enable this activity to be predicted. We describe here a new proteome-wide approach to identify RNA-protein interactions based on in vitro binding of RNA samples to yeast protein microarrays that represent over 80% of the yeast proteome. We used this procedure to screen for novel RBPs and RNA-protein interactions. A complementary mass spectrometry technique also identified proteins that associate with yeast mRNAs. Both the protein microarray and mass spectrometry methods successfully identify previously annotated RBPs, suggesting that other proteins identified in these assays might be novel RBPs. Of 35 putative novel RBPs identified by either or both of these methods, 12, including 75% of the eight most highly-ranked candidates, reproducibly associated with specific cellular RNAs. Surprisingly, most of the 12 newly discovered RBPs were enzymes. Functional characteristics of the RNA targets of some of the novel RBPs suggest coordinated post-transcriptional regulation of subunits of protein complexes and a possible link between mRNA trafficking and vesicle transport. Our results suggest that many more RBPs still remain to be identified and provide a set of candidates for further investigation. Set of arrays organized by shared biological context, such as organism, tumors types, processes, etc.
2011-03-10 | E-GEOD-22876 | biostudies-arrayexpress