MassIVE MSV000096455

Partial Public

SPIDR: a highly multiplexed method for mapping RNA-protein interactions uncovers a mechanism for selective translational suppression upon cellular stress

Subscribe Comment Reanalyze Spectra Add Reanalysis

Description

RNA binding proteins (RBPs) play crucial roles in regulating every stage of the mRNA life cycle and mediating non-coding RNA functions. Despite their importance, the specific roles of most RBPs remain unexplored because we do not know their specific RNA binding partners. Current methods, such as crosslinking and immunoprecipitation followed by sequencing (CLIP-seq), have expanded our knowledge of RBP-RNA interactions but are generally limited by their ability to map only one RBP at a time. To address this limitation, we developed SPIDR (Split and Pool Identification of RBP targets), a massively multiplexed method to simultaneously profile global RNA binding sites of dozens to hundreds of RBPs in a single experiment. SPIDR employs antibody-bead labeling coupled with split-pool barcoding to increase the throughput of current CLIP methods by two orders of magnitude. SPIDR identifies precise, single-nucleotide RNA binding sites for diverse classes of RBPs simultaneously. We identified several novel ribosomal RNA binders, including a novel interaction between LARP1 and 18S ribosomal RNA located within the mRNA entry site on the 40S small ribosomal subunit, and resolved this structure at 2.8 A using single-particle cryo-electron microscopy (cryo-EM). This structure provides a potential mechanistic explanation for the role of LARP1 in translational suppression. We next explored SPIDR ability to detect changes in RBP binding upon mTOR inhibition and identified that 4EBP1 acts as a dynamic RBP that selectively binds to 5-untranslated regions of translationally repressed mRNAs only upon mTOR inhibition. These observations provide a potential mechanism to explain the specificity of translational regulation controlled by mTOR signaling. SPIDR enables rapid, de novo discovery of RNA-protein interactions at an unprecedented scale and has the potential to transform our understanding of RNA biology and both transcriptional and post-transcriptional gene regulation. [doi:10.25345/C5SX64N69] [dataset license: CC0 1.0 Universal (CC0 1.0)]

Keywords: IP ; DDA ; DIA ; DatasetType:Proteomics

Contact

Principal Investigators:
(in alphabetical order) 		Marko Jovanovic, Columbia University, USA
Submitting User: ak3952
Number of Files:
Total Size:
Spectra:
Subscribers:
 
Owner Reanalyses
Experimental Design
    Conditions:
    Biological Replicates:
    Technical Replicates:
 
Identification Results
    Proteins (Human, Remapped):
    Proteins (Reported):
    Peptides:
    Variant Peptides:
    PSMs:
 
Quantification Results
    Differential Proteins:
    Quantified Proteins:
 
Browse Dataset Files
 
FTP Download Link (click to copy):

Species

Instrument

Modifications

- Dataset Reanalyses

+ Dataset History

Click here to queue conversion of this dataset's submitted spectrum files to open formats (e.g. mzML). This process may take some time.

When complete, the converted files will be available in the "ccms_peak" subdirectory of the dataset's FTP space (accessible via the "FTP Download" link to the right).
Number of distinct conditions across all analyses (original submission and reanalyses) associated with this dataset.

Distinct condition labels are counted across all files submitted in the "Metadata" category having a "Condition" column in this dataset.

"N/A" means no results of this type were submitted.
Number of distinct biological replicates across all analyses (original submission and reanalyses) associated with this dataset.

Distinct replicate labels are counted across all files submitted in the "Metadata" category having a "BioReplicate" or "Replicate" column in this dataset.

"N/A" means no results of this type were submitted.
Number of distinct technical replicates across all analyses (original submission and reanalyses) associated with this dataset.

The technical replicate count is defined as the maximum number of times any one distinct combination of condition and biological replicate was analyzed across all files submitted in the "Metadata" category. In the case of fractionated experiments, only the first fraction is considered.

"N/A" means no results of this type were submitted.
Originally identified proteins that were automatically remapped by MassIVE to proteins in the SwissProt human reference database.

"N/A" means no results of this type were submitted.
Number of distinct protein accessions reported across all analyses (original submission and reanalyses) associated with this dataset.

"N/A" means no results of this type were submitted.
Number of distinct unmodified peptide sequences reported across all analyses (original submission and reanalyses) associated with this dataset.

"N/A" means no results of this type were submitted.
Number of distinct peptide sequences (including modified variants or peptidoforms) reported across all analyses (original submission and reanalyses) associated with this dataset.

"N/A" means no results of this type were submitted.
Total number of peptide-spectrum matches (i.e. spectrum identifications) reported across all analyses (original submission and reanalyses) associated with this dataset.

"N/A" means no results of this type were submitted.
Number of distinct proteins quantified across all analyses (original submission and reanalyses) associated with this dataset.

Distinct protein accessions are counted across all files submitted in the "Statistical Analysis of Quantified Analytes" category having a "Protein" column in this dataset.

"N/A" means no results of this type were submitted.
Number of distinct proteins found to be differentially abundant in at least one comparison across all analyses (original submission and reanalyses) associated with this dataset.

A protein is differentially abundant if its change in abundance across conditions is found to be statistically significant with an adjusted p-value <= 0.05 and lists no issues associated with statistical tests for differential abundance.

Distinct protein accessions are counted across all files submitted in the "Statistical Analysis of Quantified Analytes" category having a "Protein" column in this dataset.

"N/A" means no results of this type were submitted.
This dataset may not contain all raw spectra data as originally deposited in PRIDE. It has been imported to MassIVE for reanalysis purposes, so its spectra data here may consist solely of processed peak lists suitable for reanalysis with most software.