Proteomics

Dataset Information

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Pervasive functional micropeptides in humans encoded by non-canonical open reading frames


ABSTRACT: Protein-coding regions of the genome continue to expand beyond canonical annotations and remain incompletely understood. Despite recent reports demonstrating important cellular functions played by micropeptides from unannotated open reading frames (ORFs), there exists no systematic method to fully uncover the prevalence of functional ORFs and their encoded peptides. Here, using ribosome profiling and mass spectrometry based proteomics, we reveal pervasive translation of ORFs previously annotated as non-coding, including those encoded on long non-coding RNAs (lncRNA ORFs) and upstream regions of protein-coding messages (uORFs). Cross-validation of novel ORFs in six different cell lines via HLA peptidomics confirm their allotype-specific MHC receptor presentation and their engagement with the endogenous HLA processing machinery. We further developed systematic CRISPR-based screens to show that hundreds of these ORFs are essential for cellular growth and encode stable, functional peptides that play diverse roles in cells. By tagging with a split-GFP system, we show that many of the peptides have specific cellular localization patterns and form functional protein complexes. We uncovered uORFs encoding stable peptides that interact with the downstream protein encoded on the same mRNA, suggesting possible regulatory or feedback roles from the bicistronic message in addition to the previously assumed role of uORFs in translational repression. Our results uncover functional micropeptides encoded in the human genome with essential roles in diverse biological processes.

OTHER RELATED OMICS DATASETS IN: GSE131650

INSTRUMENT(S): Q Exactive

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Cell Culture

SUBMITTER: Andreas-David Brunner  

LAB HEAD: Matthias Mann

PROVIDER: PXD014031 | Pride | 2020-03-06

REPOSITORIES: Pride

Dataset's files

Source:
Action DRS
FullProteome_sORF.rar Other
HLA.rar Other
Pulldowns.rar Other
txt_FullProteome_sORF.rar Other
txt_HLA_sORF.rar Other
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Publications


Ribosome profiling has revealed pervasive but largely uncharacterized translation outside of canonical coding sequences (CDSs). In this work, we exploit a systematic CRISPR-based screening strategy to identify hundreds of noncanonical CDSs that are essential for cellular growth and whose disruption elicits specific, robust transcriptomic and phenotypic changes in human cells. Functional characterization of the encoded microproteins reveals distinct cellular localizations, specific protein bindin  ...[more]

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