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Limited Evidence for Protein Products of Noncoding Transcripts in the HEK293T Cellular Cytosol

Ribosome profiling has revealed translation outside canonical coding sequences, including translation of short upstream ORFs, long noncoding RNAs, overlapping ORFs, ORFs in UTRs, or ORFs in alternative reading frames. Studies combining mass spectrometry, ribosome profiling, and CRISPR-based screens...

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Detalles Bibliográficos
Autores principales: Bogaert, Annelies, Fijalkowska, Daria, Staes, An, Van de Steene, Tessa, Demol, Hans, Gevaert, Kris
Formato: Online Artículo Texto
Lenguaje:English
Publicado: American Society for Biochemistry and Molecular Biology 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9396073/
https://www.ncbi.nlm.nih.gov/pubmed/35788065
http://dx.doi.org/10.1016/j.mcpro.2022.100264
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author Bogaert, Annelies
Fijalkowska, Daria
Staes, An
Van de Steene, Tessa
Demol, Hans
Gevaert, Kris
author_facet Bogaert, Annelies
Fijalkowska, Daria
Staes, An
Van de Steene, Tessa
Demol, Hans
Gevaert, Kris
author_sort Bogaert, Annelies
collection PubMed
description Ribosome profiling has revealed translation outside canonical coding sequences, including translation of short upstream ORFs, long noncoding RNAs, overlapping ORFs, ORFs in UTRs, or ORFs in alternative reading frames. Studies combining mass spectrometry, ribosome profiling, and CRISPR-based screens showed that hundreds of ORFs derived from noncoding transcripts produce (micro)proteins, whereas other studies failed to find evidence for such types of noncanonical translation products. Here, we attempted to discover translation products from noncoding regions by strongly reducing the complexity of the sample prior to mass spectrometric analysis. We used an extended database as the search space and applied stringent filtering of the identified peptides to find evidence for novel translation events. We show that, theoretically our strategy facilitates the detection of translation events of transcripts from noncoding regions but experimentally only find 19 peptides that might originate from such translation events. Finally, Virotrap-based interactome analysis of two N-terminal proteoforms originating from noncoding regions showed the functional potential of these novel proteins.
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spelling pubmed-93960732022-08-25 Limited Evidence for Protein Products of Noncoding Transcripts in the HEK293T Cellular Cytosol Bogaert, Annelies Fijalkowska, Daria Staes, An Van de Steene, Tessa Demol, Hans Gevaert, Kris Mol Cell Proteomics Research Ribosome profiling has revealed translation outside canonical coding sequences, including translation of short upstream ORFs, long noncoding RNAs, overlapping ORFs, ORFs in UTRs, or ORFs in alternative reading frames. Studies combining mass spectrometry, ribosome profiling, and CRISPR-based screens showed that hundreds of ORFs derived from noncoding transcripts produce (micro)proteins, whereas other studies failed to find evidence for such types of noncanonical translation products. Here, we attempted to discover translation products from noncoding regions by strongly reducing the complexity of the sample prior to mass spectrometric analysis. We used an extended database as the search space and applied stringent filtering of the identified peptides to find evidence for novel translation events. We show that, theoretically our strategy facilitates the detection of translation events of transcripts from noncoding regions but experimentally only find 19 peptides that might originate from such translation events. Finally, Virotrap-based interactome analysis of two N-terminal proteoforms originating from noncoding regions showed the functional potential of these novel proteins. American Society for Biochemistry and Molecular Biology 2022-07-02 /pmc/articles/PMC9396073/ /pubmed/35788065 http://dx.doi.org/10.1016/j.mcpro.2022.100264 Text en © 2022 The Authors https://creativecommons.org/licenses/by/4.0/This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).
spellingShingle Research
Bogaert, Annelies
Fijalkowska, Daria
Staes, An
Van de Steene, Tessa
Demol, Hans
Gevaert, Kris
Limited Evidence for Protein Products of Noncoding Transcripts in the HEK293T Cellular Cytosol
title Limited Evidence for Protein Products of Noncoding Transcripts in the HEK293T Cellular Cytosol
title_full Limited Evidence for Protein Products of Noncoding Transcripts in the HEK293T Cellular Cytosol
title_fullStr Limited Evidence for Protein Products of Noncoding Transcripts in the HEK293T Cellular Cytosol
title_full_unstemmed Limited Evidence for Protein Products of Noncoding Transcripts in the HEK293T Cellular Cytosol
title_short Limited Evidence for Protein Products of Noncoding Transcripts in the HEK293T Cellular Cytosol
title_sort limited evidence for protein products of noncoding transcripts in the hek293t cellular cytosol
topic Research
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9396073/
https://www.ncbi.nlm.nih.gov/pubmed/35788065
http://dx.doi.org/10.1016/j.mcpro.2022.100264
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