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Efficient Detection of the Alternative Spliced Human Proteome Using Translatome Sequencing

Alternative splicing (AS) isoforms create numerous proteoforms, expanding the complexity of the genome. Highly similar sequences, incomplete reference databases and the insufficient sequence coverage of mass spectrometry limit the identification of AS proteoforms. Here, we demonstrated full-length t...

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Autores principales: Wu, Chun, Lu, Xiaolong, Lu, Shaohua, Wang, Hongwei, Li, Dehua, Zhao, Jing, Jin, Jingjie, Sun, Zhenghua, He, Qing-Yu, Chen, Yang, Zhang, Gong
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Frontiers Media S.A. 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9201276/
https://www.ncbi.nlm.nih.gov/pubmed/35720116
http://dx.doi.org/10.3389/fmolb.2022.895746
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author Wu, Chun
Lu, Xiaolong
Lu, Shaohua
Wang, Hongwei
Li, Dehua
Zhao, Jing
Jin, Jingjie
Sun, Zhenghua
He, Qing-Yu
Chen, Yang
Zhang, Gong
author_facet Wu, Chun
Lu, Xiaolong
Lu, Shaohua
Wang, Hongwei
Li, Dehua
Zhao, Jing
Jin, Jingjie
Sun, Zhenghua
He, Qing-Yu
Chen, Yang
Zhang, Gong
author_sort Wu, Chun
collection PubMed
description Alternative splicing (AS) isoforms create numerous proteoforms, expanding the complexity of the genome. Highly similar sequences, incomplete reference databases and the insufficient sequence coverage of mass spectrometry limit the identification of AS proteoforms. Here, we demonstrated full-length translating mRNAs (ribosome nascent-chain complex-bound mRNAs, RNC-mRNAs) sequencing (RNC-seq) strategy to sequence the entire translating mRNA using next-generation sequencing, including short-read and long-read technologies, to construct a protein database containing all translating AS isoforms. Taking the advantage of read length, short-read RNC-seq identified up to 15,289 genes and 15,906 AS isoforms in a single human cell line, much more than the Ribo-seq. The single-molecule long-read RNC-seq supplemented 4,429 annotated AS isoforms that were not identified by short-read datasets, and 4,525 novel AS isoforms that were not included in the public databases. Using such RNC-seq-guided database, we identified 6,766 annotated protein isoforms and 50 novel protein isoforms in mass spectrometry datasets. These results demonstrated the potential of full-length RNC-seq in investigating the proteome of AS isoforms.
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spelling pubmed-92012762022-06-17 Efficient Detection of the Alternative Spliced Human Proteome Using Translatome Sequencing Wu, Chun Lu, Xiaolong Lu, Shaohua Wang, Hongwei Li, Dehua Zhao, Jing Jin, Jingjie Sun, Zhenghua He, Qing-Yu Chen, Yang Zhang, Gong Front Mol Biosci Molecular Biosciences Alternative splicing (AS) isoforms create numerous proteoforms, expanding the complexity of the genome. Highly similar sequences, incomplete reference databases and the insufficient sequence coverage of mass spectrometry limit the identification of AS proteoforms. Here, we demonstrated full-length translating mRNAs (ribosome nascent-chain complex-bound mRNAs, RNC-mRNAs) sequencing (RNC-seq) strategy to sequence the entire translating mRNA using next-generation sequencing, including short-read and long-read technologies, to construct a protein database containing all translating AS isoforms. Taking the advantage of read length, short-read RNC-seq identified up to 15,289 genes and 15,906 AS isoforms in a single human cell line, much more than the Ribo-seq. The single-molecule long-read RNC-seq supplemented 4,429 annotated AS isoforms that were not identified by short-read datasets, and 4,525 novel AS isoforms that were not included in the public databases. Using such RNC-seq-guided database, we identified 6,766 annotated protein isoforms and 50 novel protein isoforms in mass spectrometry datasets. These results demonstrated the potential of full-length RNC-seq in investigating the proteome of AS isoforms. Frontiers Media S.A. 2022-06-02 /pmc/articles/PMC9201276/ /pubmed/35720116 http://dx.doi.org/10.3389/fmolb.2022.895746 Text en Copyright © 2022 Wu, Lu, Lu, Wang, Li, Zhao, Jin, Sun, He, Chen and Zhang. https://creativecommons.org/licenses/by/4.0/This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
spellingShingle Molecular Biosciences
Wu, Chun
Lu, Xiaolong
Lu, Shaohua
Wang, Hongwei
Li, Dehua
Zhao, Jing
Jin, Jingjie
Sun, Zhenghua
He, Qing-Yu
Chen, Yang
Zhang, Gong
Efficient Detection of the Alternative Spliced Human Proteome Using Translatome Sequencing
title Efficient Detection of the Alternative Spliced Human Proteome Using Translatome Sequencing
title_full Efficient Detection of the Alternative Spliced Human Proteome Using Translatome Sequencing
title_fullStr Efficient Detection of the Alternative Spliced Human Proteome Using Translatome Sequencing
title_full_unstemmed Efficient Detection of the Alternative Spliced Human Proteome Using Translatome Sequencing
title_short Efficient Detection of the Alternative Spliced Human Proteome Using Translatome Sequencing
title_sort efficient detection of the alternative spliced human proteome using translatome sequencing
topic Molecular Biosciences
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9201276/
https://www.ncbi.nlm.nih.gov/pubmed/35720116
http://dx.doi.org/10.3389/fmolb.2022.895746
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