Combination of Proteogenomics with Peptide <italic toggle="yes">De Novo</italic> Sequencing Identifies New Genes and Hidden Posttranscriptional Modifications

ABSTRACT Proteogenomics combines proteomics, genomics, and transcriptomics and has considerably improved genome annotation in poorly investigated phylogenetic groups for which homology information is lacking. Furthermore, it can be advantageous when reinvestigating well-annotated genomes. Here, we a...

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Autores principales:	B. Blank-Landeshammer, I. Teichert, R. Märker, M. Nowrousian, U. Kück, A. Sickmann
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2019
Materias:	proteogenomics peptide de novo sequencing RNA editing alternative splicing phylostratigraphy gene ontology Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/6b6994fd6554481e83bddc1e3025fd81
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spelling	oai:doaj.org-article:6b6994fd6554481e83bddc1e3025fd812021-11-15T15:59:42ZCombination of Proteogenomics with Peptide <italic toggle="yes">De Novo</italic> Sequencing Identifies New Genes and Hidden Posttranscriptional Modifications10.1128/mBio.02367-192150-7511https://doaj.org/article/6b6994fd6554481e83bddc1e3025fd812019-10-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mBio.02367-19https://doaj.org/toc/2150-7511ABSTRACT Proteogenomics combines proteomics, genomics, and transcriptomics and has considerably improved genome annotation in poorly investigated phylogenetic groups for which homology information is lacking. Furthermore, it can be advantageous when reinvestigating well-annotated genomes. Here, we applied an advanced proteogenomics approach, combining standard proteogenomics with peptide de novo sequencing, to refine annotation of the well-studied model fungus Sordaria macrospora. We investigated samples from different developmental and physiological conditions, resulting in the detection of 104 so-far hidden proteins and annotation changes in 575 genes, including 389 splice site refinements. Significantly, our approach provides peptide-level evidence for 113 single-amino-acid variations and 15 C-terminal protein elongations originating from A-to-I RNA editing, a phenomenon recently detected in fungi. Coexpression and phylostratigraphic analysis of the refined proteome suggest that new functions in evolutionarily young genes correlate with distinct developmental stages. In conclusion, our advanced proteogenomics approach supports and promotes functional studies of fungal model systems. IMPORTANCE Next-generation sequencing techniques have considerably increased the number of completely sequenced eukaryotic genomes. These genomes are mostly automatically annotated, and ab initio gene prediction is commonly combined with homology-based search approaches and often supported by transcriptomic data. The latter in particular improve the prediction of intron splice sites and untranslated regions. However, correct prediction of translation initiation sites (TIS), alternative splice junctions, and protein-coding potential remains challenging. Here, we present an advanced proteogenomics approach, namely, the combination of proteogenomics and de novo peptide sequencing analysis, in conjunction with Blast2GO and phylostratigraphy. Using the model fungus Sordaria macrospora as an example, we provide a comprehensive view of the proteome that not only increases the functional understanding of this multicellular organism at different developmental stages but also immensely enhances the genome annotation quality.B. Blank-LandeshammerI. TeichertR. MärkerM. NowrousianU. KückA. SickmannAmerican Society for Microbiologyarticleproteogenomicspeptide de novo sequencingRNA editingalternative splicingphylostratigraphygene ontologyMicrobiologyQR1-502ENmBio, Vol 10, Iss 5 (2019)
institution	DOAJ
collection	DOAJ
language	EN
topic	proteogenomics peptide de novo sequencing RNA editing alternative splicing phylostratigraphy gene ontology Microbiology QR1-502
spellingShingle	proteogenomics peptide de novo sequencing RNA editing alternative splicing phylostratigraphy gene ontology Microbiology QR1-502 B. Blank-Landeshammer I. Teichert R. Märker M. Nowrousian U. Kück A. Sickmann Combination of Proteogenomics with Peptide <italic toggle="yes">De Novo</italic> Sequencing Identifies New Genes and Hidden Posttranscriptional Modifications
description	ABSTRACT Proteogenomics combines proteomics, genomics, and transcriptomics and has considerably improved genome annotation in poorly investigated phylogenetic groups for which homology information is lacking. Furthermore, it can be advantageous when reinvestigating well-annotated genomes. Here, we applied an advanced proteogenomics approach, combining standard proteogenomics with peptide de novo sequencing, to refine annotation of the well-studied model fungus Sordaria macrospora. We investigated samples from different developmental and physiological conditions, resulting in the detection of 104 so-far hidden proteins and annotation changes in 575 genes, including 389 splice site refinements. Significantly, our approach provides peptide-level evidence for 113 single-amino-acid variations and 15 C-terminal protein elongations originating from A-to-I RNA editing, a phenomenon recently detected in fungi. Coexpression and phylostratigraphic analysis of the refined proteome suggest that new functions in evolutionarily young genes correlate with distinct developmental stages. In conclusion, our advanced proteogenomics approach supports and promotes functional studies of fungal model systems. IMPORTANCE Next-generation sequencing techniques have considerably increased the number of completely sequenced eukaryotic genomes. These genomes are mostly automatically annotated, and ab initio gene prediction is commonly combined with homology-based search approaches and often supported by transcriptomic data. The latter in particular improve the prediction of intron splice sites and untranslated regions. However, correct prediction of translation initiation sites (TIS), alternative splice junctions, and protein-coding potential remains challenging. Here, we present an advanced proteogenomics approach, namely, the combination of proteogenomics and de novo peptide sequencing analysis, in conjunction with Blast2GO and phylostratigraphy. Using the model fungus Sordaria macrospora as an example, we provide a comprehensive view of the proteome that not only increases the functional understanding of this multicellular organism at different developmental stages but also immensely enhances the genome annotation quality.
format	article
author	B. Blank-Landeshammer I. Teichert R. Märker M. Nowrousian U. Kück A. Sickmann
author_facet	B. Blank-Landeshammer I. Teichert R. Märker M. Nowrousian U. Kück A. Sickmann
author_sort	B. Blank-Landeshammer
title	Combination of Proteogenomics with Peptide <italic toggle="yes">De Novo</italic> Sequencing Identifies New Genes and Hidden Posttranscriptional Modifications
title_short	Combination of Proteogenomics with Peptide <italic toggle="yes">De Novo</italic> Sequencing Identifies New Genes and Hidden Posttranscriptional Modifications
title_full	Combination of Proteogenomics with Peptide <italic toggle="yes">De Novo</italic> Sequencing Identifies New Genes and Hidden Posttranscriptional Modifications
title_fullStr	Combination of Proteogenomics with Peptide <italic toggle="yes">De Novo</italic> Sequencing Identifies New Genes and Hidden Posttranscriptional Modifications
title_full_unstemmed	Combination of Proteogenomics with Peptide <italic toggle="yes">De Novo</italic> Sequencing Identifies New Genes and Hidden Posttranscriptional Modifications
title_sort	combination of proteogenomics with peptide <italic toggle="yes">de novo</italic> sequencing identifies new genes and hidden posttranscriptional modifications
publisher	American Society for Microbiology
publishDate	2019
url	https://doaj.org/article/6b6994fd6554481e83bddc1e3025fd81
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Combination of Proteogenomics with Peptide <italic toggle="yes">De Novo</italic> Sequencing Identifies New Genes and Hidden Posttranscriptional Modifications

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