ProTInSeq: transposon insertion tracking by ultra-deep DNA sequencing to identify translated large and small ORFs
Abstract Identifying open reading frames (ORFs) being translated is not a trivial task. ProTInSeq is a technique designed to characterize proteomes by sequencing transposon insertions engineered to express a selection marker when they occur in-frame within a protein-coding gene. In the bacterium Myc...
| Main Authors: | , , , , , |
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| Format: | Article |
| Language: | English |
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Nature Portfolio
2024-03-01
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| Series: | Nature Communications |
| Online Access: | https://doi.org/10.1038/s41467-024-46112-2 |
| _version_ | 1827301044659421184 |
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| author | Samuel Miravet-Verde Rocco Mazzolini Carolina Segura-Morales Alicia Broto Maria Lluch-Senar Luis Serrano |
| author_facet | Samuel Miravet-Verde Rocco Mazzolini Carolina Segura-Morales Alicia Broto Maria Lluch-Senar Luis Serrano |
| author_sort | Samuel Miravet-Verde |
| collection | DOAJ |
| description | Abstract Identifying open reading frames (ORFs) being translated is not a trivial task. ProTInSeq is a technique designed to characterize proteomes by sequencing transposon insertions engineered to express a selection marker when they occur in-frame within a protein-coding gene. In the bacterium Mycoplasma pneumoniae, ProTInSeq identifies 83% of its annotated proteins, along with 5 proteins and 153 small ORF-encoded proteins (SEPs; ≤100 aa) that were not previously annotated. Moreover, ProTInSeq can be utilized for detecting translational noise, as well as for relative quantification and transmembrane topology estimation of fitness and non-essential proteins. By integrating various identification approaches, the number of initially annotated SEPs in this bacterium increases from 27 to 329, with a quarter of them predicted to possess antimicrobial potential. Herein, we describe a methodology complementary to Ribo-Seq and mass spectroscopy that can identify SEPs while providing other insights in a proteome with a flexible and cost-effective DNA ultra-deep sequencing approach. |
| first_indexed | 2024-04-24T16:16:24Z |
| format | Article |
| id | doaj.art-4737d8ad11ee4bcea5a868ec42cd75f1 |
| institution | Directory Open Access Journal |
| issn | 2041-1723 |
| language | English |
| last_indexed | 2024-04-24T16:16:24Z |
| publishDate | 2024-03-01 |
| publisher | Nature Portfolio |
| record_format | Article |
| series | Nature Communications |
| spelling | doaj.art-4737d8ad11ee4bcea5a868ec42cd75f12024-03-31T11:24:35ZengNature PortfolioNature Communications2041-17232024-03-0115111710.1038/s41467-024-46112-2ProTInSeq: transposon insertion tracking by ultra-deep DNA sequencing to identify translated large and small ORFsSamuel Miravet-Verde0Rocco Mazzolini1Carolina Segura-Morales2Alicia Broto3Maria Lluch-Senar4Luis Serrano5Centre for Genomic Regulation (CRG), The Barcelona Institute of Science and Technology, Dr Aiguader 88Pulmobiotics, Dr Aiguader 88Centre for Genomic Regulation (CRG), The Barcelona Institute of Science and Technology, Dr Aiguader 88Centre for Genomic Regulation (CRG), The Barcelona Institute of Science and Technology, Dr Aiguader 88Pulmobiotics, Dr Aiguader 88Centre for Genomic Regulation (CRG), The Barcelona Institute of Science and Technology, Dr Aiguader 88Abstract Identifying open reading frames (ORFs) being translated is not a trivial task. ProTInSeq is a technique designed to characterize proteomes by sequencing transposon insertions engineered to express a selection marker when they occur in-frame within a protein-coding gene. In the bacterium Mycoplasma pneumoniae, ProTInSeq identifies 83% of its annotated proteins, along with 5 proteins and 153 small ORF-encoded proteins (SEPs; ≤100 aa) that were not previously annotated. Moreover, ProTInSeq can be utilized for detecting translational noise, as well as for relative quantification and transmembrane topology estimation of fitness and non-essential proteins. By integrating various identification approaches, the number of initially annotated SEPs in this bacterium increases from 27 to 329, with a quarter of them predicted to possess antimicrobial potential. Herein, we describe a methodology complementary to Ribo-Seq and mass spectroscopy that can identify SEPs while providing other insights in a proteome with a flexible and cost-effective DNA ultra-deep sequencing approach.https://doi.org/10.1038/s41467-024-46112-2 |
| spellingShingle | Samuel Miravet-Verde Rocco Mazzolini Carolina Segura-Morales Alicia Broto Maria Lluch-Senar Luis Serrano ProTInSeq: transposon insertion tracking by ultra-deep DNA sequencing to identify translated large and small ORFs Nature Communications |
| title | ProTInSeq: transposon insertion tracking by ultra-deep DNA sequencing to identify translated large and small ORFs |
| title_full | ProTInSeq: transposon insertion tracking by ultra-deep DNA sequencing to identify translated large and small ORFs |
| title_fullStr | ProTInSeq: transposon insertion tracking by ultra-deep DNA sequencing to identify translated large and small ORFs |
| title_full_unstemmed | ProTInSeq: transposon insertion tracking by ultra-deep DNA sequencing to identify translated large and small ORFs |
| title_short | ProTInSeq: transposon insertion tracking by ultra-deep DNA sequencing to identify translated large and small ORFs |
| title_sort | protinseq transposon insertion tracking by ultra deep dna sequencing to identify translated large and small orfs |
| url | https://doi.org/10.1038/s41467-024-46112-2 |
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