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Transposon mutagenesis in Mycobacterium abscessus identifies an essential penicillin-binding protein involved in septal peptidoglycan synthesis and antibiotic sensitivity

Mycobacterium abscessus (Mab) is a rapidly growing non-tuberculous mycobacterium (NTM) that causes a wide range of infections. Treatment of Mab infections is difficult because the bacterium is intrinsically resistant to many classes of antibiotics. Developing new and effective treatments against Mab...

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Autores principales: Akusobi, Chidiebere, Benghomari, Bouchra S, Zhu, Junhao, Wolf, Ian D, Singhvi, Shreya, Dulberger, Charles L, Ioerger, Thomas R, Rubin, Eric J
Formato: Online Artículo Texto
Lenguaje:English
Publicado: eLife Sciences Publications, Ltd 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9170245/
https://www.ncbi.nlm.nih.gov/pubmed/35659317
http://dx.doi.org/10.7554/eLife.71947
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author Akusobi, Chidiebere
Benghomari, Bouchra S
Zhu, Junhao
Wolf, Ian D
Singhvi, Shreya
Dulberger, Charles L
Ioerger, Thomas R
Rubin, Eric J
author_facet Akusobi, Chidiebere
Benghomari, Bouchra S
Zhu, Junhao
Wolf, Ian D
Singhvi, Shreya
Dulberger, Charles L
Ioerger, Thomas R
Rubin, Eric J
author_sort Akusobi, Chidiebere
collection PubMed
description Mycobacterium abscessus (Mab) is a rapidly growing non-tuberculous mycobacterium (NTM) that causes a wide range of infections. Treatment of Mab infections is difficult because the bacterium is intrinsically resistant to many classes of antibiotics. Developing new and effective treatments against Mab requires a better understanding of the unique vulnerabilities that can be targeted for future drug development. To achieve this, we identified essential genes in Mab by conducting transposon sequencing (TnSeq) on the reference Mab strain ATCC 19977. We generated ~51,000 unique transposon mutants and used this high-density library to identify 362 essential genes for in vitro growth. To investigate species-specific vulnerabilities in Mab, we further characterized MAB_3167c, a predicted penicillin-binding protein and hypothetical lipoprotein (PBP-lipo) that is essential in Mab and non-essential in Mycobacterium tuberculosis (Mtb). We found that PBP-lipo primarily localizes to the subpolar region and later to the septum as cells prepare to divide. Depletion of Mab PBP-lipo causes cells to elongate, develop ectopic branches, and form multiple septa. Knockdown of PBP-lipo along with PbpB, DacB1, and a carboxypeptidase, MAB_0519 lead to synergistic growth arrest. In contrast, these genetic interactions were absent in the Mtb model organism, Mycobacterium smegmatis, indicating that the PBP-lipo homologs in the two species exist in distinct genetic networks. Finally, repressing PBP-lipo sensitized the reference strain and 11 Mab clinical isolates to several classes of antibiotics, including the β-lactams, ampicillin, and amoxicillin by greater than 128-fold. Altogether, this study presents PBP-lipo as a key enzyme to study Mab-specific processes in cell wall synthesis and importantly positions PBP-lipo as an attractive drug target to treat Mab infections.
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spelling pubmed-91702452022-06-07 Transposon mutagenesis in Mycobacterium abscessus identifies an essential penicillin-binding protein involved in septal peptidoglycan synthesis and antibiotic sensitivity Akusobi, Chidiebere Benghomari, Bouchra S Zhu, Junhao Wolf, Ian D Singhvi, Shreya Dulberger, Charles L Ioerger, Thomas R Rubin, Eric J eLife Microbiology and Infectious Disease Mycobacterium abscessus (Mab) is a rapidly growing non-tuberculous mycobacterium (NTM) that causes a wide range of infections. Treatment of Mab infections is difficult because the bacterium is intrinsically resistant to many classes of antibiotics. Developing new and effective treatments against Mab requires a better understanding of the unique vulnerabilities that can be targeted for future drug development. To achieve this, we identified essential genes in Mab by conducting transposon sequencing (TnSeq) on the reference Mab strain ATCC 19977. We generated ~51,000 unique transposon mutants and used this high-density library to identify 362 essential genes for in vitro growth. To investigate species-specific vulnerabilities in Mab, we further characterized MAB_3167c, a predicted penicillin-binding protein and hypothetical lipoprotein (PBP-lipo) that is essential in Mab and non-essential in Mycobacterium tuberculosis (Mtb). We found that PBP-lipo primarily localizes to the subpolar region and later to the septum as cells prepare to divide. Depletion of Mab PBP-lipo causes cells to elongate, develop ectopic branches, and form multiple septa. Knockdown of PBP-lipo along with PbpB, DacB1, and a carboxypeptidase, MAB_0519 lead to synergistic growth arrest. In contrast, these genetic interactions were absent in the Mtb model organism, Mycobacterium smegmatis, indicating that the PBP-lipo homologs in the two species exist in distinct genetic networks. Finally, repressing PBP-lipo sensitized the reference strain and 11 Mab clinical isolates to several classes of antibiotics, including the β-lactams, ampicillin, and amoxicillin by greater than 128-fold. Altogether, this study presents PBP-lipo as a key enzyme to study Mab-specific processes in cell wall synthesis and importantly positions PBP-lipo as an attractive drug target to treat Mab infections. eLife Sciences Publications, Ltd 2022-06-06 /pmc/articles/PMC9170245/ /pubmed/35659317 http://dx.doi.org/10.7554/eLife.71947 Text en © 2022, Akusobi et al https://creativecommons.org/licenses/by/4.0/This article is distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use and redistribution provided that the original author and source are credited.
spellingShingle Microbiology and Infectious Disease
Akusobi, Chidiebere
Benghomari, Bouchra S
Zhu, Junhao
Wolf, Ian D
Singhvi, Shreya
Dulberger, Charles L
Ioerger, Thomas R
Rubin, Eric J
Transposon mutagenesis in Mycobacterium abscessus identifies an essential penicillin-binding protein involved in septal peptidoglycan synthesis and antibiotic sensitivity
title Transposon mutagenesis in Mycobacterium abscessus identifies an essential penicillin-binding protein involved in septal peptidoglycan synthesis and antibiotic sensitivity
title_full Transposon mutagenesis in Mycobacterium abscessus identifies an essential penicillin-binding protein involved in septal peptidoglycan synthesis and antibiotic sensitivity
title_fullStr Transposon mutagenesis in Mycobacterium abscessus identifies an essential penicillin-binding protein involved in septal peptidoglycan synthesis and antibiotic sensitivity
title_full_unstemmed Transposon mutagenesis in Mycobacterium abscessus identifies an essential penicillin-binding protein involved in septal peptidoglycan synthesis and antibiotic sensitivity
title_short Transposon mutagenesis in Mycobacterium abscessus identifies an essential penicillin-binding protein involved in septal peptidoglycan synthesis and antibiotic sensitivity
title_sort transposon mutagenesis in mycobacterium abscessus identifies an essential penicillin-binding protein involved in septal peptidoglycan synthesis and antibiotic sensitivity
topic Microbiology and Infectious Disease
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9170245/
https://www.ncbi.nlm.nih.gov/pubmed/35659317
http://dx.doi.org/10.7554/eLife.71947
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