Competitive fitness of essential gene knockdowns reveals a broad-spectrum antibacterial inhibitor of the cell division protein FtsZ

Andrew M. Hogan, Viola C. Scoffone, Vadim Makarov, April S. Gislason, Haben Tesfu, Maria S. Stietz, Ann Karen C. Brassinga, Michael Domaratzki, Xuan Li, Alberto Azzalin, Marco Biggiogera, Olga Riabova, Natalia Monakhova, Laurent R. Chiarelli, Giovanna Riccardi, Silvia Buroni, Silvia T. Cardona

Research output: Contribution to journalArticle

4 Citations (Scopus)

Abstract

To streamline the elucidation of antibacterial compounds’ mechanism of action, comprehensive high-throughput assays interrogating multiple putative targets are necessary. However, current chemogenomic approaches for antibiotic target identification have not fully utilized the multiplexing potential of next-generation sequencing. Here, we used Illumina sequencing of transposon insertions to track the competitive fitness of a Burkholderia cenocepacia library containing essential gene knockdowns. Using this method, we characterized a novel benzothiadiazole derivative, 10126109 (C109), with antibacterial activity against B. cenocepacia, for which whole-genome sequencing of low-frequency spontaneous drug-resistant mutants had failed to identify the drug target. By combining the identification of hypersusceptible mutants and morphology screening, we show that C109 targets cell division. Furthermore, fluorescence microscopy of bacteria harboring green fluorescent protein (GFP) cell division protein fusions revealed that C109 prevents divisome formation by altering the localization of the essential cell division protein FtsZ. In agreement with this, C109 inhibited both the GTPase and polymerization activities of purified B. cenocepacia FtsZ. C109 displayed antibacterial activity against Gram-positive and Gram-negative cystic fibrosis pathogens, including Mycobacterium abscessus. C109 effectively cleared B. cenocepacia infection in the Caenorhabditis elegans model and exhibited additive interactions with clinically relevant antibiotics. Hence, C109 is an enticing candidate for further drug development.

Original languageEnglish (US)
Article numbere01231-18
JournalAntimicrobial agents and chemotherapy
Volume62
Issue number12
DOIs
StatePublished - Dec 1 2018

Fingerprint

Burkholderia cenocepacia
Gene Knockdown Techniques
Essential Genes
Cell Division
Burkholderia Infections
Proteins
Pharmaceutical Preparations
Anti-Bacterial Agents
GTP Phosphohydrolases
Caenorhabditis elegans
Mycobacterium
Green Fluorescent Proteins
Fluorescence Microscopy
Cystic Fibrosis
Polymerization
Libraries
Genome
Bacteria

Keywords

  • Burkholderia
  • Drug targets
  • Essential genes
  • Fluorescent image analysis
  • FtsZ
  • Mechanisms of action
  • Tn-seq

PubMed: MeSH publication types

  • Journal Article
  • Research Support, Non-U.S. Gov't

Cite this

Hogan, A. M., Scoffone, V. C., Makarov, V., Gislason, A. S., Tesfu, H., Stietz, M. S., ... Cardona, S. T. (2018). Competitive fitness of essential gene knockdowns reveals a broad-spectrum antibacterial inhibitor of the cell division protein FtsZ. Antimicrobial agents and chemotherapy, 62(12), [e01231-18]. https://doi.org/10.1128/AAC.01231-18

Competitive fitness of essential gene knockdowns reveals a broad-spectrum antibacterial inhibitor of the cell division protein FtsZ. / Hogan, Andrew M.; Scoffone, Viola C.; Makarov, Vadim; Gislason, April S.; Tesfu, Haben; Stietz, Maria S.; Brassinga, Ann Karen C.; Domaratzki, Michael; Li, Xuan; Azzalin, Alberto; Biggiogera, Marco; Riabova, Olga; Monakhova, Natalia; Chiarelli, Laurent R.; Riccardi, Giovanna; Buroni, Silvia; Cardona, Silvia T.

In: Antimicrobial agents and chemotherapy, Vol. 62, No. 12, e01231-18, 01.12.2018.

Research output: Contribution to journalArticle

Hogan, AM, Scoffone, VC, Makarov, V, Gislason, AS, Tesfu, H, Stietz, MS, Brassinga, AKC, Domaratzki, M, Li, X, Azzalin, A, Biggiogera, M, Riabova, O, Monakhova, N, Chiarelli, LR, Riccardi, G, Buroni, S & Cardona, ST 2018, 'Competitive fitness of essential gene knockdowns reveals a broad-spectrum antibacterial inhibitor of the cell division protein FtsZ', Antimicrobial agents and chemotherapy, vol. 62, no. 12, e01231-18. https://doi.org/10.1128/AAC.01231-18
Hogan, Andrew M. ; Scoffone, Viola C. ; Makarov, Vadim ; Gislason, April S. ; Tesfu, Haben ; Stietz, Maria S. ; Brassinga, Ann Karen C. ; Domaratzki, Michael ; Li, Xuan ; Azzalin, Alberto ; Biggiogera, Marco ; Riabova, Olga ; Monakhova, Natalia ; Chiarelli, Laurent R. ; Riccardi, Giovanna ; Buroni, Silvia ; Cardona, Silvia T. / Competitive fitness of essential gene knockdowns reveals a broad-spectrum antibacterial inhibitor of the cell division protein FtsZ. In: Antimicrobial agents and chemotherapy. 2018 ; Vol. 62, No. 12.
@article{06a87d694be34fc9b924cb47bc6f3f53,
title = "Competitive fitness of essential gene knockdowns reveals a broad-spectrum antibacterial inhibitor of the cell division protein FtsZ",
abstract = "To streamline the elucidation of antibacterial compounds’ mechanism of action, comprehensive high-throughput assays interrogating multiple putative targets are necessary. However, current chemogenomic approaches for antibiotic target identification have not fully utilized the multiplexing potential of next-generation sequencing. Here, we used Illumina sequencing of transposon insertions to track the competitive fitness of a Burkholderia cenocepacia library containing essential gene knockdowns. Using this method, we characterized a novel benzothiadiazole derivative, 10126109 (C109), with antibacterial activity against B. cenocepacia, for which whole-genome sequencing of low-frequency spontaneous drug-resistant mutants had failed to identify the drug target. By combining the identification of hypersusceptible mutants and morphology screening, we show that C109 targets cell division. Furthermore, fluorescence microscopy of bacteria harboring green fluorescent protein (GFP) cell division protein fusions revealed that C109 prevents divisome formation by altering the localization of the essential cell division protein FtsZ. In agreement with this, C109 inhibited both the GTPase and polymerization activities of purified B. cenocepacia FtsZ. C109 displayed antibacterial activity against Gram-positive and Gram-negative cystic fibrosis pathogens, including Mycobacterium abscessus. C109 effectively cleared B. cenocepacia infection in the Caenorhabditis elegans model and exhibited additive interactions with clinically relevant antibiotics. Hence, C109 is an enticing candidate for further drug development.",
keywords = "Burkholderia, Drug targets, Essential genes, Fluorescent image analysis, FtsZ, Mechanisms of action, Tn-seq",
author = "Hogan, {Andrew M.} and Scoffone, {Viola C.} and Vadim Makarov and Gislason, {April S.} and Haben Tesfu and Stietz, {Maria S.} and Brassinga, {Ann Karen C.} and Michael Domaratzki and Xuan Li and Alberto Azzalin and Marco Biggiogera and Olga Riabova and Natalia Monakhova and Chiarelli, {Laurent R.} and Giovanna Riccardi and Silvia Buroni and Cardona, {Silvia T.}",
year = "2018",
month = "12",
day = "1",
doi = "10.1128/AAC.01231-18",
language = "English (US)",
volume = "62",
journal = "Antimicrobial Agents and Chemotherapy",
issn = "0066-4804",
publisher = "American Society for Microbiology",
number = "12",

}

TY - JOUR

T1 - Competitive fitness of essential gene knockdowns reveals a broad-spectrum antibacterial inhibitor of the cell division protein FtsZ

AU - Hogan, Andrew M.

AU - Scoffone, Viola C.

AU - Makarov, Vadim

AU - Gislason, April S.

AU - Tesfu, Haben

AU - Stietz, Maria S.

AU - Brassinga, Ann Karen C.

AU - Domaratzki, Michael

AU - Li, Xuan

AU - Azzalin, Alberto

AU - Biggiogera, Marco

AU - Riabova, Olga

AU - Monakhova, Natalia

AU - Chiarelli, Laurent R.

AU - Riccardi, Giovanna

AU - Buroni, Silvia

AU - Cardona, Silvia T.

PY - 2018/12/1

Y1 - 2018/12/1

N2 - To streamline the elucidation of antibacterial compounds’ mechanism of action, comprehensive high-throughput assays interrogating multiple putative targets are necessary. However, current chemogenomic approaches for antibiotic target identification have not fully utilized the multiplexing potential of next-generation sequencing. Here, we used Illumina sequencing of transposon insertions to track the competitive fitness of a Burkholderia cenocepacia library containing essential gene knockdowns. Using this method, we characterized a novel benzothiadiazole derivative, 10126109 (C109), with antibacterial activity against B. cenocepacia, for which whole-genome sequencing of low-frequency spontaneous drug-resistant mutants had failed to identify the drug target. By combining the identification of hypersusceptible mutants and morphology screening, we show that C109 targets cell division. Furthermore, fluorescence microscopy of bacteria harboring green fluorescent protein (GFP) cell division protein fusions revealed that C109 prevents divisome formation by altering the localization of the essential cell division protein FtsZ. In agreement with this, C109 inhibited both the GTPase and polymerization activities of purified B. cenocepacia FtsZ. C109 displayed antibacterial activity against Gram-positive and Gram-negative cystic fibrosis pathogens, including Mycobacterium abscessus. C109 effectively cleared B. cenocepacia infection in the Caenorhabditis elegans model and exhibited additive interactions with clinically relevant antibiotics. Hence, C109 is an enticing candidate for further drug development.

AB - To streamline the elucidation of antibacterial compounds’ mechanism of action, comprehensive high-throughput assays interrogating multiple putative targets are necessary. However, current chemogenomic approaches for antibiotic target identification have not fully utilized the multiplexing potential of next-generation sequencing. Here, we used Illumina sequencing of transposon insertions to track the competitive fitness of a Burkholderia cenocepacia library containing essential gene knockdowns. Using this method, we characterized a novel benzothiadiazole derivative, 10126109 (C109), with antibacterial activity against B. cenocepacia, for which whole-genome sequencing of low-frequency spontaneous drug-resistant mutants had failed to identify the drug target. By combining the identification of hypersusceptible mutants and morphology screening, we show that C109 targets cell division. Furthermore, fluorescence microscopy of bacteria harboring green fluorescent protein (GFP) cell division protein fusions revealed that C109 prevents divisome formation by altering the localization of the essential cell division protein FtsZ. In agreement with this, C109 inhibited both the GTPase and polymerization activities of purified B. cenocepacia FtsZ. C109 displayed antibacterial activity against Gram-positive and Gram-negative cystic fibrosis pathogens, including Mycobacterium abscessus. C109 effectively cleared B. cenocepacia infection in the Caenorhabditis elegans model and exhibited additive interactions with clinically relevant antibiotics. Hence, C109 is an enticing candidate for further drug development.

KW - Burkholderia

KW - Drug targets

KW - Essential genes

KW - Fluorescent image analysis

KW - FtsZ

KW - Mechanisms of action

KW - Tn-seq

UR - http://www.scopus.com/inward/record.url?scp=85057189615&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=85057189615&partnerID=8YFLogxK

U2 - 10.1128/AAC.01231-18

DO - 10.1128/AAC.01231-18

M3 - Article

C2 - 30297366

AN - SCOPUS:85057189615

VL - 62

JO - Antimicrobial Agents and Chemotherapy

JF - Antimicrobial Agents and Chemotherapy

SN - 0066-4804

IS - 12

M1 - e01231-18

ER -