Composition and liquid-to-solid maturation of protein aggregates contribute to bacterial dormancy development and recovery.

Recalcitrant bacterial infections can be caused by various types of dormant bacteria, including persisters and viable but nonculturable (VBNC) cells. Despite their clinical importance, we know fairly little about bacterial dormancy development and recovery. Previously, we established a correlation between protein aggregation and dormancy in Escherichia coli.

The role of the essential GTPase ObgE in regulating lipopolysaccharide synthesis in Escherichia coli.

During growth, cells need to synthesize and expand their envelope, a process that requires careful regulation. Here, we show that the GTPase ObgE of E. coli contributes to the regulation of lipopolysaccharide (LPS) synthesis, an essential component of the Gram-negative outer membrane.

The rise and future of CRISPR-based approaches for high-throughput genomics.

Clustered regularly interspaced short palindromic repeats (CRISPR) has revolutionized the field of genome editing. To circumvent the permanent modifications made by traditional CRISPR techniques and facilitate the study of both essential and nonessential genes, CRISPR interference (CRISPRi) was developed. This gene-silencing technique employs a deactivated Cas effector protein and a guide RNA to block transcription initiation or elongation.

YbiB: a novel interactor of the GTPase ObgE.

Obg is a widely conserved and essential GTPase in bacteria, which plays a central role in a large range of important cellular processes, such as ribosome biogenesis, DNA replication, cell division and bacterial persistence. Nevertheless, the exact function of Obg in these processes and the interactions it makes within the associated pathways remain largely unknown. Here, we identify the DNA-binding TrpD2 protein YbiB as an interactor of the Escherichia coli Obg (ObgE).

Deep mutational scanning of essential bacterial proteins can guide antibiotic development.

Deep mutational scanning is a powerful approach to investigate a wide variety of research questions including protein function and stability. Here, we perform deep mutational scanning on three essential E. coli proteins (FabZ, LpxC and MurA) involved in cell envelope synthesis using high-throughput CRISPR genome editing, and study the effect of the mutations in their original genomic context.

Amoxicillin-resistant can be resensitized by targeting the mevalonate pathway as indicated by sCRilecs-seq.

Antibiotic resistance in the important opportunistic human pathogen is on the rise. This is particularly problematic in the case of the β-lactam antibiotic amoxicillin, which is the first-line therapy. It is therefore crucial to uncover targets that would kill or resensitize amoxicillin-resistant pneumococci.

Mutations in respiratory complex I promote antibiotic persistence through alterations in intracellular acidity and protein synthesis.

Antibiotic persistence describes the presence of phenotypic variants within an isogenic bacterial population that are transiently tolerant to antibiotic treatment. Perturbations of metabolic homeostasis can promote antibiotic persistence, but the precise mechanisms are not well understood. Here, we use laboratory evolution, population-wide sequencing and biochemical characterizations to identify mutations in respiratory complex I and discover how they promote persistence in Escherichia coli.

The Dynamic Transition of Persistence toward the Viable but Nonculturable State during Stationary Phase Is Driven by Protein Aggregation.

Decades of research into bacterial persistence has been unable to fully characterize this antibiotic-tolerant phenotype, thereby hampering the development of therapies effective against chronic infections. Although some active persister mechanisms have been identified, the prevailing view is that cells become persistent because they enter a dormant state. We therefore characterized starvation-induced dormancy in Escherichia coli.

Protein Aggregation as a Bacterial Strategy to Survive Antibiotic Treatment.

While protein aggregation is predominantly associated with loss of function and toxicity, it is also known to increase survival of bacteria under stressful conditions. Indeed, protein aggregation not only helps bacteria to cope with proteotoxic stresses like heat shocks or oxidative stress, but a growing number of studies suggest that it also improves survival during antibiotic treatment by inducing dormancy. A well-known example of dormant cells are persisters, which are transiently refractory to the action of antibiotics.

GTP Binding is Necessary for the Activation of a Toxic Mutant Isoform of the Essential GTPase ObgE.

Even though the Obg protein is essential for bacterial viability, the cellular functions of this universally conserved GTPase remain enigmatic. Moreover, the influence of GTP and GDP binding on the activity of this protein is largely unknown. Previously, we identified a mutant isoform of ObgE (the Obg protein of ) that triggers cell death.

Bacterial Heterogeneity and Antibiotic Survival: Understanding and Combatting Persistence and Heteroresistance.

For decades, mankind has dominated the battle against bacteria, yet the tide is slowly turning. Our antibacterial strategies are becoming less effective, allowing bacteria to get the upper hand. The alarming rise in antibiotic resistance is an important cause of anti-infective therapy failure.

Biochemical determinants of ObgE-mediated persistence.

Obg is a versatile GTPase that plays a pivotal role in bacterial persistence. We previously showed that the Escherichia coli homolog ObgE exerts this activity through transcriptional activation of a toxin-antitoxin module and subsequent membrane depolarization. Here, we assessed the role of G-domain functionality in ObgE-mediated persistence.

HokB Monomerization and Membrane Repolarization Control Persister Awakening.

Every bacterial population harbors a small subpopulation of so-called persisters that are transiently antibiotic tolerant. These persisters are associated with the recalcitrance of chronic infections because they can recolonize the host after antibiotic removal. Although several effectors have been described to induce persistence, persister cell awakening is poorly understood.

The Persistence-Inducing Toxin HokB Forms Dynamic Pores That Cause ATP Leakage.

Bacterial populations harbor a small fraction of cells that display transient multidrug tolerance. These so-called persister cells are extremely difficult to eradicate and contribute to the recalcitrance of chronic infections. Several signaling pathways leading to persistence have been identified.

An integrative view of cell cycle control in Escherichia coli.

Bacterial proliferation depends on the cells' capability to proceed through consecutive rounds of the cell cycle. The cell cycle consists of a series of events during which cells grow, copy their genome, partition the duplicated DNA into different cell halves and, ultimately, divide to produce two newly formed daughter cells. Cell cycle control is of the utmost importance to maintain the correct order of events and safeguard the integrity of the cell and its genomic information.

A Mutant Isoform of ObgE Causes Cell Death by Interfering with Cell Division.

Cell division is a vital part of the cell cycle that is fundamental to all life. Despite decades of intense investigation, this process is still incompletely understood. Previously, the essential GTPase ObgE, which plays a role in a myriad of basic cellular processes (such as initiation of DNA replication, chromosome segregation, and ribosome assembly), was proposed to act as a cell cycle checkpoint in by licensing chromosome segregation.

The bacterial cell cycle checkpoint protein Obg and its role in programmed cell death.

The phenomenon of programmed cell death (PCD), in which cells initiate their own demise, is not restricted to multicellular organisms. Unicellular organisms, both eukaryotes and prokaryotes, also possess pathways that mediate PCD. We recently identified a PCD mechanism in that is triggered by a mutant isoform of the essential GTPase ObgE (Obg of ).

Reactive oxygen species do not contribute to ObgE*-mediated programmed cell death.

Programmed cell death (PCD) in bacteria is considered an important target for developing novel antimicrobials. Development of PCD-specific therapies requires a deeper understanding of what drives this process. We recently discovered a new mode of PCD in Escherichia coli that is triggered by expression of a mutant isoform of the essential ObgE protein, ObgE*.

A Single-Amino-Acid Substitution in Obg Activates a New Programmed Cell Death Pathway in Escherichia coli.

Unlabelled: Programmed cell death (PCD) is an important hallmark of multicellular organisms. Cells self-destruct through a regulated series of events for the benefit of the organism as a whole. The existence of PCD in bacteria has long been controversial due to the widely held belief that only multicellular organisms would profit from this kind of altruistic behavior at the cellular level.

Obg and Membrane Depolarization Are Part of a Microbial Bet-Hedging Strategy that Leads to Antibiotic Tolerance.

Within bacterial populations, a small fraction of persister cells is transiently capable of surviving exposure to lethal doses of antibiotics. As a bet-hedging strategy, persistence levels are determined both by stochastic induction and by environmental stimuli called responsive diversification. Little is known about the mechanisms that link the low frequency of persisters to environmental signals.