Time-Resolved Proteome Analysis of
            <i>Listeria monocytogenes</i>
            during Infection Reveals the Role of the AAA+ Chaperone ClpC for Host Cell Adaptation

Birk, Marlène S; Ahmed-Begrich, Rina; Tran, Stefan; Elsholz, Alexander K. W.; Frese, Christian K.; Charpentier, Emmanuelle

doi:10.1128/msystems.00215-21

Cited by 9 publications

(8 citation statements)

References 76 publications

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“…In L . monocytogenes and Bacillus subtilis , MurA is a substrate of the ClpCP protease [ 30 , 31 ] and reoM and reoY mutants accumulate MurA to a similar extent as a clpC mutant [ 24 , 27 ]. Analysis of phospho-ablative reoM strains and mutants depleted for PrkA and the corresponding protein phosphatase PrpC showed that genetic constellations, in which ReoM is locked in the phosphorylated state, cause MurA stabilization and vice versa [ 24 ].…”

Section: Introductionmentioning

confidence: 99%

MurA escape mutations uncouple peptidoglycan biosynthesis from PrkA signaling

et al. 2022

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Gram-positive bacteria are protected by a thick mesh of peptidoglycan (PG) completely engulfing their cells. This PG network is the main component of the bacterial cell wall, it provides rigidity and acts as foundation for the attachment of other surface molecules. Biosynthesis of PG consumes a high amount of cellular resources and therefore requires careful adjustments to environmental conditions. An important switch in the control of PG biosynthesis of Listeria monocytogenes, a Gram-positive pathogen with a high infection fatality rate, is the serine/threonine protein kinase PrkA. A key substrate of this kinase is the small cytosolic protein ReoM. We have shown previously that ReoM phosphorylation regulates PG formation through control of MurA stability. MurA catalyzes the first step in PG biosynthesis and the current model suggests that phosphorylated ReoM prevents MurA degradation by the ClpCP protease. In contrast, conditions leading to ReoM dephosphorylation stimulate MurA degradation. How ReoM controls degradation of MurA and potential other substrates is not understood. Also, the individual contribution of the ~20 other known PrkA targets to PG biosynthesis regulation is unknown. We here present murA mutants which escape proteolytic degradation. The release of MurA from ClpCP-dependent proteolysis was able to activate PG biosynthesis and further enhances the intrinsic cephalosporin resistance of L. monocytogenes. This latter effect required the RodA3/PBP B3 transglycosylase/transpeptidase pair. One murA escape mutation not only fully rescued an otherwise non-viable prkA mutant during growth in batch culture and inside macrophages but also overcompensated cephalosporin hypersensitivity. Our data collectively indicate that the main purpose of PrkA-mediated signaling in L. monocytogenes is control of MurA stability during standard laboratory growth conditions and intracellular growth in macrophages. These findings have important implications for the understanding of PG biosynthesis regulation and β-lactam resistance of L. monocytogenes and related Gram-positive bacteria.

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Section: Introductionmentioning

confidence: 99%

MurA escape mutations uncouple peptidoglycan biosynthesis from PrkA signaling

et al. 2022

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“…Protein XL-MS and computational modeling has also gained attention for unveiling protein interactions. Early this year Birk et al developed another new method combining metabolic and chemical isobaric peptide labeling to simultaneously determine time-resolved protein decay and de novo synthesis in Listeria monocytogenes ( 85 ). Employing this new method, the authors identified more than 100 ClpC target proteins and observed indirect effects of the clp C deletion on the protein abundance in diverse cellular and metabolic pathways, highlighting the crucial role of ClpC for L. monocytogenes adaptation to the host environment through proteome remodeling.…”

Section: Microorganisms and Their Proteomesmentioning

confidence: 99%

“…And the Liu group also profiled the intracellular proteome of a Salmonella mutant lacking a putative regulatory gene, ydcR , which was found to transcriptionally control the expression of a known virulence factor, SrfN ( 109 ). Other than Salmonella typhimurium, a number of groups also examined the proteome of other bacteria residing within host cells, such as Shigella flexneri ( 110 ), Listeria monocytogenes ( 85 ), Bordetella pertussis ( 111 ), Brucella abortus ( 112 ), and Staphylococcus aureus ( 113 ). Last but not least, there have also been attempts to study bacterial proteomes from in vivo infections ( 93 ), though thus far such practice can still be technically challenging mostly due to limited amounts of sample recovered from infected animal tissues.…”

Section: The Dynamic Host-pathogen Interactions During Infectionmentioning

confidence: 99%

Proteomic Approaches to Unravel Mechanisms of Antibiotic Resistance and Immune Evasion of Bacterial Pathogens

et al. 2022

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The profound effects of and distress caused by the global COVID-19 pandemic highlighted what has been known in the health sciences a long time ago: that bacteria, fungi, viruses, and parasites continue to present a major threat to human health. Infectious diseases remain the leading cause of death worldwide, with antibiotic resistance increasing exponentially due to a lack of new treatments. In addition to this, many pathogens share the common trait of having the ability to modulate, and escape from, the host immune response. The challenge in medical microbiology is to develop and apply new experimental approaches that allow for the identification of both the microbe and its drug susceptibility profile in a time-sensitive manner, as well as to elucidate their molecular mechanisms of survival and immunomodulation. Over the last three decades, proteomics has contributed to a better understanding of the underlying molecular mechanisms responsible for microbial drug resistance and pathogenicity. Proteomics has gained new momentum as a result of recent advances in mass spectrometry. Indeed, mass spectrometry-based biomedical research has been made possible thanks to technological advances in instrumentation capability and the continuous improvement of sample processing and workflows. For example, high-throughput applications such as SWATH or Trapped ion mobility enable the identification of thousands of proteins in a matter of minutes. This type of rapid, in-depth analysis, combined with other advanced, supportive applications such as data processing and artificial intelligence, presents a unique opportunity to translate knowledge-based findings into measurable impacts like new antimicrobial biomarkers and drug targets. In relation to the Research Topic “Proteomic Approaches to Unravel Mechanisms of Resistance and Immune Evasion of Bacterial Pathogens,” this review specifically seeks to highlight the synergies between the powerful fields of modern proteomics and microbiology, as well as bridging translational opportunities from biomedical research to clinical practice.

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“…The proteomes of many other intracellular bacteria, including S. flexneri , L. monocytogenes , B. pertussis , Brucella ( B .) abortus , R. prowazekii , etc., were also investigated for vaccine development by many groups [ 194 , 195 , 196 , 197 , 198 ]. This research opens the door to the development of a potent vaccine and offers a broad strategy for the design and development of vaccines against other intracellular diseases.…”

Section: Proteomics: a Genomic Complement For Vaccine Developmentmentioning

confidence: 99%

Recent Advances in Genomics-Based Approaches for the Development of Intracellular Bacterial Pathogen Vaccines

et al. 2022

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Infectious diseases continue to be a leading cause of morbidity and mortality worldwide. The majority of infectious diseases are caused by intracellular pathogenic bacteria (IPB). Historically, conventional vaccination drives have helped control the pathogenesis of intracellular bacteria and the emergence of antimicrobial resistance, saving millions of lives. However, in light of various limitations, many diseases that involve IPB still do not have adequate vaccines. In response to increasing demand for novel vaccine development strategies, a new area of vaccine research emerged following the advent of genomics technology, which changed the paradigm of vaccine development by utilizing the complete genomic data of microorganisms against them. It became possible to identify genes related to disease virulence, genetic patterns linked to disease virulence, as well as the genetic components that supported immunity and favorable vaccine responses. Complete genomic databases, and advancements in transcriptomics, metabolomics, structural genomics, proteomics, immunomics, pan-genomics, synthetic genomics, and population biology have allowed researchers to identify potential vaccine candidates and predict their effects in patients. New vaccines have been created against diseases for which previously there were no vaccines available, and existing vaccines have been improved. This review highlights the key issues and explores the evolution of vaccines. The increasing volume of IPB genomic data, and their application in novel genome-based techniques for vaccine development, were also examined, along with their characteristics, and the opportunities and obstacles involved. Critically, the application of genomics technology has helped researchers rapidly select and evaluate candidate antigens. Novel vaccines capable of addressing the limitations associated with conventional vaccines have been developed and pressing healthcare issues are being addressed.

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Time-Resolved Proteome Analysis of Listeria monocytogenes during Infection Reveals the Role of the AAA+ Chaperone ClpC for Host Cell Adaptation

Cited by 9 publications

References 76 publications

MurA escape mutations uncouple peptidoglycan biosynthesis from PrkA signaling

MurA escape mutations uncouple peptidoglycan biosynthesis from PrkA signaling

Proteomic Approaches to Unravel Mechanisms of Antibiotic Resistance and Immune Evasion of Bacterial Pathogens

Recent Advances in Genomics-Based Approaches for the Development of Intracellular Bacterial Pathogen Vaccines

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