scholarly journals ppGpp Ribosome Dimerization Model for Bacterial Persister Formation and Resuscitation

2019 ◽  
Author(s):  
Sooyeon Song ◽  
Thomas K. Wood
Keyword(s):  
Cystic Fibrosis ◽  
Lyme Disease ◽  
Single Cell ◽  
Direct Connection ◽  
Persister Cells ◽  
Dormant State ◽  
Modulation Factor ◽  
Ribosome Modulation Factor

ABSTRACTStress is ubiquitous for bacteria and converts a subpopulation of cells into a dormant state known as persistence, in which cells are tolerant to antimicrobials. These cells revive rapidly when the stress is removed and are likely the cause of many recurring infections such as those associated with tuberculosis, cystic fibrosis, and Lyme disease. However, how persister cells are formed is not understood well. Here we propose the ppGpp ribosome dimerization persister (PRDP) model in which the alarmone guanosine pentaphosphate/tetraphosphate (henceforth ppGpp) generates persister cells directly by inactivating ribosomes via the ribosome modulation factor (RMF) and the hibernation promoting factor (Hpf). We demonstrate both RMF and HpF increase persistence and reduce single-cell persister resuscitation and that ppGpp has no effect on single-cell persister resuscitation. Hence, a direct connection between ppGpp and persistence is shown.

scholarly journals An Antipersister Strategy for Treatment of Chronic Pseudomonas aeruginosa Infections

2017 ◽  
Vol 61 (12) ◽  
Author(s):  
Martina Koeva ◽  
Alina D. Gutu ◽  
Wesley Hebert ◽  
Jeffrey D. Wager ◽  
Lael M. Yonker ◽  
...  
Keyword(s):  
Cystic Fibrosis ◽  
Pseudomonas Aeruginosa ◽  
Lactate Dehydrogenase ◽  
Stationary Phase ◽  
Persister Cells ◽  
Content Type ◽  
Ph Values ◽  
Potentiation Effect ◽  
Metabolite Concentrations ◽  
Ldh Assay

ABSTRACTBacterial persisters are a quasidormant subpopulation of cells that are tolerant to antibiotic treatment. The combination of the aminoglycoside tobramycin with fumarate as an antibacterial potentiator utilizes an antipersister strategy that is aimed at reducing recurrentPseudomonas aeruginosainfections by enhancing the killing ofP. aeruginosapersisters. Stationary-phase cultures ofP. aeruginosawere used to generate persister cells. A range of tobramycin concentrations was tested with a range of metabolite concentrations to determine the potentiation effect of the metabolite under a variety of conditions, including a range of pH values and in the presence of azithromycin or cystic fibrosis (CF) patient sputum. In addition, 96-well dish biofilm and colony biofilm assays were performed, and the cytotoxicity of the tobramycin-fumarate combination was determined utilizing a lactate dehydrogenase (LDH) assay. Enhanced killing of up to 6 orders of magnitude ofP. aeruginosapersisters over a range of CF isolates, including mucoid and nonmucoid strains, was observed for the tobramycin-fumarate combination compared to killing with tobramycin alone. Furthermore, significant fumarate-mediated potentiation was seen in the presence of azithromycin or CF patient sputum. Fumarate also reduced the cytotoxicity of tobramycin-treatedP. aeruginosato human epithelial airway cells. Finally, in mucoid and nonmucoid CF isolates, complete eradication ofP. aeruginosabiofilm was observed in the colony biofilm assay due to fumarate potentiation. These data suggest that a combination of tobramycin with fumarate as an antibacterial potentiator may be an attractive therapeutic for eliminating recurrentP. aeruginosainfections in CF patients through the eradication of bacterial persisters.

scholarly journals Modulation of mRNA Stability Participates in Stationary-Phase-Specific Expression of Ribosome Modulation Factor

2005 ◽  
Vol 187 (6) ◽  
pp. 1951-1958 ◽  
Author(s):  
Toshiko Aiso ◽  
Hideji Yoshida ◽  
Akira Wada ◽  
Reiko Ohki
Keyword(s):  
Stationary Phase ◽  
Growth Phase ◽  
Mrna Stability ◽  
De Novo ◽  
Fresh Medium ◽  
Specific Expression ◽  
Modulation Factor ◽  
Inactive Form ◽  
Regulation Of Growth ◽  
Ribosome Modulation Factor

ABSTRACT The expression of ribosome modulation factor (RMF) is induced during stationary phase in Escherichia coli. RMF participates in the dimerization of 70S ribosomes to form the 100S ribosome, which is the translationally inactive form of the ribosome. To elucidate the involvement of the control of mRNA stability in growth-phase-specific rmf expression, we investigated rmf mRNA stability in stationary-phase cells and cells inoculated into fresh medium. The rmf mRNA was found to have an extremely long half-life during stationary phase, whereas destabilization of this mRNA took place after the culture was inoculated into fresh medium. RMF and 100S ribosomes disappeared from cells 1 min after inoculation. In addition to control by ppGpp-dependent transcription, these results indicate that the modulation of rmf mRNA stability is also involved in the regulation of growth-phase-specific rmf expression. Unexpectedly, the postinoculation degradation of rmf mRNA was suppressed by the addition of rifampin, suggesting that de novo RNA synthesis is necessary for degradation. This degradation was also suppressed in both a poly(A) polymerase-deficient and an rne-131 mutant strain. We cloned and sequenced the 3′-proximal regions of rmf mRNAs and found that most of these 3′ ends terminated at the ρ-independent terminator with the addition of a one- to five-A oligo(A) tail in either stationary-phase or inoculated cells. No difference was observed in the length of the poly(A) tail between stationary-phase and inoculated cells. These results suggest that a certain postinoculation-specific regulatory factor participates in the destabilization of rmf mRNA and is dependent on polyadenylation.

scholarly journals Bacterial Persister Cell Formation and Dormancy

2013 ◽  
Vol 79 (23) ◽  
pp. 7116-7121 ◽  
Author(s):  
Thomas K. Wood ◽  
Stephen J. Knabel ◽  
Brian W. Kwan
Keyword(s):  
Stationary Phase ◽  
Cell Formation ◽  
Genetic Change ◽  
Bacterial Cells ◽  
Mechanistic Studies ◽  
Chronic Infections ◽  
Persister Cells ◽  
Metabolic State ◽  
Dormant State ◽  
Persister Cell

ABSTRACTBacterial cells may escape the effects of antibiotics without undergoing genetic change; these cells are known as persisters. Unlike resistant cells that grow in the presence of antibiotics, persister cells do not grow in the presence of antibiotics. These persister cells are a small fraction of exponentially growing cells (due to carryover from the inoculum) but become a significant fraction in the stationary phase and in biofilms (up to 1%). Critically, persister cells may be a major cause of chronic infections. The mechanism of persister cell formation is not well understood, and even the metabolic state of these cells is debated. Here, we review studies relevant to the formation of persister cells and their metabolic state and conclude that the best model for persister cells is still dormancy, with the latest mechanistic studies shedding light on how cells reach this dormant state.

scholarly journals Functional Sites of Ribosome Modulation Factor (RMF) Involved in the Formation of 100S Ribosome

2021 ◽  
Vol 8 ◽  
Author(s):  
Hideji Yoshida ◽  
Hideki Nakayama ◽  
Yasushi Maki ◽  
Masami Ueta ◽  
Chieko Wada ◽  
...  
Keyword(s):  
Amino Acids ◽  
Gradient Centrifugation ◽  
Essential Amino Acids ◽  
Sucrose Density Gradient ◽  
Bacterial Survival ◽  
Sucrose Density Gradient Centrifugation ◽  
Functional Sites ◽  
Ribosome Binding ◽  
Modulation Factor ◽  
Ribosome Modulation Factor

One of the important cellular events in all organisms is protein synthesis, which is catalyzed by ribosomes. The ribosomal activity is dependent on the environmental situation of the cell. Bacteria form 100S ribosomes, lacking translational activity, to survive under stress conditions such as nutrient starvation. The 100S ribosome is a dimer of two 70S ribosomes bridged through the 30S subunits. In some pathogens of gammaproteobacteria, such as Escherichia coli, Yersinia pestis, and Vibrio cholerae, the key factor for ribosomal dimerization is the small protein, ribosome modulation factor (RMF). When ribosomal dimerization by RMF is impaired, long-term bacterial survival is abolished. This shows that the interconversion system between active 70S ribosomes and inactive 100S ribosomes is an important survival strategy for bacteria. According to the results of several structural analyses, RMF does not directly connect two ribosomes, but binds to them and changes the conformation of their 30S subunits, thus promoting ribosomal dimerization. In this study, conserved RMF amino acids among 50 bacteria were selectively altered by mutagenesis to identify the residues involved in ribosome binding and dimerization. The activities of mutant RMF for ribosome binding and ribosome dimerization were measured using the sucrose density gradient centrifugation (SDGC) and western blotting methods. As a result, some essential amino acids of RMF for the ribosomal binding and dimerization were elucidated. Since the induction of RMF expression inhibits bacterial growth, the data on this protein could serve as information for the development of antibiotic or bacteriostatic agents.

3. Mutations and gene variants

2014 ◽  
Author(s):  
Jonathan Slack
Keyword(s):  
Cystic Fibrosis ◽  
Single Cell ◽  
Genetic Diseases ◽  
The Body ◽  
Gene Variants ◽  
Single Mutation ◽  
Gene Variant ◽  
Do So ◽  
New Mutations

All gene variants originate as mutations. Most variants in the genome of any given individual are not new mutations but have been inherited from previous generations. ‘Mutations and gene variants’ shows that mutations can occur in any cell of the body, but in order to be inherited they must occur in the DNA of the reproductive cells. There are numerous genetic diseases caused by a single mutation in one gene, and the examples considered here are cystic fibrosis, haemophilia, achondroplasia, and Holt-Oram Syndrome. In such cases, the inheritance of the abnormal gene variant follows simple Mendelian rules. The origin of cancer is explained as a combination of mutations occurring in a single cell of the body. Inherited gene variants predisposing to cancer do so because they reduce the number of new mutations required.

scholarly journals Single cell observations show persister cells wake based on ribosome content

2018 ◽  
Vol 20 (6) ◽  
pp. 2085-2098 ◽  
Author(s):  
Jun‐Seob Kim ◽  
Ryota Yamasaki ◽  
Sooyeon Song ◽  
Weiwei Zhang ◽  
Thomas K. Wood
Keyword(s):  
Single Cell ◽  
Persister Cells
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