scholarly journals The choline dehydrogenase BetA of Acinetobacter baumannii: a flavo‐protein responsible for osmotic stress protection

2021 ◽  
Author(s):  
Jennifer Breisch ◽  
Melanie Bendel ◽  
Beate Averhoff
Keyword(s):  
Osmotic Stress ◽  
Acinetobacter Baumannii ◽  
Choline Dehydrogenase ◽  
Stress Protection

scholarly journals Defining Gene-Phenotype Relationships in Acinetobacter baumannii through One-Step Chromosomal Gene Inactivation

mBio ◽  
2014 ◽  
Vol 5 (4) ◽  
Author(s):  
Ashley T. Tucker ◽  
Emily M. Nowicki ◽  
Joseph M. Boll ◽  
Gregory A. Knauf ◽  
Nora C. Burdis ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Antibiotic Resistance ◽  
Acinetobacter Baumannii ◽  
Genome Editing ◽  
Biofilm Formation ◽  
Content Type ◽  
Stress Protection ◽  
Biocide Resistance ◽  
Hospital Acquired ◽  
Oxidative Stress Protection

ABSTRACTRates of infection with hospital-acquiredAcinetobacter baumanniihave exploded over the past decade due to our inability to limit persistence and effectively treat disease.A. baumanniiquickly acquires antibiotic resistance, and its genome encodes mechanisms to tolerate biocides and desiccation, which enhance its persistence in hospital settings. With depleted antibiotic options, new methods to treatA. baumanniiinfections are desperately needed. A comprehensive understanding detailingA. baumanniicellular factors that contribute to its resiliency at genetic and mechanistic levels is vital to the development of new treatment options. Tools to rapidly dissect theA. baumanniigenome will facilitate this goal by quickly advancing our understanding ofA. baumanniigene-phenotype relationships. We describe here a recombination-mediated genetic engineering (recombineering) system for targeted genome editing ofA. baumannii. We have demonstrated that this system can perform directed mutagenesis on wide-ranging genes and operons and is functional in various strains ofA. baumannii, indicating its broad application. We utilized this system to investigate key gene-phenotype relationships inA. baumanniibiology important to infection and persistence in hospitals, including oxidative stress protection, biocide resistance mechanisms, and biofilm formation. In addition, we have demonstrated that both the formation and movement of type IV pili play an important role inA. baumanniibiofilm.IMPORTANCEAcinetobacter baumanniiis the causative agent of hospital-acquired infections, including pneumonia and serious blood and wound infections.A. baumanniiis an emerging pathogen and has become a threat to public health because it quickly develops antibiotic resistance, making treatment difficult or impossible. While the threat ofA. baumanniiis well recognized, our understanding of even its most basic biology lags behind. Analysis ofA. baumanniicellular functions to identify potential targets for drug development has stalled due in part to laborious genetic techniques. Here we have pioneered a novel recombineering system that facilitates efficient genome editing inA. baumanniiby single PCR products. This technology allows for rapid genome editing to quickly ascertain gene-phenotype relationships. To demonstrate the power of recombineering in dissectingA. baumanniibiology, we use this system to establish key gene-phenotype relationships important to infection and persistence in hospitals, including oxidative stress protection, biocide resistance, and biofilm formation.

DksA is a central regulatory switch for stress protection and virulence in Acinetobacter baumannii

2021 ◽  
Author(s):  
Ram Maharjan ◽  
Geraldine Sulivan ◽  
Felise Adams ◽  
Natasha Delgado ◽  
Lucie Semenec ◽  
...  
Keyword(s):  
Stress Response ◽  
Acinetobacter Baumannii ◽  
Galleria Mellonella ◽  
Resistance Mechanisms ◽  
Term Survival ◽  
In Vivo Models ◽  
Molecular Control ◽  
Stress Protection

Abstract Bacterial coordination of stress resistance mechanisms in harsh environments is key to long-term survival and evolutionary success. In many Gram-negative pathogens, both general- and specific-stress response are controlled by alternative sigma factors such as RpoS. The critically important pathogen Acinetobacter baumannii is notoriously recalcitrant to external stressors, yet it lacks RpoS, so the molecular control of its resilience remains unclear. Here, we used transposon insertion sequencing to characterize the molecular responses of Acinetobacter baumannii to two biologically-important metals stressors, zinc and copper, and discovered that the transcriptional regulator DksA acts as a major regulatory stress-protection switch. We mapped the highly pleiotropic nature of DksA using transcriptomics and phenomics and found that it controls ribosomal protein expression, metabolism of gluconeogenic substrates and survival in stresses that cause oxidative damage. A. baumannii strains lacking DksA were no longer virulent in both murine and Galleria mellonella in vivo models. In vitro, DksA mutants exhibited increased sensitivity to human serum and antibiotics yet promoted biofilm and capsule formation. Our study provides detailed insight into the unique role that DksA plays in stress protection and virulence for A. baumannii and lays the groundwork for understanding of RpoS-independent regulatory general stress response.

scholarly journals In vivo analysis of LEA-1 in C. elegans desiccation tolerance

2021 ◽  
Author(s):  
Jonathan D. Hibshman ◽  
Bob Goldstein
Keyword(s):  
Osmotic Stress ◽  
Late Embryogenesis Abundant ◽  
The Body ◽  
Disordered Proteins ◽  
Lea Proteins ◽  
Fluorescent Reporters ◽  
C Elegans ◽  
Stress Protection

AbstractCells and organisms typically cannot survive in the absence of water. However, there are some notable exceptions, including animals such as nematodes, tardigrades, rotifers, and some arthropods. One class of proteins known to play a role in desiccation resistance is the late embryogenesis abundant (LEA) proteins. These largely disordered proteins protect plants and animals alike from desiccation. A multitude of studies have characterized stress-protective functions of LEA proteins in vitro and in heterologous systems. However, the extent to which LEA proteins exhibit the same functions in their native contexts in animals is unclear. Furthermore, nothing is known about the distribution of LEA proteins in multicellular organisms or tissue-specific requirements in conferring stress protection. To study the endogenous function of LEA proteins in an animal, we created a true null mutant of C. elegans LEA-1, as well as endogenous fluorescent reporters of the protein. We confirmed that C. elegans lacking LEA-1 are sensitive to desiccation. LEA-1 mutant animals were also sensitive to heat and osmotic stress and were prone to protein aggregation. During desiccation, LEA-1 expression increased and became more widespread throughout the body. LEA-1 was required at high levels in body wall muscle for animals to survive desiccation and osmotic stress. We identified minimal motifs within LEA-1 that are sufficient to increase desiccation survival of E. coli. Our results provide insights into the endogenous functions and expression dynamics of an LEA protein in a multicellular animal. We show that LEA-1 buffers animals from a broad range of stresses. Our identification of functional motifs within the protein suggests the possibility of engineering LEA-1-derived peptides for desiccation protection.

scholarly journals Human pleural fluid triggers global changes in the transcriptional landscape of Acinetobacter baumannii as an adaptive response to stress

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Jasmine Martinez ◽  
Jennifer S. Fernandez ◽  
Christine Liu ◽  
Amparo Hoard ◽  
Anthony Mendoza ◽  
...  
Keyword(s):  
Osmotic Stress ◽  
Acinetobacter Baumannii ◽  
Pleural Fluid ◽  
Biofilm Formation ◽  
Membrane Vesicle ◽  
Global Changes ◽  
Expression Of Genes ◽  
Response To Stress ◽  
The Impact ◽  
Dna Acquisition

AbstractAcinetobacter baumannii is a feared, drug-resistant pathogen, characterized by its ability to resist extreme environmental and nutrient-deprived conditions. Previously, we showed that human serum albumin (HSA) can increase foreign DNA acquisition specifically and alter the expression of genes associated with pathogenicity. Moreover, in a recent genome-wide transcriptomic study, we observed that pleural fluid (PF), an HSA-containing fluid, increases DNA acquisition, can modulate cytotoxicity, and control immune responses by eliciting changes in the A. baumannii metabolic profile. In the present work, using more stringent criteria and focusing on the analysis of genes related to pathogenicity and response to stress, we analyzed our previous RNA-seq data and performed phenotypic assays to further explore the impact of PF on A. baumannii’s microbial behavior and the strategies used to overcome environmental stress. We observed that PF triggered differential expression of genes associated with motility, efflux pumps, antimicrobial resistance, biofilm formation, two-component systems (TCSs), capsule synthesis, osmotic stress, and DNA-damage response, among other categories. Phenotypic assays of A. baumannii A118 and two other clinical A. baumannii strains, revealed differences in their responses to PF in motility, biofilm formation, antibiotic susceptibility, osmotic stress, and outer membrane vesicle (OMV) production, suggesting that these changes are strain specific. We conclude that A. baumannii’s pathoadaptive responses is induced by HSA-containing fluids and must be part of this bacterium armamentarium to persist in hostile environments.

scholarly journals Improving l-threonine production in Escherichia coli by elimination of transporters ProP and ProVWX

2021 ◽  
Vol 20 (1) ◽  
Author(s):  
Shuaiwen Wang ◽  
Yu Fang ◽  
Zhen Wang ◽  
Shuyan Zhang ◽  
Liangjia Wang ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Osmotic Stress ◽  
Abc Transporter ◽  
Glycine Betaine ◽  
Compatible Solute ◽  
E Coli ◽  
Stress Protection ◽  
Threonine Production ◽  
Glucose 6 Phosphate Dehydrogenase ◽  
Betaine Supplementation

Abstract Background Betaine, an osmoprotective compatible solute, has been used to improve l-threonine production in engineered Escherichia colil-threonine producer. Betaine supplementation upregulates the expression of zwf encoding glucose-6-phosphate dehydrogenase, leading to the increase of NADPH, which is beneficial for l-threonine production. In E. coli, betaine can be taken through ProP encoded by proP or ProVWX encoded by proVWX. ProP is a H+-osmolyte symporter, whereas ProVWX is an ABC transporter. ProP and ProVWX mediate osmotic stress protection by transporting zwitterionic osmolytes, including glycine betaine. Betaine can also be synthesized in E. coli by enzymes encoded by betABIT. However, the influence of ProP, ProVWX and betABIT on l-threonine production in E. coli has not been investigated. Results In this study, the influence of ProP, ProVWX and betABIT on l-threonine production in E. coli has been investigated. Addition of betaine slightly improved the growth of the l-threonine producing E. coli strain TWF001 as well as the l-threonine production. Deletion of betABIT retarded the growth of TWF001 and slightly decreased the l-threonine production. However, deletion of proP or/and proVWX significantly increased the l-threonine production. When proP was deleted, the l-threonine production increased 33.3%; when proVWX was deleted, the l-threonine production increased 40.0%. When both proP and proVWX were deleted, the resulting strain TSW003 produced 23.5 g/l l-threonine after 36 h flask cultivation. The genes betABIT, proC, fadR, crr and ptsG were individually deleted from TSW003, and it was found that further absence of either crr (TWS008) or ptsG (TWS009) improved l-threonine production. TSW008 produced 24.9 g/l l-threonine after 36 h flask cultivation with a yield of 0.62 g/g glucose and a productivity of 0.69 g/l/h. TSW009 produced 26 g/l l-threonine after 48 h flask cultivation with a yield of 0.65 g/g glucose and a productivity of 0.54 g/l/h, which is 116% increase compared to the control TWF001. Conclusions In this study, l-threonine-producing E. coli strains TSW008 and TSW009 with high l-threonine productivity were developed by regulating the intracellular osmotic pressure. This strategy could be used to improve the production of other products in microorganisms.

White clover sensitivity to UV-B radiation - biochemical relationships and interaction with drought

2003 ◽  
pp. 273-277
Author(s):  
R.W. Hofmann ◽  
B.D. Campbell ◽  
E.E. Swinny ◽  
S.J. Bloor ◽  
K.R. Markham ◽  
...  
Keyword(s):  
New Zealand ◽  
White Clover ◽  
Ultraviolet B ◽  
Summer Drought ◽  
Stress Protection ◽  
Trade Offs ◽  
Population Comparisons ◽  
Potential Benefits ◽  
Quercetin Glycosides ◽  
Balance Trade

During summertime in New Zealand, white clover experiences high levels of ultraviolet-B (UV-B) radiation. This frequently coincides with periods of summer drought. We investigated responses to UV-B and to the combination of UV-B and drought in various white clover populations, including New Zealand cultivars and ecotypes as well as overseas germplasm. The results were obtained under controlled environmental conditions in three independent trials. Overall, white clover growth was reduced by UV-B. The population comparisons indicated that low growth rate and adaptation to other forms of stress may be related to UV-B tolerance under well-watered conditions, but not during extended periods of drought. Flavonoid pigments that are involved in stress protection were strongly increased under UV-B and were further enhanced in the combination of UV-B and drought. The responses among these flavonoids were highly specific, with more pronounced UV-B-induced increases in quercetin glycosides, compared to their closely related kaempferol counterparts. UV-B toler ance of the less productive white clover populations was linked to the accumulation of quercetin compounds. In conclusion, these studies suggest (i) that slow-growing white clover ecotypes adapted to other stresses have higher capacity for biochemical acclimation to UV-B under well-watered conditions and (ii) that these biochemical attributes may also contribute to decreased UV-B sensitivity across white clover populations under drought. The findings alert plant breeders to potential benefits of selecting productive germplasm for high levels of specific flavonoids to balance trade-offs between plant productivity and stress tolerance. Keywords: Drought, flavonoids, genetic variation, HPLC, kaempferol, quercetin, str ess, Trifolium repens L., ultraviolet-B, white clover

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