scholarly journals Heightened virulence of Yersinia is associated with decreased function of the YopJ protein

2021 ◽  
Author(s):  
Chris A Mares ◽  
Fernando P Lugo ◽  
Mohammad Albataineh ◽  
Beth Goins ◽  
Irene Newton ◽  
...  
Keyword(s):  
Positive Selection ◽  
Sequence Divergence ◽  
Bacterial Pathogenesis ◽  
Effector Protein ◽  
Evolutionary Analysis ◽  
Yersinia Species ◽  
Macrophage Cytotoxicity ◽  
A Minor ◽  
Pathogenic Yersinia

Despite the maintenance of YopP/J alleles throughout the human-pathogenic Yersinia lineage, the benefit of YopP/J-induced phagocyte death for Yersinia pathogenesis in animals is not obvious. To determine how sequence divergence of YopP/J has impacted Yersinia virulence, we examined protein polymorphisms in this Type III secreted effector protein across 17 Yersinia species, and tested the consequences of polymorphism in a murine model of sub-acute systemic yersiniosis. Our evolutionary analysis revealed that codon 177 has been subjected to positive selection - the Y. enterocolitica residue had been altered from a leucine to a phenylalanine in nearly all Y. pseudotuberculosis and Y. pestis strains examined. Despite being a minor change, as both leucine and phenylalanine have hydrophobic side chains, reversion of YopJF177 to the ancestral YopJL177 variant yielded a Y. pseudotuberculosis strain with enhanced cytotoxicity towards macrophages, consistent with previous findings. Surprisingly, expression of YopJF177L in the mildly attenuated ksgA- background rendered the strain completely avirulent in mice. Consistent with this hypothesis that YopJ activity indirectly relates to Yersinia pathogenesis in vivo, ksgA- strains lacking functional YopJ failed to kill macrophages but actually regained virulence in animals. Also, treatment with the anti-apoptosis drug suramin prevented YopJ-mediated macrophage cytotoxicity and enhanced Y. pseudotuberculosis virulence in vivo. Our results demonstrate that Yersinia-induced cell death is detrimental for bacterial pathogenesis in this animal model of illness, and indicate that positive selection has driven YopJ/P and Yersinia evolution towards diminished cytotoxicity and increased virulence, respectively.

scholarly journals Heightened virulence of Yersinia is associated with decreased function of the YopJ protein

2021 ◽  
Author(s):  
Chris A. Mares ◽  
Fernando P. Lugo ◽  
Mohammad Albataineh ◽  
Beth A. Goins ◽  
Irene G. Newton ◽  
...  
Keyword(s):  
Animal Model ◽  
Cell Death ◽  
Positive Selection ◽  
Sequence Divergence ◽  
Bacterial Pathogenesis ◽  
Effector Protein ◽  
Evolutionary Analysis ◽  
Macrophage Cytotoxicity ◽  
A Minor

Despite the maintenance of YopP/J alleles throughout the human-pathogenic Yersinia lineage, the benefit of YopP/J-induced phagocyte death for Yersinia pathogenesis in animals is not obvious. To determine how sequence divergence of YopP/J has impacted Yersinia virulence, we examined protein polymorphisms in this Type III secreted effector protein across 17 Yersinia species, and tested the consequences of polymorphism in a murine model of sub-acute systemic yersiniosis. Our evolutionary analysis revealed that codon 177 has been subjected to positive selection - the Y. enterocolitica residue had been altered from a leucine to a phenylalanine in nearly all Y. pseudotuberculosis and Y. pestis strains examined. Despite being a minor change, as both leucine and phenylalanine have hydrophobic side chains, reversion of YopJ F177 to the ancestral YopJ L177 variant yielded a Y. pseudotuberculosis strain with enhanced cytotoxicity towards macrophages, consistent with previous findings. Surprisingly, expression of YopJ F177L in the mildly attenuated ksgA - background rendered the strain completely avirulent in mice. Consistent with this hypothesis that YopJ activity indirectly relates to Yersinia pathogenesis in vivo , ksgA - strains lacking functional YopJ failed to kill macrophages but actually regained virulence in animals. Also, treatment with the anti-apoptosis drug suramin prevented YopJ-mediated macrophage cytotoxicity and enhanced Y. pseudotuberculosis virulence in vivo . Our results demonstrate that Yersinia -induced cell death is detrimental for bacterial pathogenesis in this animal model of illness, and indicate that positive selection has driven YopJ/P and Yersinia evolution towards diminished cytotoxicity and increased virulence, respectively.

scholarly journals Metabolism of rat bone proteoglycans in vivo

1983 ◽  
Vol 216 (3) ◽  
pp. 589-596 ◽  
Author(s):  
C W Prince ◽  
F Rahemtulla ◽  
W T Butler
Keyword(s):  
Extraction Procedure ◽  
Total Radioactivity ◽  
Mineralized Tissue ◽  
Guanidinium Chloride ◽  
Disaccharide Composition ◽  
A Minor ◽  
Papain Digestion ◽  
Rat Bone

Former evaluations of the role of proteoglycans in mineralization have neglected to address the possibility that the metabolism of proteoglycans may be of significance in this regard. This problem was studied by using radiolabeling in vivo of rat calvaria with [35Sulphate for 2-72 h and a sequential extraction procedure to yield two pools of newly synthesized proteoglycans: one obtained from non-mineralized tissue by extraction with guanidinium chloride (GdmCl) and another obtained only after demineralization with EDTA. Total radioactivity in calvaria was maximal after 12 h of incorporation, but by 36 h had declined to a level that was about 55-65% of maximum. Radioactivity in the GdmCl extract declined steadily after 12 h, whereas that in the EDTA extract remained constant until 36 h, when it began to increase. Each extract contained a minor proteoglycan that eluted at the void volume (Vo) of a Sepharose CL-6B column. Unlike in the EDTA extract, this proteoglycan gradually disappeared from the GdmCl extract. Each extract also contained a major, smaller proteoglycan, with a Kav. of 0.24 and 0.36 in the GdmCl and EDTA extracts respectively. Papain digestion of each extract yielded glycosaminoglycan chains with Kav. values of 0.32 and 0.50 on CL-6B in the GdmCl and EDTA extracts respectively. Digestion of each extract with chondroitinase ABC and chondroitinase AC showed that the glycosaminoglycans were of similar disaccharide composition, with about 85% being 4-sulphated and the remainder 6-sulphated and/or iduronic acid-containing. These data suggest that about 45% of the newly synthesized proteoglycans are removed from the tissue during the course of mineralization.

scholarly journals Chronic Effects of a Salmonella Type III Secretion Effector Protein AvrA In Vivo

PLoS ONE ◽  
2010 ◽  
Vol 5 (5) ◽  
pp. e10505 ◽  
Author(s):  
Rong Lu ◽  
Shaoping Wu ◽  
Xingyin Liu ◽  
Yinglin Xia ◽  
Yong-guo Zhang ◽  
...  
Keyword(s):  
Type Iii Secretion ◽  
Effector Protein ◽  
Type Iii ◽  
Chronic Effects ◽  
Type Iii Secretion Effector

Potassium depletion improves myocardial potassium uptake in vivo

2004 ◽  
Vol 287 (1) ◽  
pp. C135-C141 ◽  
Author(s):  
Henning Bundgaard
Keyword(s):  
Uptake Rate ◽  
High Plasma ◽  
Potassium Depletion ◽  
Uptake Capacity ◽  
K Uptake ◽  
Impulse Generation ◽  
Uptake Rates ◽  
Clinical Interest ◽  
A Minor

Potassium depletion (KD) is a very common clinical entity often associated with adverse cardiac effects. KD is generally considered to reduce muscular Na-K-ATPase density and secondarily reduce K uptake capacity. In KD rats we evaluated myocardial Na-K-ATPase density, ion content, and myocardial K reuptake. KD for 2 wk reduced plasma K to 1.8 ± 0.1 vs. 3.5 ± 0.2 mM in controls ( P < 0.01, n = 7), myocardial K to 80 ± 1 vs. 86 ± 1 μmol/g wet wt ( P < 0.05, n = 7), increased Mg, and induced a tendency to increased Na. Myocardial Na-K-ATPase α2-subunit abundance was reduced by ∼30%, whereas increases in α1- and K-dependent pNPPase activity of 24% ( n = 6) and 13% ( n = 6), respectively, were seen. This indicates an overall upregulation of the myocardial Na-K pump pool. KD rats tolerated a higher intravenous KCl dose. KCl infusion until animals died increased myocardial K by 34% in KD rats and 18% in controls ( P < 0.05, n = 6 for both) but did not induce different net K uptake rates between groups. However, clamping plasma K at ∼5.5 mM by KCl infusion caused a higher net K uptake rate in KD rats (0.22 ± 0.04 vs. 0.10 ± 0.03 μmol·g wet wt−1·min−1; P < 0.05, n = 8). In conclusion, a minor KD-induced decrease in myocardial K increased Na-K pump density and in vivo increased K tolerance and net myocardial K uptake rate during K repletion. Thus the heart is protected from major K losses and accumulates considerable amounts of K during exposure to high plasma K. This is of clinical interest, because a therapeutically induced rise in myocardial K may affect contractility and impulse generation-propagation and may attenuate increased myocardial Na, the hallmark of heart failure.

scholarly journals Flagellar Morphogenesis: Protein Targeting and Assembly in the Paraflagellar Rod of Trypanosomes

1999 ◽  
Vol 19 (12) ◽  
pp. 8191-8200 ◽  
Author(s):  
Philippe Bastin ◽  
Thomas H. MacRae ◽  
Susan B. Francis ◽  
Keith R. Matthews ◽  
Keith Gull
Keyword(s):  
Cell Cycle ◽  
Trypanosoma Brucei ◽  
Protein Targeting ◽  
Fluorescent Protein ◽  
Green Fluorescent Protein Reporter ◽  
Mutant Proteins ◽  
Green Fluorescent ◽  
A Minor ◽  
Fluorescent Protein Reporter

ABSTRACT The paraflagellar rod (PFR) of the African trypanosomeTrypanosoma brucei represents an excellent model to study flagellum assembly. The PFR is an intraflagellar structure present alongside the axoneme and is composed of two major proteins, PFRA and PFRC. By inducible expression of a functional epitope-tagged PFRA protein, we have been able to monitor PFR assembly in vivo. As T. brucei cells progress through their cell cycle, they possess both an old and a new flagellum. The induction of expression of tagged PFRA in trypanosomes growing a new flagellum provided an excellent marker of newly synthesized subunits. This procedure showed two different sites of addition: a major, polar site at the distal tip of the flagellum and a minor, nonpolar site along the length of the partially assembled PFR. Moreover, we have observed turnover of epitope-tagged PFRA in old flagella that takes place throughout the length of the PFR structure. Expression of truncated PFRA mutant proteins identified a sequence necessary for flagellum localization by import or binding. This sequence was not sufficient to confer full flagellum localization to a green fluorescent protein reporter. A second sequence, necessary for the addition of PFRA protein to the distal tip, was also identified. In the absence of this sequence, the mutant PFRA proteins were localized both in the cytosol and in the flagellum where they could still be added along the length of the PFR. This seven-amino-acid sequence is conserved in all PFRA and PFRC proteins and shows homology to a sequence in the flagellar dynein heavy chain of Chlamydomonas reinhardtii.

scholarly journals Structural defects in rat liver deoxyribonucleic acid. Endogenous single-strained regions in comparison with damage induced in vivo by a carcinogen

1979 ◽  
Vol 179 (2) ◽  
pp. 341-352 ◽  
Author(s):  
B W Stewart ◽  
P H Huang ◽  
M J Brian
Keyword(s):  
Rat Liver ◽  
Structural Damage ◽  
Deae Cellulose ◽  
Minor Component ◽  
Structural Defects ◽  
Structural Abnormality ◽  
Denaturation Kinetics ◽  
Limited Digestion ◽  
A Minor

Rat liver DNA may be separated into two fractions by stepwise elution from benzoylated-DEAE-cellulose with NaCl and caffeine solutions respectively. Other studies using bacterical and yeast DNA suggested that the first fraction contains native DNA, whereas the second may exhibit some degree of single-stranded character. In the present experiments, chromatography of DNA was monitored by labelling in vivo with [methyl-3H]thymidine in rats previously subjected to partial hepatectomy. In animals killed up to 1 h after thymidine injection, radioactivity eluted in the second fraction was inversely related to the incorporation time, being greatest when animals were killed 10 min after radioisotope injection. However, for most experiments, animals were allowed to survive 2-4 weeks after surgery before use, analysis being made on non-dividing DNA. Under these conditions, the proportion of caffeine-eluted DNA was decreased by subjecting the preparation to shear, before chromatography. A procedure that resulted in 12% of the recovered radioactivity being eluted with caffeine was adopted for experiments involving comparisons of the two DNA fractions. Under these conditions, cross-contamination could be detected by rechromatography, but this did not preclude distinction being made between the two fractions in terms of DNA structure. NaCl-eluted DNA did not bind to nitrocellulose filters. Caffeine-eluted DNA was retained by the filters and released by washing with 3mM-Tris/HCl, pH9.4. The fractions did not differ in terms of isopycnic centrifugation in CsCl. The NaCl-eluted fraction migrated as a single band in polyacrylamide gels, and this pattern was not modified by prior digestion with Neurospora crassa endonuclease. In contrast, caffeine-eluted DNA contained a minor component having a wide molecular-weight distribution and was subject to limited digestion by the endonuclease. The kinetics of denaturation of NaCi-eluted DNA in the presence of formaldehyde, in common with unfractionated DNA, were consistent with double-stranded structure. The same analysis of caffeine-eluted DNA revealed structural abnormality equivalent to two defects per 10000 base-pairs. The data are consistent with the minor fraction of rat liver DNA, separated by using benzoylated-DEAE-cellulose, containing regions of local denaturation. We previously showed that administration of the hepatocarcinogen dimethylnitrosamine is associated with an increase in the proportion of caffeine-eluted DNA. In terms of most analysis, differences between DNA fraction from nitrosamine-treated rats were similar to differences exhibited by preparations from control animals. However, structural analysis using denaturation kinetics indicated defects in both the NaCl- and caffeine-eluted DNA isolated from nitrosamine-treated rats. The two fractions differed from each other in that caffeine-eluted DNA exhibited a degree of structural damage far greater than that detected in any preparation from control animals...

scholarly journals Evolutionary Analysis of Plastid Genomes of Seven Lonicera L. Species: Implications for Sequence Divergence and Phylogenetic Relationships

2018 ◽  
Vol 19 (12) ◽  
pp. 4039 ◽  
Author(s):  
Mi-Li Liu ◽  
Wei-Bing Fan ◽  
Ning Wang ◽  
Peng-Bin Dong ◽  
Ting-Ting Zhang ◽  
...  
Keyword(s):  
Molecular Genetic ◽  
Phylogenetic Reconstruction ◽  
Sequence Divergence ◽  
Genomic Analysis ◽  
Repeat Sequence ◽  
Biological Research ◽  
Comparative Genomic ◽  
Sequence Evolution ◽  
Evolutionary Analysis ◽  
Plastid Genomes

Plant plastomes play crucial roles in species evolution and phylogenetic reconstruction studies due to being maternally inherited and due to the moderate evolutionary rate of genomes. However, patterns of sequence divergence and molecular evolution of the plastid genomes in the horticulturally- and economically-important Lonicera L. species are poorly understood. In this study, we collected the complete plastomes of seven Lonicera species and determined the various repeat sequence variations and protein sequence evolution by comparative genomic analysis. A total of 498 repeats were identified in plastid genomes, which included tandem (130), dispersed (277), and palindromic (91) types of repeat variations. Simple sequence repeat (SSR) elements analysis indicated the enriched SSRs in seven genomes to be mononucleotides, followed by tetra-nucleotides, dinucleotides, tri-nucleotides, hex-nucleotides, and penta-nucleotides. We identified 18 divergence hotspot regions (rps15, rps16, rps18, rpl23, psaJ, infA, ycf1, trnN-GUU-ndhF, rpoC2-rpoC1, rbcL-psaI, trnI-CAU-ycf2, psbZ-trnG-UCC, trnK-UUU-rps16, infA-rps8, rpl14-rpl16, trnV-GAC-rrn16, trnL-UAA intron, and rps12-clpP) that could be used as the potential molecular genetic markers for the further study of population genetics and phylogenetic evolution of Lonicera species. We found that a large number of repeat sequences were distributed in the divergence hotspots of plastid genomes. Interestingly, 16 genes were determined under positive selection, which included four genes for the subunits of ribosome proteins (rps7, rpl2, rpl16, and rpl22), three genes for the subunits of photosystem proteins (psaJ, psbC, and ycf4), three NADH oxidoreductase genes (ndhB, ndhH, and ndhK), two subunits of ATP genes (atpA and atpB), and four other genes (infA, rbcL, ycf1, and ycf2). Phylogenetic analysis based on the whole plastome demonstrated that the seven Lonicera species form a highly-supported monophyletic clade. The availability of these plastid genomes provides important genetic information for further species identification and biological research on Lonicera.

Functional specificity of jejunal brush-border pteroylpolyglutamate hydrolase in pig

1991 ◽  
Vol 260 (6) ◽  
pp. G865-G872 ◽  
Author(s):  
C. J. Chandler ◽  
D. A. Harrison ◽  
C. A. Buffington ◽  
N. A. Santiago ◽  
C. H. Halsted
Keyword(s):  
Brush Border ◽  
Protein Band ◽  
Major Protein ◽  
Functional Specificity ◽  
Major Protein Band ◽  
Regional Location ◽  
Minor Protein ◽  
A Minor ◽  
Hydrolysis Of

To determine the functional specificity of intestinal brush-border pteroylpolyglutamate hydrolase (PPH), we compared the regional location of in vivo hydrolysis of pteroyltriglutamate (PteGlu3) with the location of activity and immunoreactivity of the enzyme in the pig. After in vivo incubations, PteGlu3 hydrolytic products were recovered from intestinal segments in the jejunum but not from the ileum. Brush-border PPH activity in fractionated mucosa was 10-fold greater in the jejunum than in the ileum, whereas the activity of intracellular PPH was increased in the distal ileum. Antibodies to purified brush-border PPH identified a major protein band at 120 kDa and a minor protein band at 195 kDa in solubilized jejunal brush border. Immunohistochemistry identified the enzyme only on the brush-border surface of the jejunum, whereas an immunoblot of solubilized brush-border membranes identified brush-border PPH in the jejunum but not in the ileum. The parallel of the regional location of in vivo hydrolysis of PteGlu3 with the location of brush-border PPH activity and immunoreactivity demonstrates the functional specificity of this enzyme in folate digestion.

scholarly journals Optogenetic Modulation of a Minor Fraction of Parvalbumin-Positive Interneurons Specifically Affects Spatiotemporal Dynamics of Spontaneous and Sensory-Evoked Activity in Mouse Somatosensory Cortex in Vivo

2017 ◽  
Vol 27 (12) ◽  
pp. 5784-5803 ◽  
Author(s):  
Jenq-Wei Yang ◽  
Pierre-Hugues Prouvot ◽  
Vicente Reyes-Puerta ◽  
Maik C Stüttgen ◽  
Albrecht Stroh ◽  
...  
Keyword(s):  
Somatosensory Cortex ◽  
Spatiotemporal Dynamics ◽  
Minor Fraction ◽  
Evoked Activity ◽  
Sensory Evoked ◽  
A Minor

scholarly journals Heterogeneity of Mycolactones Produced by Clinical Isolates of Mycobacterium ulcerans: Implications for Virulence

2003 ◽  
Vol 71 (2) ◽  
pp. 774-783 ◽  
Author(s):  
Armand Mve-Obiang ◽  
Richard E. Lee ◽  
Françoise Portaels ◽  
P. L. C. Small
Keyword(s):  
Clinical Evidence ◽  
Buruli Ulcer ◽  
Mycobacterium Ulcerans ◽  
Biologically Active ◽  
In Vivo Studies ◽  
Tissue Destruction ◽  
The Core ◽  
A Minor ◽  
Layer Chromatography

ABSTRACT Mycobacterium ulcerans is the causative agent of Buruli ulcer, a severe necrotizing skin disease endemic in tropical countries. Clinical evidence suggests that M. ulcerans isolates from Asia, Mexico, and Australia may be less virulent than isolates from Africa. In vivo studies suggest that mycolactone, a polyketide-derived macrolide toxin, plays a major role in the tissue destruction and immune suppression which occur in cases of Buruli ulcer. Mycolactones were extracted from 34 isolates of M. ulcerans representing strains from Africa, Malaysia, Asia, Australia, and Mexico. Thin-layer chromatography, mass spectroscopic analysis, and cytopathic assays of partially purified mycolactones from these isolates revealed that M. ulcerans produces a heterogeneous mixture of mycolactone variants. Mycolactone A/B, the most biologically active mycolactone species, was identified by mass spectroscopy as [M+Na]+ at m/z 765.5 in all cytotoxic isolates except for those from Mexico. Mycolactone C [M+Na]+ at m/z 726.3 was the dominant mycolactone species in eight Australian isolates, and mycolactone D [M+Na]+ m/z 781.2 was characteristic of two Asian strains. Mycolactone species are conserved within specific geographic areas, suggesting that there may be a correlation between mycolactone profile and virulence. In addition, the core lactone, [M+Na]+ m/z 447.4, was identified as a minor species, supporting the hypothesis that mycolactones are synthesized by two polyketide synthases. A cytopathic assay of the core lactone showed that this molecule is sufficient for cytotoxicity, although it is much less potent than the complete mycolactone.

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