scholarly journals Toxin, toxin-coregulated pili, and the toxR regulon are essential for Vibrio cholerae pathogenesis in humans.

1988 ◽  
Vol 168 (4) ◽  
pp. 1487-1492 ◽  
Author(s):  
D A Herrington ◽  
R H Hall ◽  
G Losonsky ◽  
J J Mekalanos ◽  
R K Taylor ◽  
...  
Keyword(s):  
Cholera Toxin ◽  
Regulatory Protein ◽  
Deletion Mutation ◽  
Intestinal Colonization ◽  
Gene Encoding ◽  
Disease Symptoms ◽  
Mutant Strains ◽  
A Subunit ◽  
First Time

Isogenic mutant strains of V. cholerae O1 lacking elements of a genetic regulon controlled by toxR and implicated in virulence were tested in volunteers. A deletion mutation in ctxA, the gene encoding the A subunit of cholera toxin, markedly attenuated disease symptoms without affecting intestinal colonization. Deletion of toxR, the gene encoding the cholera toxin-positive regulatory protein resulted in a diminution in colonizing capacity. A deletion mutation in tcpA, encoding the major subunit of the toxin coregulated pilus (regulated by toxR), abolished the colonizing capacity of this strain. These results show for the first time the role of a specific pilus structure in colonization of the human intestine by V. cholerae O1 and exemplify the significance of a genetic regulon in pathogenesis.

scholarly journals Expression of Mycobacterium smegmatisPyrazinamidase in Mycobacterium tuberculosis Confers Hypersensitivity to Pyrazinamide and Related Amides

2000 ◽  
Vol 182 (19) ◽  
pp. 5479-5485 ◽  
Author(s):  
Helena I. M. Boshoff ◽  
Valerie Mizrahi
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Mycobacterium Smegmatis ◽  
Intracellular Localization ◽  
Wild Type ◽  
Gene Encoding ◽  
Allelic Exchange ◽  
Mutant Strains ◽  
Established Role ◽  
Amidase Gene

ABSTRACT A pyrazinamidase (PZase)-deficient pncA mutant ofMycobacterium tuberculosis, constructed by allelic exchange, was used to investigate the effects of heterologous amidase gene expression on the susceptibility of this organism to pyrazinamide (PZA) and related amides. The mutant was highly resistant to PZA (MIC, >2,000 μg/ml), in accordance with the well-established role ofpncA in the PZA susceptibility of M. tuberculosis (A. Scorpio and Y. Zhang, Nat. Med. 2:662–667, 1996). Integration of the pzaA gene encoding the major PZase/nicotinamidase from Mycobacterium smegmatis (H. I. M. Boshoff and V. Mizrahi, J. Bacteriol. 180:5809–5814, 1998) or the M. tuberculosis pncA gene into the pncAmutant complemented its PZase/nicotinamidase defect. In bothpzaA- and pncA-complemented mutant strains, the PZase activity was detected exclusively in the cytoplasm, suggesting an intracellular localization for PzaA and PncA. ThepzaA-complemented strain was hypersensitive to PZA (MIC, ≤10 μg/ml) and nicotinamide (MIC, ≥20 μg/ml) and was also sensitive to benzamide (MIC, 20 μg/ml), unlike the wild-type andpncA-complemented mutant strains, which were highly resistant to this amide (MIC, >500 μg/ml). This finding was consistent with the observation that benzamide is hydrolyzed by PzaA but not by PncA. Overexpression of PzaA also conferred sensitivity to PZA, nicotinamide, and benzamide on M. smegmatis (MIC, 150 μg/ml in all cases) and rendered Escherichia colihypersensitive for growth at low pH.

scholarly journals The Orphan Receptor CRF2-4 Is an Essential Subunit of the Interleukin 10 Receptor

1998 ◽  
Vol 187 (4) ◽  
pp. 571-578 ◽  
Author(s):  
Susan D. Spencer ◽  
Francesco Di Marco ◽  
Jeff Hooley ◽  
Sharon Pitts-Meek ◽  
Michele Bauer ◽  
...  
Keyword(s):  
Interleukin 10 ◽  
Mutant Mice ◽  
Chromosome 21 ◽  
Orphan Receptor ◽  
Type I ◽  
Gene Encoding ◽  
Mediated Effects ◽  
A Subunit ◽  
Essential Subunit

The orphan receptor CRF2-4 is a member of the class II cytokine receptor family (CRF2), which includes the interferon receptors, the interleukin (IL) 10 receptor, and tissue factor. CRFB4, the gene encoding CRF2-4, is located within a gene cluster on human chromosome 21 that comprises three interferon receptor subunits. To elucidate the role of CRF2-4, we disrupted the CRFB4 gene in mice by means of homologous recombination. Mice lacking CRF2-4 show no overt abnormalities, grow normally, and are fertile. CRF2-4 deficient cells are normally responsive to type I and type II interferons, but lack responsiveness to IL-10. By ∼12 wk of age, the majority of mutant mice raised in a conventional facility developed a chronic colitis and splenomegaly. Thus, CRFB4 mutant mice recapitulate the phenotype of IL-10–deficient mice. These findings suggest that CRF2-4 is essential for IL-10–mediated effects and is a subunit of the IL-10 receptor.

scholarly journals Adenylate Cyclase Activation by Cholera Toxin in Pig Epidermis: An Obligatory Role of the GTP-Regulatory Protein

1983 ◽  
Vol 81 (2) ◽  
pp. 131-136 ◽  
Author(s):  
Junji Takeda ◽  
Kenji Adachi ◽  
Kenneth M. Halprin ◽  
Victor Levine ◽  
Clyde Woodyard
Keyword(s):  
Adenylate Cyclase ◽  
Cholera Toxin ◽  
Regulatory Protein ◽  
Cyclase Activation ◽  
Adenylate Cyclase Activation

THE ROLE OF PROTON ATPase SPECIFIC INHIBITOR $N,N'$-DICYCLOHEXYLCARBODIIMIDE AND EXTERNAL FORMATE CONCENTRATION ON $E.~COLI$ GROWTH DURING MIXED CARBON SOURCES FERMENTATION AT DIFFERENT pHs.

2021 ◽  
Vol 55 (1 (254)) ◽  
pp. 67-74
Author(s):  
Heghine Kh. Gevorgyan ◽  
Anait V. Vassilian ◽  
Karen A. Trchounian
Keyword(s):  
Regulatory Protein ◽  
Carbon Sources ◽  
Specific Inhibitor ◽  
Inhibitory Effect ◽  
E Coli ◽  
Proton Atpase ◽  
Mutant Strains ◽  
Atpase Inhibition ◽  
Compensatory Effect

This research is focused on the investigation of specific growth rate changes of $E.~coli$ wild type and mutant strains with defect of Hyd, FDH enzymes and FhlA regulatory protein in the presence of $N,N'$-dicyclohexylcarbodiimide (DCCD) and external formate various concentration during co-fermentation of glucose, glycerol and formate at pHs $5.5-7.5.$ The highest value of SGR was observed at pH 7.5. It was revealed that SGR depends on external formate concentration at all pHs. DCCD inhibitory effect was shown mainly at pH 7.5 and partially at pH 6.5 and 5.5. In the case of the F0F1-ATPase inhibition FhlA compensatory effect on SGR was revealed.

scholarly journals Molecular Analysis of Vibrio choleraeO1, O139, non-O1, and non-O139 Strains: Clonal Relationships between Clinical and Environmental Isolates

2001 ◽  
Vol 67 (2) ◽  
pp. 910-921 ◽  
Author(s):  
D. V. Singh ◽  
Maria H. Matte ◽  
G. R. Matte ◽  
Sunny Jiang ◽  
F. Sabeena ◽  
...  
Keyword(s):  
Cholera Toxin ◽  
Consensus Sequence ◽  
Regulatory Protein ◽  
Fluid Accumulation ◽  
Environmental Isolates ◽  
Gene Encoding ◽  
Ileal Loop ◽  
Heat Stable ◽  
Genes Encoding ◽  
Toxigenic Strains

ABSTRACT A total of 26 strains of Vibrio cholerae, including members of the O1, O139, and non-O1, non-O139 serogroups from both clinical and environmental sources, were examined for the presence of genes encoding cholera toxin (ctxA), zonula occludens toxin (zot), accessory cholera enterotoxin (ace), hemolysin (hlyA), NAG-specific heat-stable toxin (st), toxin-coregulated pilus (tcpA), and outer membrane protein (ompU), for genomic organization, and for the presence of the regulatory protein genes tcpI andtoxR in order to determine relationships between epidemic serotypes and sources of isolation. While 22 of the 26 strains were hemolytic on 5% sheep blood nutrient agar, all strains were PCR positive for hlyA, the hemolysin gene. When multiplex PCR was used, all serogroup O1 and O139 strains were positive fortcpA, ompU, and tcpI. All O1 and O139 strains except one O1 strain and one O139 strain were positive for the ctxA, zot, and ace genes. Also, O1 strain VO3 was negative for the zot gene. All of the non-O1, non-O139 strains were negative for the ctxA,zot, ace, tcpA, andtcpI genes, and all of the non-O1, non-O139 strains except strain VO26 were negative for ompU. All of the strains except non-O1, non-O139 strain VO22 were PCR positive for the gene encoding the central regulatory protein, toxR. All V. cholerae strains were negative for the NAG-specificst gene. Of the nine non-ctx-producing strains of V. cholerae, only one, non-O1, non-O139 strain VO24, caused fluid accumulation in the rabbit ileal loop assay. The other eight strains, including an O1 strain, an O139 strain, and six non-O1, non-O139 strains, regardless of the source of isolation, caused fluid accumulation after two to five serial passages through the rabbit gut. Culture filtrates of all non-cholera-toxigenic strains grown in AKI media also caused fluid accumulation, suggesting that a new toxin was produced in AKI medium by these strains. Studies of clonality performed by using enterobacterial repetitive intergenic consensus sequence PCR, Box element PCR, amplified fragment length polymorphism (AFLP), and pulsed-field gel electrophoresis (PFGE) collectively indicated that theV. cholerae O1 and O139 strains had a clonal origin, whereas the non-O1, non-O139 strains belonged to different clones. The clinical isolates closely resembled environmental isolates in their genomic patterns. Overall, there was an excellent correlation among the results of the PCR, AFLP, and PFGE analyses, and individual strains derived from clinical and environmental sources produced similar fingerprint patterns. From the results of this study, we concluded that the non-cholera-toxin-producing strains of V. cholerae, whether of clinical or environmental origin, possess the ability to produce a new secretogenic toxin that is entirely different from the toxin produced by toxigenic V. cholerae O1 and O139 strains. We also concluded that the aquatic environment is a reservoir for V. cholerae O1, O139, non-O1, and non-O139 serogroup strains.

scholarly journals Contribution of Fibronectin-Binding Protein to Pathogenesis of Streptococcus suis Serotype 2

2002 ◽  
Vol 70 (3) ◽  
pp. 1319-1325 ◽  
Author(s):  
Astrid de Greeff ◽  
Herma Buys ◽  
Robin Verhaar ◽  
Janny Dijkstra ◽  
Loek van Alphen ◽  
...  
Keyword(s):  
Binding Protein ◽  
Streptococcus Suis ◽  
Infection Model ◽  
Organ Cultures ◽  
Gene Encoding ◽  
E Coli ◽  
Mutant Strains ◽  
Fibronectin Binding Protein ◽  
Suis Serotype

ABSTRACT In the present study we investigated the role of the fibronectin (FN)- and fibrinogen (FGN)-binding protein (FBPS) in the pathogenesis of Streptococcus suis serotype 2 in piglets. The complete gene encoding FBPS from S. suis serotype 2 was cloned in Escherichia coli and sequenced. The occurrence of the gene in various serotypes was analyzed by hybridization studies. The FBPS protein was expressed in E. coli and purified, and binding to human FN and FGN was demonstrated. The induction of antibodies in piglets was studied upon infection. An isogenic mutant unable to produce FBPS was constructed, and the levels of virulence of the wild-type and mutant strains were compared in a competitive infection model in young piglets. Organ cultures showed that FBPS was not required for colonization of the tonsils but that FBPS played a role in the colonization of the specific organs involved in an S. suis infection. Therefore, the FBPS mutant was considered as an attenuated mutant.

scholarly journals Understanding the Role of Prevotella Genus in the Digestion of Lignocellulose and Other Substrates in Vietnamese Native Goats’ Rumen by Metagenomic Deep Sequencing

Animals ◽  
2021 ◽  
Vol 11 (11) ◽  
pp. 3257
Author(s):  
Trong-Khoa Dao ◽  
Thi-Huyen Do ◽  
Ngoc-Giang Le ◽  
Hong-Duong Nguyen ◽  
Thi-Quy Nguyen ◽  
...  
Keyword(s):  
Deep Sequencing ◽  
Gene Encoding ◽  
Amylolytic Enzymes ◽  
E Coli ◽  
Lignocellulose Conversion ◽  
Genes Encoding ◽  
Abundant Genus ◽  
First Time ◽  
Goat Rumen

Bacteria in rumen play pivotal roles in the digestion of nutrients to support energy for the host. In this study, metagenomic deep sequencing of bacterial metagenome extracted from the goats’ rumen generated 48.66 GB of data with 3,411,867 contigs and 5,367,270 genes. The genes were mainly functionally annotated by Kyoto Encyclopedia of Genes and Genomes (KEGG) Carbohydrate-Active enZYmes (CAZy), and HMMER database, and taxonomically classified by MEGAN. As a result, 65,554 genes encoding for 30 enzymes/proteins related to lignocellulose conversion were exploited, in which nine enzymes were seen for the first time in goat rumen. Prevotella was the most abundant genus, contributing 30% hemicellulases and 36% enzymes/proteins for lignocellulose pretreatment, and supporting 98.8% of feruloyl esterases and 71.7% acetylxylan esterases. In addition, 18 of the 22 most lignocellulose digesting- potential contigs belonged to Prevotella. Besides, Prevotella possessed many genes coding for amylolytic enzymes. One gene encoding for endoxylanase was successfully expressed in E. coli. The recombinant enzyme had high Vmax, was tolerant to some salts and detergents, worked better at pH 5.5–6.5, temperature 40–50 °C, and was capable to be used in practices. Based on these findings, we confirm that Prevotella plays a pivotal role for hemicellulose digestion and significantly participates in starch, cellulose, hemicellulose, and pectin digestion in the goat rumen.

scholarly journals Suppressors of translation initiation defect in hem12 locus of Saccharomyces cerevisiae.

2000 ◽  
Vol 47 (1) ◽  
pp. 181-190 ◽  
Author(s):  
M Góra ◽  
K Pluta ◽  
A Chelstowska ◽  
T Zoładek
Keyword(s):  
Mutational Analysis ◽  
Carbon Sources ◽  
Open Reading Frames ◽  
Initiation Factor ◽  
Uroporphyrinogen Decarboxylase ◽  
Gene Encoding ◽  
A Subunit ◽  
Extragenic Suppressors ◽  
Cis And Trans

A system for the positive selection of transational initiation suppressors in S. cerevisiae has been developed. A mutant with an ATA initiation codon in the HEM12 gene, encoding uroporphyrinogen decarboxylase, was used to select cis- and trans-acting suppressors. These suppressors partially restore growth on nonfermentable carbon sources, such as glycerol, but still allow the accumulation of porphyrins. All extragenic suppressors are mapped to the SUI1 locus, encoding initiation factor eIF1. The effect of the hem12 mutation is also partially reversed by the known SUI3 suppressor encoding the beta subunit of eIF2. In contrast, the sui2 suppressor encoding the a subunit of eIF2 does not affect the hem12 phenotype. The intragenic suppressors are able to restore the translation of hem12 due to the generation of additional, in frame AUG codons upstream of the hem12-14 mutation. Mutational analysis of the HEM12 leader sequence was also performed to determine the role of small open reading frames (uORFs) present upstream of the HEM12 ORF. Studies on the expression of integrated hem12-1/4-lacZ fusion, devoid of all upstream ATGs, indicate a lack of regulatory effect of uORFs on HEM12 translation.

scholarly journals Novel insights into N-glycan fucosylation and core xylosylation in C. reinhardtii

2019 ◽  
Author(s):  
Anne Oltmanns ◽  
Lara Hoepfner ◽  
Martin Scholz ◽  
Karen Zinzius ◽  
Stefan Schulze ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Candidate Gene ◽  
Chlamydomonas Reinhardtii ◽  
Candidate Genes ◽  
Protein Function ◽  
Double Mutant ◽  
Gene Encoding ◽  
Mutant Strains ◽  
Glycosylation Pathway

AbstractChlamydomonas reinhardtii N-glycans carry plant typical β1,2-core xylose, α1,3-fucose residues as well as plant atypical terminal β1,4-xylose and methylated mannoses. In a recent study, XylT1A was shown to act as core xylosyltransferase, whereby its action was of importance for an inhibition of excessive Man1A dependent trimming. N-Glycans found in a XylT1A/Man1A double mutant carried core xylose residues, suggesting the existence of a second core xylosyltransferase in C. reinhardtii. To further elucidate enzymes important for N-glycosylation, novel single knockdown mutants of candidate genes involved in the N-glycosylation pathway were characterized. In addition, double, triple and quadruple mutants affecting already known N-glycosylation pathway genes were generated. By characterizing N-glycan compositions of intact N-glycopeptides from these mutant strains by mass spectrometry, a candidate gene encoding for a second putative core xylosyltransferase (XylT1B) was identified. Additionally, the role of a putative fucosyltransferase was revealed. Mutant strains with knockdown of both xylosyltransferases and the fucosyltransferase resulted in the formation of N-glycans with strongly diminished core modifications. Thus, the mutant strains generated will pave the way for further investigations on how single N-glycan core epitopes modulate protein function in C. reinhardtii.Significance StatementOur data provide novel insights into the function of XylT1B and FucT in C. reinhardtii as N-glycan core modifying enzymes. In the course of our study, different mutants were created by genetic crosses showing either varying or a lack of N-glycan core modification, enabling comparative analyses in relation to single N-glycan core epitope and overall protein function in C. reinhardtii.

scholarly journals C-Terminal Truncation of α-COP Affects Functioning of Secretory Organelles and Calcium Homeostasis in Hansenula polymorpha

2004 ◽  
Vol 3 (1) ◽  
pp. 52-60 ◽  
Author(s):  
Maria B. Chechenova ◽  
Nina V. Romanova ◽  
Alexander V. Deev ◽  
Anna N. Packeiser ◽  
Vladimir N. Smirnov ◽  
...  
Keyword(s):  
Intracellular Transport ◽  
Hansenula Polymorpha ◽  
Gene Encoding ◽  
Golgi Transport ◽  
Type Plasminogen Activator ◽  
Urokinase Type Plasminogen Activator ◽  
A Subunit ◽  
Terminal Amino ◽  
Mutant Phenotypes

ABSTRACT In eukaryotic cells, COPI vesicles retrieve resident proteins to the endoplasmic reticulum and mediate intra-Golgi transport. Here, we studied the Hansenula polymorpha homologue of the Saccharomyces cerevisiae RET1 gene, encoding α-COP, a subunit of the COPI protein complex. H. polymorpha ret1 mutants, which expressed truncated α-COP lacking more than 300 C-terminal amino acids, manifested an enhanced ability to secrete human urokinase-type plasminogen activator (uPA) and an inability to grow with a shortage of Ca2+ ions, whereas a lack of α-COP expression was lethal. The α-COP defect also caused alteration of intracellular transport of the glycosylphosphatidylinositol-anchored protein Gas1p, secretion of abnormal uPA forms, and reductions in the levels of Pmr1p, a Golgi Ca2+-ATPase. Overexpression of Pmr1p suppressed some ret1 mutant phenotypes, namely, Ca2+ dependence and enhanced uPA secretion. The role of COPI-dependent vesicular transport in cellular Ca2+ homeostasis is discussed.

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