scholarly journals AmiC Functions as an N-Acetylmuramyl-l-Alanine Amidase Necessaryfor Cell Separation and Can Promote Autolysis in Neisseria gonorrhoeae

2006 ◽  
Vol 188 (20) ◽  
pp. 7211-7221 ◽  
Author(s):  
Daniel L. Garcia ◽  
Joseph P. Dillard
Keyword(s):  
Neisseria Gonorrhoeae ◽  
Stationary Phase ◽  
Growth Characteristics ◽  
Wild Type ◽  
Cell Components ◽  
Section Electron ◽  
Cell Wall Breakdown ◽  
Mutant Cells

ABSTRACT Neisseria gonorrhoeae is prone to undergo autolysis under many conditions not conducive to growth. The role of autolysis during gonococcal infection is not known, but possible advantages for the bacterial population include provision of nutrients to a starving population, modulation of the host immune response by released cell components, and donation of DNA for natural transformation. Biochemical studies indicated that an N-acetylmuramyl-l-alanine amidase is responsible for cell wall breakdown during autolysis. In order to better understand autolysis and in hopes of creating a nonautolytic mutant, we mutated amiC, the gene for a putative peptidoglycan-degrading amidase in N. gonorrhoeae. Characterization of peptidoglycan fragments released during growth showed that an amiC mutant did not produce free disaccharide, consistent with a role for AmiC as an N-acetylmuramyl-l-alanine amidase. Compared to the wild-type parent, the mutant exhibited altered growth characteristics, including slowed exponential-phase growth, increased turbidity in stationary phase, and increased colony opacity. Thin-section electron micrographs showed that mutant cells did not fully separate but grew as clumps. Complementation of the amiC deletion mutant with wild-type amiC restored wild-type growth characteristics and transparent colony morphology. Overexpression of amiC resulted in increased cell lysis, supporting AmiC's purported function as a gonococcal autolysin. However, amiC mutants still underwent autolysis in stationary phase, indicating that other gonococcal enzymes are also involved in this process.

The in vivo role of annexin VII (synexin): characterization of an annexin VII-deficient Dictyostelium mutant indicates an involvement in Ca(2+)-regulated processes

1995 ◽  
Vol 108 (5) ◽  
pp. 2065-2076 ◽  
Author(s):  
V. Doring ◽  
F. Veretout ◽  
R. Albrecht ◽  
B. Muhlbauer ◽  
C. Schlatterer ◽  
...  
Keyword(s):  
Developmental Cycle ◽  
Cell Fractionation ◽  
Wild Type ◽  
Camp Phosphodiesterase ◽  
In The Wild ◽  
Extracellular Camp ◽  
Mutant Cells

Dictyostelium discoideum cells harbor two annexin VII isoforms of 47 and 51 kDa which are present throughout development. In immunofluorescence and cell fractionation studies annexin VII was found in the cytoplasm and on the plasma membrane. In gene disruption mutants lacking both annexin VII isoforms growth, pinocytosis, phagocytosis, chemotaxis and motility were not significantly impaired under routine laboratory conditions, and the cells were able to complete the developmental cycle on bacterial plates. On non-nutrient agar plates development was delayed by three to four hours and a significant number of aggregates was no longer able to form fruiting bodies. Exocytosis as determined by measuring extracellular cAMP phosphodiesterase, alpha-fucosidase and alpha-mannosidase activity was unaltered, the total amounts of these enzymes were however lower in the mutant than in the wild type. The mutant cells were markedly impaired when they were exposed to low Ca2+ concentrations by adding EGTA to the nutrient medium. Under these conditions growth, motility and chemotaxis were severely affected. The Ca2+ concentrations were similar in mutant and wild-type cells both under normal and Ca2+ limiting conditions; however, the distribution was altered under low Ca2+ conditions in SYN-cells. The data suggest that annexin VII is not required for membrane fusion events but rather contributes to proper Ca2+ homeostasis in the cell.

The role of catalase isozymes in the culturability of the root colonizer Pseudomonas putida after exposure to hydrogen peroxide and antibiotics

1994 ◽  
Vol 40 (5) ◽  
pp. 382-387 ◽  
Author(s):  
Martin G. Klotz ◽  
Anne J. Anderson
Keyword(s):  
Hydrogen Peroxide ◽  
Stationary Phase ◽  
Pseudomonas Putida ◽  
Growth Phase ◽  
Stationary Growth Phase ◽  
Exponential Phase ◽  
Wild Type ◽  
Early Stationary Growth Phase ◽  
Mutant Cells

The culturability of Pseudomonas putida cells after exposure to hydrogen peroxide and antibiotics was correlated with growth-dependent expression of catalase isozymes. Exponential phase wild-type cells, which contained catalase isozyme A, survived a 15-min treatment with less than 4 mM hydrogen peroxide, but were killed by higher concentrations. The culturability of P. putida mutant JIM, which lacked any functional catalase in exponential phase, was reduced by more than 75% after a 15-min exposure to ≥ 0.25 mM hydrogen peroxide. Because submillimolar concentrations of hydrogen peroxide are physiologically relevant in the bacterial cell, our results demonstrate that catalase isozyme A has essential housekeeping functions for growing cultures of P. putida. The accumulation of catalase isozymes B and C during growth into stationary phase coincided with a decrease in the sensitivity of wild-type and JIM cells of P. putida to hydrogen peroxide. Late stationary phase wild-type cells survived a 15-min exposure to even 50 mM hydrogen peroxide and mutant J1M cells survived exposure to 20 mM but not 50 mM hydrogen peroxide. The antibiotics tetracycline and kanamycin, which inhibit protein synthesis, were used to study the role of catalase induction in resistance to hydrogen peroxide. More than 40 and 80% of exponential phase cells of P. putida wild-type and J1M strains, respectively, were rendered nonculturable after a 20-min exposure to 45 μM tetracycline. Surprisingly, stationary phase cells of both P. putida strains were culturable after a 20-min exposure to tetracycline but remained sensitive to kanamycin. Exposure to tetracycline of stationary phase cells did not reduce the resistance of these cells to hydrogen peroxide. Tetracycline but not kanamycin increased the activity of catalase in lysates prepared from P. putida wild-type and mutant cells in early stationary growth phase. At this growth phase, only catalase isozyme B is operational in both strains, which suggests that tetracycline affects the activity of this enzyme.Key words: Pseudomonas putida, antibiotics, catalase, culturability, growth phase.

Characterization of a steroid hormone receptor gene and mRNA in wild-type and mutant cells

Nature ◽  
1984 ◽  
Vol 312 (5996) ◽  
pp. 779-781 ◽  
Author(s):  
Roger Miesfeld ◽  
Sam Okret ◽  
Ann-Charlotte Wikström ◽  
Örjan Wrange ◽  
Jan-Åke Gustafsson ◽  
...  
Keyword(s):  
Hormone Receptor ◽  
Steroid Hormone ◽  
Receptor Gene ◽  
Steroid Hormone Receptor ◽  
Wild Type ◽  
Mutant Cells ◽  
Hormone Receptor Gene

Characterization of the Role of the FluG Protein in Asexual Development of Aspergillus nidulans

Genetics ◽  
2001 ◽  
Vol 158 (3) ◽  
pp. 1027-1036 ◽  
Author(s):  
Cletus A D'Souza ◽  
Bee Na Lee ◽  
Thomas H Adams
Keyword(s):  
Aspergillus Nidulans ◽  
Negative Influence ◽  
Asexual Development ◽  
Wild Type ◽  
Significant Similarity ◽  
Developmental Defects ◽  
Asexual Sporulation ◽  
Type Strains

Abstract We showed previously that a ΔfluG mutation results in a block in Aspergillus nidulans asexual sporulation and that overexpression of fluG activates sporulation in liquid-submerged culture, a condition that does not normally support sporulation of wild-type strains. Here we demonstrate that the entire N-terminal region of FluG (∼400 amino acids) can be deleted without affecting sporulation, indicating that FluG activity resides in the C-terminal half of the protein, which bears significant similarity with GSI-type glutamine synthetases. While FluG has no apparent role in glutamine biosynthesis, we propose that it has an enzymatic role in sporulation factor production. We also describe the isolation of dominant suppressors of ΔfluG(dsg) that should identify components acting downstream of FluG and thereby define the function of FluG in sporulation. The dsgA1 mutation also suppresses the developmental defects resulting from ΔflbA and dominant activating fadA mutations, which both cause constitutive induction of the mycelial proliferation pathway. However, dsgA1 does not suppress the negative influence of these mutations on production of the aflatoxin precursor, sterigmatocystin, indicating that dsgA1 is specific for asexual development. Taken together, our studies define dsgA as a novel component of the asexual sporulation pathway.

scholarly journals Genetic analysis of the Arabidopsis TIR1/AFB auxin receptors reveals both overlapping and specialized functions

eLife ◽  
2020 ◽  
Vol 9 ◽  
Author(s):  
Michael J Prigge ◽  
Matthieu Platre ◽  
Nikita Kadakia ◽  
Yi Zhang ◽  
Kathleen Greenham ◽  
...  
Keyword(s):  
Genetic Analysis ◽  
Early Embryo ◽  
Wild Type ◽  
The Family ◽  
Dependent Inhibition ◽  
Root Gravitropism ◽  
Specialized Function ◽  
Mutant Lines

The TIR1/AFB auxin co-receptors mediate diverse responses to the plant hormone auxin. The Arabidopsis genome encodes six TIR1/AFB proteins representing three of the four clades that were established prior to angiosperm radiation. To determine the role of these proteins in plant development we performed an extensive genetic analysis involving the generation and characterization of all possible multiply-mutant lines. We find that loss of all six TIR1/AFB proteins results in early embryo defects and eventually seed abortion, and yet a single wild-type allele of TIR1 or AFB2 is sufficient to support growth throughout development. Our analysis reveals extensive functional overlap between even the most distantly related TIR1/AFB genes except for AFB1. Surprisingly, AFB1 has a specialized function in rapid auxin-dependent inhibition of root growth and early phase of root gravitropism. This activity may be related to a difference in subcellular localization compared to the other members of the family.

Effect of temperature on transition and transversion mutagenesis: characterization of wild type and a mutator T4 DNA polymerase mutant

1984 ◽  
Vol 26 (3) ◽  
pp. 386-389 ◽  
Author(s):  
Linda J. Reha-Krantz ◽  
Sükran Parmaksizoglu
Keyword(s):  
Dna Polymerase ◽  
Low Temperatures ◽  
Bacteriophage T4 ◽  
Effect Of Temperature ◽  
In Vitro Mutagenesis ◽  
Wild Type ◽  
Mutational Site

The effect of temperature on genetically well-defined mutational pathways was examined in the bacteriophage T4. The mutational site was a T4 rII ochre mutant which could revert to rII+ via a transversion or to the amber convertant via a transition. Temperature did not strongly affect any of the pathways examined in a wild-type background; however, increased temperature reduced the mutational activity of a mutator DNA polymerase mutant. Possible models to explain the role of temperature in mutagenesis are discussed as well as the significance of low temperatures for in vitro mutagenesis reactions.Key words: bacteriophage T4, mutator, transition, transversion, temperature effects.

The role of the flagellar transition region: inferences from the analysis of a Chlamydomonas mutant with defective transition region structures

1991 ◽  
Vol 99 (4) ◽  
pp. 731-740
Author(s):  
JONATHAN W. JARVIK ◽  
JOSEPH P. SUHAN
Keyword(s):  
Transition Region ◽  
Basal Body ◽  
Wild Type ◽  
Central Pair ◽  
Normal Transition ◽  
Thin Section Electron Microscopy ◽  
Concomitant Reduction ◽  
Section Electron ◽  
Type Variable

Thin-section electron microscopy of the Chlamydomonas reinhardtii mutant vfl-2 revealed striking defects in the transition region between basal body and flagellum. In place of the highly organized transition cylinders and stellate fibers characteristic of wild type, variable quantities of poorly organized electron-dense material were present. In many cases the transition region was penetrated by central pair microtubules that passed from the axoneme into the basal body. On the basis of these observations we propose that an important function of the structures present in the normal transition region is to physically exclude the central pair microtubules from the basal body. The transition region is the site of flagellar autotomy – the process by which doublet microtubules are severed and flagella are released from the cell. It has been claimed that autotomy is caused by contraction of the centrin-containing stellate fibers, resulting in the mechanical severing of the doublet microtubules and a concomitant reduction of the diameter of the axoneme adjacent to the abscission point. Our observations do not support this claim in that vfl-2 cells, which lack organized stellate fibers, display effective autotomy unaccompanied by detectable narrowing of the axoneme.

scholarly journals Characterization of Trypanosoma cruzi Sirtuins as Possible Drug Targets for Chagas Disease

2015 ◽  
Vol 59 (8) ◽  
pp. 4669-4679 ◽  
Author(s):  
Nilmar Silvio Moretti ◽  
Leonardo da Silva Augusto ◽  
Tatiana Mordente Clemente ◽  
Raysa Paes Pinto Antunes ◽  
Nobuko Yoshida ◽  
...  
Keyword(s):  
Trypanosoma Cruzi ◽  
Chagas Disease ◽  
Drug Targets ◽  
Wild Type ◽  
Content Type ◽  
Damage Repair ◽  
Lysine Deacetylases

ABSTRACTAcetylation of lysine is a major posttranslational modification of proteins and is catalyzed by lysine acetyltransferases, while lysine deacetylases remove acetyl groups. Among the deacetylases, the sirtuins are NAD+-dependent enzymes, which modulate gene silencing, DNA damage repair, and several metabolic processes. As sirtuin-specific inhibitors have been proposed as drugs for inhibiting the proliferation of tumor cells, in this study, we investigated the role of these inhibitors in the growth and differentiation ofTrypanosoma cruzi, the agent of Chagas disease. We found that the use of salermide during parasite infection prevented growth and initial multiplication after mammalian cell invasion byT. cruziat concentrations that did not affect host cell viability. In addition,in vivoinfection was partially controlled upon administration of salermide. There are two sirtuins inT. cruzi, TcSir2rp1 and TcSir2rp3. By using specific antibodies and cell lines overexpressing the tagged versions of these enzymes, we found that TcSir2rp1 is localized in the cytosol and TcSir2rp3 in the mitochondrion. TcSir2rp1 overexpression acts to impair parasite growth and differentiation, whereas the wild-type version of TcSir2rp3 and not an enzyme mutated in the active site improves both. The effects observed with TcSir2rp3 were fully reverted by adding salermide, which inhibited TcSir2rp3 expressed inEscherichia coliwith a 50% inhibitory concentration (IC50) ± standard error of 1 ± 0.5 μM. We concluded that sirtuin inhibitors targeting TcSir2rp3 could be used in Chagas disease chemotherapy.

scholarly journals Characterization of Biofilm Formation and the Role of BCR1 in Clinical Isolates of Candida parapsilosis

2013 ◽  
Vol 13 (4) ◽  
pp. 438-451 ◽  
Author(s):  
Srisuda Pannanusorn ◽  
Bernardo Ramírez-Zavala ◽  
Heinrich Lünsdorf ◽  
Birgitta Agerberth ◽  
Joachim Morschhäuser ◽  
...  
Keyword(s):  
Biofilm Formation ◽  
Candida Parapsilosis ◽  
Clinical Isolates ◽  
Wild Type ◽  
Content Type ◽  
Initial Adhesion ◽  
High Level ◽  
Increased Susceptibility

ABSTRACT In Candida parapsilosis , biofilm formation is considered to be a major virulence factor. Previously, we determined the ability of 33 clinical isolates causing bloodstream infection to form biofilms and identified three distinct groups of biofilm-forming strains (negative, low, and high). Here, we establish two different biofilm structures among strains forming large amounts of biofilm in which strains with complex spider-like structures formed robust biofilms on different surface materials with increased resistance to fluconazole. Surprisingly, the transcription factor Bcr1, required for biofilm formation in Candida albicans and C. parapsilosis , has an essential role only in strains with low capacity for biofilm formation. Although BCR1 leads to the formation of more and longer pseudohyphae, it was not required for initial adhesion and formation of mature biofilms in strains with a high level of biofilm formation. Furthermore, an additional phenotype affected by BCR1 was the switch in colony morphology from rough to crepe, but only in strains forming high levels of biofilm. All bcr1 Δ/Δ mutants showed increased proteolytic activity and increased susceptibility to the antimicrobial peptides protamine and RP-1 compared to corresponding wild-type and complemented strains. Taken together, our results demonstrate that biofilm formation in clinical isolates of C. parapsilosis is both dependent and independent of BCR1 , but even in strains which showed a BCR1 -independent biofilm phenotype, BCR1 has alternative physiological functions.

scholarly journals Characterization of the Opp Peptide Transporter ofCorynebacterium pseudotuberculosisand Its Role in Virulence and Pathogenicity

2014 ◽  
Vol 2014 ◽  
pp. 1-7 ◽  
Author(s):  
Pablo M. R. O. Moraes ◽  
Nubia Seyffert ◽  
Wanderson M. Silva ◽  
Thiago L. P. Castro ◽  
Renata F. Silva ◽  
...  
Keyword(s):  
Pathogenic Bacteria ◽  
Etiologic Agent ◽  
Peptide Transporter ◽  
Intercellular Signaling ◽  
Wild Type ◽  
Cell Nutrition ◽  
Oligopeptide Permease ◽  
Extracellular Environment

Despite the economic importance of caseous lymphadenitis (CLA), a chronic disease caused byCorynebacterium pseudotuberculosis, few genes related to the virulence of its etiologic agent have been characterized. The oligopeptide permease (Opp) transporters are located in the plasma membrane and have functions generally related to the uptake of peptides from the extracellular environment. These peptide transporters, in addition to having an important role in cell nutrition, also participate in the regulation of various processes involving intercellular signaling, including the control of the expression of virulence genes in pathogenic bacteria. To study the role of Opp inC. pseudotuberculosis, an OppD deficient strain was constructed via simple crossover with a nonreplicative plasmid carrying part of theoppDgene sequence. As occurred to the wild-type, the ΔoppDstrain showed impaired growth when exposed to the toxic glutathione peptide (GSH), indicating two possible scenarios: (i) that this component can be internalized by the bacterium through an Opp-independent pathway or (ii) that there is toxicity while the peptide is extracellular. Additionally, the ΔoppDmutant presented a reduced ability to adhere to and infect macrophages compared to the wild-type, although both strains exhibit the same potential to colonize spleens and cause injury and death to infected mice.

Sign in / Sign up

Export Citation Format

Share Document