An Extracellular Glycoprotein Is Implicated in Cell-Cell Contacts in the Toxic Cyanobacterium Microcystis aeruginosa PCC 7806

ABSTRACT Microcystins are the most common cyanobacterial toxins found in freshwater lakes and reservoirs throughout the world. They are frequently produced by the unicellular, colonial cyanobacterium Microcystis aeruginosa; however, the role of the peptide for the producing organism is poorly understood. Differences in the cellular aggregation of M. aeruginosa PCC 7806 and a microcystin-deficient ΔmcyB mutant guided the discovery of a surface-exposed protein that shows increased abundance in PCC 7806 mutants deficient in microcystin production compared to the abundance of this protein in the wild type. Mass spectrometric and immunoblot analyses revealed that the protein, designated microcystin-related protein C (MrpC), is posttranslationally glycosylated, suggesting that it may be a potential target of a putative O-glycosyltransferase of the SPINDLY family encoded downstream of the mrpC gene. Immunofluorescence microscopy detected MrpC at the cell surface, suggesting an involvement of the protein in cellular interactions in strain PCC 7806. Further analyses of field samples of Microcystis demonstrated a strain-specific occurrence of MrpC possibly associated with distinct Microcystis colony types. Our results support the implication of microcystin in the colony specificity of and colony formation by Microcystis.

Download Full-text

Transcriptome Changes in Pseudomonas putida KT2440 during Medium-Chain-Length Polyhydroxyalkanoate Synthesis Induced by Nitrogen Limitation

International Journal of Molecular Sciences ◽

10.3390/ijms22010152 ◽

2020 ◽

Vol 22 (1) ◽

pp. 152

Author(s):

Dorota Dabrowska ◽

Justyna Mozejko-Ciesielska ◽

Tomasz Pokój ◽

Slawomir Ciesielski

Keyword(s):

Chain Length ◽

Nitrogen Limitation ◽

Stringent Response ◽

Wild Type ◽

Pseudomonas Putida Kt2440 ◽

Medium Chain ◽

Environmental Stimuli ◽

Medium Chain Length ◽

In The Wild

Pseudomonas putida’s versatility and metabolic flexibility make it an ideal biotechnological platform for producing valuable chemicals, such as medium-chain-length polyhydroxyalkanoates (mcl-PHAs), which are considered the next generation bioplastics. This bacterium responds to environmental stimuli by rearranging its metabolism to improve its fitness and increase its chances of survival in harsh environments. Mcl-PHAs play an important role in central metabolism, serving as a reservoir of carbon and energy. Due to the complexity of mcl-PHAs’ metabolism, the manner in which P. putida changes its transcriptome to favor mcl-PHA synthesis in response to environmental stimuli remains unclear. Therefore, our objective was to investigate how the P. putida KT2440 wild type and mutants adjust their transcriptomes to synthesize mcl-PHAs in response to nitrogen limitation when supplied with sodium gluconate as an external carbon source. We found that, under nitrogen limitation, mcl-PHA accumulation is significantly lower in the mutant deficient in the stringent response than in the wild type or the rpoN mutant. Transcriptome analysis revealed that, under N-limiting conditions, 24 genes were downregulated and 21 were upregulated that were common to all three strains. Additionally, potential regulators of these genes were identified: the global anaerobic regulator (Anr, consisting of FnrA, Fnrb, and FnrC), NorR, NasT, the sigma54-dependent transcriptional regulator, and the dual component NtrB/NtrC regulator all appear to play important roles in transcriptome rearrangement under N-limiting conditions. The role of these regulators in mcl-PHA synthesis is discussed.

Download Full-text

Cytosolic Phospholipase A2α–deficient Mice Are Resistant to Collagen-induced Arthritis

Journal of Experimental Medicine ◽

10.1084/jem.20030016 ◽

2003 ◽

Vol 197 (10) ◽

pp. 1297-1302 ◽

Cited By ~ 105

Author(s):

Martin Hegen ◽

Linhong Sun ◽

Naonori Uozumi ◽

Kazuhiko Kume ◽

Mary E. Goad ◽

...

Keyword(s):

Critical Role ◽

Wild Type ◽

Collagen Induced Arthritis ◽

Cytosolic Phospholipase ◽

Severity Scores ◽

In The Wild ◽

Cytosolic Phospholipase A2α ◽

Antibody Levels ◽

Deficient Mice

Pathogenic mechanisms relevant to rheumatoid arthritis occur in the mouse model of collagen-induced arthritis (CIA). Cytosolic phospholipase A2α (cPLA2α) releases arachidonic acid from cell membranes to initiate the production of prostaglandins and leukotrienes. These inflammatory mediators have been implicated in the development of CIA. To test the hypothesis that cPLA2α plays a key role in the development of CIA, we backcrossed cPLA2α-deficient mice on the DBA/1LacJ background that is susceptible to CIA. The disease severity scores and the incidence of disease were markedly reduced in cPLA2α-deficient mice compared with wild-type littermates. At completion of the study, >90% of the wild-type mice had developed disease whereas none of the cPLA2α-deficient mice had more than one digit inflamed. Furthermore, visual disease scores correlated with severity of disease determined histologically. Pannus formation, articular fibrillation, and ankylosis were all dramatically reduced in the cPLA2α-deficient mice. Although the disease scores differed significantly between cPLA2α mutant and wild-type mice, anti-collagen antibody levels were similar in the wild-type mice and mutant littermates. These data demonstrate the critical role of cPLA2α in the pathogenesis of CIA.

Download Full-text

The development of the sensory neuron pattern in the antennal disc of wild-type and mutant (lz3, ssa) Drosophila melanogaster

Development ◽

10.1242/dev.112.4.1063 ◽

1991 ◽

Vol 112 (4) ◽

pp. 1063-1075

Author(s):

M.C. Lienhard ◽

R.F. Stocker

Keyword(s):

Drosophila Melanogaster ◽

Sensory Neuron ◽

Antennal Segment ◽

Wild Type ◽

Homeotic Mutant ◽

In The Wild ◽

The Brain ◽

Antennal Disc ◽

Antennal Nerve

The development of the sensory neuron pattern in the antennal disc of Drosophila melanogaster was studied with a neuron-specific monoclonal antibody (22C10). In the wild type, the earliest neurons become visible 3 h after pupariation, much later than in other imaginal discs. They lie in the center of the disc and correspond to the neurons of the adult aristal sensillum. Their axons join the larval antennal nerve and seem to establish the first connection towards the brain. Later on, three clusters of neurons appear in the periphery of the disc. Two of them most likely give rise to the Johnston's organ in the second antennal segment. Neurons of the olfactory third antennal segment are formed only after eversion of the antennal disc (clusters t1-t3). The adult pattern of antennal neurons is established at about 27% of metamorphosis. In the mutant lozenge3 (lz3), which lacks basiconic antennal sensilla, cluster t3 fails to develop. This indicates that, in the wild type, a homogeneous group of basiconic sensilla is formed by cluster t3. The possible role of the lozenge gene in sensillar determination is discussed. The homeotic mutant spineless-aristapedia (ssa) transforms the arista into a leg-like tarsus. Unlike leg discs, neurons are missing in the larval antennal disc of ssa. However, the first neurons differentiate earlier than in normal antennal discs. Despite these changes, the pattern of afferents in the ectopic tarsus appears leg specific, whereas in the non-transformed antennal segments a normal antennal pattern is formed. This suggests that neither larval leg neurons nor early aristal neurons are essential for the outgrowth of subsequent afferents.

Download Full-text

Pax6 regulates specification of ventral neurone subtypes in the hindbrain by establishing progenitor domains

Development ◽

10.1242/dev.129.6.1327 ◽

2002 ◽

Vol 129 (6) ◽

pp. 1327-1338 ◽

Cited By ~ 1

Author(s):

Masanori Takahashi ◽

Noriko Osumi

Keyword(s):

Expression Patterns ◽

Domain Formation ◽

Wild Type ◽

Whole Embryo Culture ◽

Neuronal Markers ◽

Signalling Molecules ◽

Neuronal Progenitors ◽

In The Wild ◽

Mutant Rat

Recent studies have shown that generation of different kinds of neurones is controlled by combinatorial actions of homeodomain (HD) proteins expressed in the neuronal progenitors. Pax6 is a HD protein that has previously been shown to be involved in the differentiation of the hindbrain somatic (SM) motoneurones and V1 interneurones in the hindbrain and/or spinal cord. To investigate in greater depth the role of Pax6 in generation of the ventral neurones, we first examined the expression patterns of HD protein genes and subtype-specific neuronal markers in the hindbrain of the Pax6 homozygous mutant rat. We found that Islet2 (SM neurone marker) and En1 (V1 interneurone marker) were transiently expressed in a small number of cells, indicating that Pax6 is not directly required for specification of these neurones. We also observed that domains of all other HD protein genes (Nkx2.2, Nkx6.1, Irx3, Dbx2 and Dbx1) were shifted and their boundaries became blurred. Thus, Pax6 is required for establishment of the progenitor domains of the ventral neurones. Next, we performed Pax6 overexpression experiments by electroporating rat embryos in whole embryo culture. Pax6 overexpression in the wild type decreased expression of Nkx2.2, but ectopically increased expression of Irx3, Dbx1 and Dbx2. Moreover, electroporation of Pax6 into the Pax6 mutant hindbrain rescued the development of Islet2-positive and En1-positive neurones. To know reasons for perturbed progenitor domain formation in Pax6 mutant, we examined expression patterns of Shh signalling molecules and states of cell death and cell proliferation. Shh was similarly expressed in the floor plate of the mutant hindbrain, while the expressions of Ptc1, Gli1 and Gli2 were altered only in the progenitor domains for the motoneurones. The position and number of TUNEL-positive cells were unchanged in the Pax6 mutant. Although the proportion of cells that were BrdU-positive slightly increased in the mutant, there was no relationship with specific progenitor domains. Taken together, we conclude that Pax6 regulates specification of the ventral neurone subtypes by establishing the correct progenitor domains.

Download Full-text

A 13C-NMR study of the role of Asn-155 in stabilizing the oxyanion of a subtilisin tetrahedral adduct

Biochemical Journal ◽

10.1042/bj3260861 ◽

1997 ◽

Vol 326 (3) ◽

pp. 861-866 ◽

Cited By ~ 14

Author(s):

Timothy P. O'CONNELL ◽

Regina M. DAY ◽

Ekaterina V. TORCHILIN ◽

William W. BACHOVCHIN ◽

J. Paul G. MALTHOUSE

Keyword(s):

13C Nmr ◽

Chemical Shifts ◽

Catalytic Efficiency ◽

Site Directed Mutagenesis ◽

Wild Type ◽

Imidazolium Cation ◽

Nmr Study ◽

In The Wild ◽

Main Factor

By removing one of the hydrogen-bond donors in the oxyanion hole of subtilisin BPN, we have been able to determine how it affects the catalytic efficiency of the enzyme and the pKa of the oxyanion formed in a choloromethane inhibitor derivative. Variant 8397 of subtilisin BPN contains five mutations which enhance its stability. Site-directed mutagenesis was used to prepare the N155A mutant of this variant. The catalytic efficiencies of wild-type and variant 8397 are similar, but replacing Asn-155 with alanine reduces catalytic efficiency approx. 300-fold. All three forms of subtilisin were alkylated using benzyloxycarbonylglycylglycyl[2-13C]phenylalanylchloromethane and examined by 13C-NMR. A single signal due to the 13C-enriched carbon was detected in all the derivatives and it was assigned to the hemiketal carbon of a tetrahedral adduct formed between the hydroxy group of Ser-221 and the inhibitor. This signal had chemical shifts in the range 98.3–103.6 p.p.m., depending on the pH. The titration shift of 4.7–4.8 p.p.m. was assigned to oxyanion formation. The oxyanion pKa values in the wild-type and 8397 variants were 6.92 and 7.00 respectively. In the N155A mutant of the 8397 variant the oxyanion pKa increased to 8.09. We explain why such a small increase is observed and we conclude that it is the interaction between the oxyanion and the imidazolium cation of the active-site histidine that is the main factor responsible for lowering the oxyanion pKa.

Download Full-text

Indoleamine 2,3-Dioxygenase 2 Deficiency Exacerbates Imiquimod-Induced Psoriasis-Like Skin Inflammation

International Journal of Molecular Sciences ◽

10.3390/ijms21155515 ◽

2020 ◽

Vol 21 (15) ◽

pp. 5515

Author(s):

Kento Fujii ◽

Yasuko Yamamoto ◽

Yoko Mizutani ◽

Kuniaki Saito ◽

Mariko Seishima

Keyword(s):

Immune Responses ◽

Immune Cells ◽

Skin Inflammation ◽

Kynurenine Pathway ◽

Immune Functions ◽

Wild Type ◽

Indoleamine 2,3 Dioxygenase ◽

Tnf Α ◽

In The Wild

Indoleamine 2,3-dioxygenase 1 (IDO1) is an enzyme known to suppress immune responses, and several reports have showed that it is associated with psoriasis. IDO2 is an isoform of IDO1, recently identified as a catalytic enzyme in the tryptophan-kynurenine pathway, which is expressed in dendritic cells and monocytes. The expression of IDO2 in immune cells suggests that IDO2 may contribute to immune functions. However, the role of IDO2 in the pathogenesis of psoriasis remains unclear. In this study, to elucidate the role of IDO2 in psoriasis, we assessed imiquimod (IMQ)-induced psoriasis-like dermatitis in IDO2 knockout (KO) mice. Skin inflammation, evaluated by scoring erythema, scaling, and ear thickness, was significantly worse in the IDO2 KO mice than in the wild-type (WT) mice. The mRNA expression levels of TNF-α, IL-23p19, and IL-17A, key cytokines involved in the development of psoriasis, were also increased in the IDO2 KO mice. Furthermore, immunohistochemistry revealed that the number of Ki67-positive cells in the epidermis and CD4-, CD8-, and IL-17-positive lymphocytes infiltrating the dermis were significantly increased in the IDO2 KO mice. These results suggest that IDO2 might decrease IL-17 expression, thereby resulting in the suppression of skin inflammation in IMQ-induced psoriasis-like dermatitis.

Download Full-text

Colonization and Inflammation Deficiencies in Mongolian Gerbils Infected by Helicobacter pylori Chemotaxis Mutants

Infection and Immunity ◽

10.1128/iai.73.3.1820-1827.2005 ◽

2005 ◽

Vol 73 (3) ◽

pp. 1820-1827 ◽

Cited By ~ 78

Author(s):

David J. McGee ◽

Melanie L. Langford ◽

Emily L. Watson ◽

J. Elliot Carter ◽

Yu-Ting Chen ◽

...

Keyword(s):

Helicobacter Pylori ◽

Immune Cell ◽

Response Regulator ◽

Critical Role ◽

Mongolian Gerbils ◽

Wild Type ◽

In The Wild ◽

H Pylori

ABSTRACT Helicobacter pylori causes disease in the human stomach and in mouse and gerbil stomach models. Previous results have shown that motility is critical for H. pylori to colonize mice, gerbils, and other animal models. The role of chemotaxis, however, in colonization and disease is less well understood. Two genes in the H. pylori chemotaxis pathway, cheY and tlpB, which encode the chemotaxis response regulator and a methyl-accepting chemoreceptor, respectively, were disrupted. The cheY mutation was complemented with a wild-type copy of cheY inserted into the chromosomal rdxA gene. The cheY mutant lost chemotaxis but retained motility, while all other strains were motile and chemotactic in vitro. These strains were inoculated into gerbils either alone or in combination with the wild-type strain, and colonization and inflammation were assessed. While the cheY mutant completely failed to colonize gerbil stomachs, the tlpB mutant colonized at levels similar to those of the wild type. With the tlpB mutant, there was a substantial decrease in inflammation in the gerbil stomach compared to that with the wild type. Furthermore, there were differences in the numbers of each immune cell in the tlpB-mutant-infected stomach: the ratio of lymphocytes to neutrophils was about 8 to 1 in the wild type but only about 1 to 1 in the mutant. These results suggest that the TlpB chemoreceptor plays an important role in the inflammatory response while the CheY chemotaxis regulator plays a critical role in initial colonization. Chemotaxis mutants may provide new insights into the steps involved in H. pylori pathogenesis.

Download Full-text

Biological role of the general control of amino acid biosynthesis in Saccharomyces cerevisiae.

Molecular and Cellular Biology ◽

10.1128/mcb.1.7.584 ◽

1981 ◽

Vol 1 (7) ◽

pp. 584-593 ◽

Cited By ~ 73

Author(s):

P Niederberger ◽

G Miozzari ◽

R Hütter

Keyword(s):

Saccharomyces Cerevisiae ◽

Amino Acid ◽

Amino Acid Biosynthesis ◽

Wild Type ◽

General Control ◽

Acid Biosynthesis ◽

Mutant Strains ◽

In The Wild ◽

Amino Acid Imbalance

The biological role of the "general control of amino acid biosynthesis" has been investigated by analyzing growth and enzyme levels in wild-type, bradytrophic, and nonderepressing mutant strains of Saccharomyces cerevisiae. Amino acid limitation was achieved by using either bradytrophic mutations or external amino acid imbalance. In the wild-type strain noncoordinate derepression of enzymes subject to the general control has been found. Derepressing factors were in the order of 2 to 4 in bradytrophic mutant strains grown under limiting conditions and only in the order of 1.5 to 2 under the influence of external amino acid imbalance. Nonderepressing mutations led to slower growth rates under conditions of amino acid limitation, and no derepression of enzymes under the general control was observed. The amino acid pools were found to be very similar in the wild type and in nonderepressing mutant strains under all conditions tested. Our results indicate that the general control affects all branched amino acid biosynthetic pathways, namely, those of the aromatic amino acids and the aspartate family, the pathways for the basic amino acids lysine, histidine, and arginine, and also the pathways of serine and valine biosyntheses.

Download Full-text

The role of catalase in hydrogen peroxide resistance in fission yeast Schizosaccharomyces pombe

Canadian Journal of Microbiology ◽

10.1139/w98-216 ◽

1999 ◽

Vol 45 (2) ◽

pp. 125-129 ◽

Cited By ~ 8

Author(s):

Norihiro Mutoh ◽

Chiaki W Nakagawa ◽

Kenichiro Yamada

Keyword(s):

Hydrogen Peroxide ◽

Schizosaccharomyces Pombe ◽

Catalase Activity ◽

Parent Strain ◽

Normal Growth ◽

Wild Type ◽

Catalase Gene ◽

In The Wild ◽

High Concentrations

The role of catalase in hydrogen peroxide resistance in Schizosaccharomyces pombe was investigated. A catalase gene disruptant completely lacking catalase activity is more sensitive to hydrogen peroxide than the parent strain. The mutant does not acquire hydrogen peroxide resistance by osmotic stress, a treatment that induces catalase activity in the wild-type cells. The growth rate of the disruptant is not different from that of the parent strain. Additionally, transformed cells that overexpress the catalase activity are more resistant to hydrogen peroxide than wild-type cells with normal catalase activity. These results indicate that the catalase of S. pombe plays an important role in resistance to high concentrations of hydrogen peroxide but offers little in the way of protection from the hydrogen peroxide generated in small amounts under normal growth conditions.Key words: catalase, gene disruption, induced hydrogen peroxide resistance, overexpression, Schizosaccharomyces pombe.

Download Full-text

Altered cytokinin distribution in the shootless maize mutant ed*41

Functional Plant Biology ◽

10.1071/pp00108 ◽

2001 ◽

Vol 28 (4) ◽

pp. 307 ◽

Cited By ~ 1

Author(s):

Adriana Chiappetta ◽

Walter de Witte ◽

Milvia L. Racchi ◽

Maria B. Bitonti ◽

Henri van Onckelen ◽

...

Keyword(s):

Zea Mays ◽

Apical Meristem ◽

Time Course ◽

Zea Mays L ◽

Wild Type ◽

Vascular Connection ◽

Vascular Tissues ◽

In The Wild ◽

Specific Synthesis

The distribution of cytokinins is strongly altered in seedlings of the shootless ed*41 mutant of maize (Zea mays L.), compared with wild-type. Immunolocalisation of zeatin and analysis of cytokinin levels clearly indicate that these hormones are not present in the shoot apex zone of the mutant. Since an anomalous differentiation of vascular tissues has also been observed in the mutant, a major role of vascular connection in hormone translocation affecting development of the shoot apical meristem is proposed. Immunolocalisation of zeatin was confined to the root cap, cortex and vascular tissues of both mutant and wild-type seedlings suggesting a tissue-specific synthesis of this hormone in the root. A time-course of cytokinin distribution in the wild-type developing shoot provided evidence that the tunica and corpus zones become competent to respond to cytokinins in subsequent periods, very probably in conjunction with photomorphogenesis. On the contrary, this pathway is totally disrupted in the mutant. Taken together, the data point to a relationship between the ed*41 mutation, inadequate vascular connection and disrupted hormone translocation.

Download Full-text