bluuebunny/crossref_metadata_2025_split huggingface dataset:
Lactate and Amino Acid Catabolism in the Cheese-Ripening Yeast
Yarrowia lipolytica
Authors: Mansour S. , Beckerich J. M. , Bonnarme P. 
Published: September 2008

ABSTRACT

The consumption of lactate and amino acids is very important for microbial development and/or aroma production during cheese ripening. A strain of
Yarrowia lipolytica
isolated from cheese was grown in a liquid medium containing lactate in the presence of a low (0.1×) or high (2×) concentration of amino acids. Our results show that there was a dramatic increase in the growth of
Y. lipolytica
in the medium containing a high amino acid concentration, but there was limited lactate consumption. Conversely, lactate was efficiently consumed in the medium containing a low concentration of amino acids after amino acid depletion was complete. These data suggest that the amino acids are used by
Y. lipolytica
as a main energy source, whereas lactate is consumed following amino acid depletion. Amino acid degradation was accompanied by ammonia production corresponding to a dramatic increase in the pH. The effect of adding amino acids to a
Y. lipolytica
culture grown on lactate was also investigated. Real-time quantitative PCR analyses were performed with specific primers for five genes involved in amino acid transport and catabolism, including an amino acid transporter gene (
GAP1
) and four aminotransferase genes (
ARO8
,
ARO9
,
BAT1
, and
BAT2
). The expression of three genes involved in lactate transport and catabolism was also studied. These genes included a lactate transporter gene (
JEN1
) and two lactate dehydrogenase genes (
CYB2
-
1
and
CYB2
-
2
). Our data showed that
GAP1
,
BAT2
,
BAT1
, and
ARO8
were maximally expressed after 15 to 30 min following addition of amino acids (
BAT2
was the most highly expressed gene), while the maximum expression of
JEN1
,
CYB2
-
1
, and
CYB2
-
2
was delayed (≥60 min).
Evidence for the Role of Calcineurin in Morphogenesis and Calcium Homeostasis during Mycelium-to-Yeast Dimorphism of
Paracoccidioides brasiliensis
Authors: Campos Claudia B. L. , Di Benedette Joao Paulo T. , Morais Flavia V. , Ovalle Rafael , Nobrega Marina P. 
Published: September 2008

ABSTRACT

Paracoccidioides brasiliensis
is a dimorphic fungus that causes paracoccidioidomycosis, the most prevalent human deep mycosis in Latin America. The dimorphic transition from mycelium to yeast (M-Y) is triggered by a temperature shift from 25°C to 37°C and is critical for pathogenicity. Intracellular Ca
2+
levels increased in hyphae immediately after temperature-induced dimorphism. The chelation of Ca
2+
with extracellular (EGTA) or intracellular (BAPTA) calcium chelators inhibited temperature-induced dimorphism, whereas the addition of extracellular Ca
2+
accelerated dimorphism. The calcineurin inhibitor cyclosporine A (CsA), but not tacrolimus (FK506), effectively decreased cell growth, halted the M-Y transition that is associated with virulence, and caused aberrant growth morphologies for all forms of
P. brasiliensis
. The difference between CsA and FK506 was ascribed by the higher levels of cyclophilins contrasted to FKBPs, the intracellular drug targets required for calcineurin suppression. Chronic exposure to CsA abolished intracellular Ca
2+
homeostasis and decreased mRNA transcription of the
CCH1
gene for the plasma membrane Ca
2+
channel in yeast-form cells. CsA had no detectable effect on multidrug resistance efflux pumps, while the effect of FK506 on rhodamine excretion was not correlated with the transition to yeast form. In this study, we present evidence that Ca
2+
/calmodulin-dependent phosphatase calcineurin controls hyphal and yeast morphology, M-Y dimorphism, growth, and Ca
2+
homeostasis in
P. brasiliensis
and that CsA is an effective chemical block for thermodimorphism in this organism. The effects of calcineurin inhibitors on
P. brasiliensis
reinforce the therapeutic potential of these drugs in a combinatory approach with antifungal drugs to treat endemic paracoccidioidomycosis.
Structural and Functional Association of
Trypanosoma brucei
MIX Protein with Cytochrome
c
Oxidase Complex
Authors: Zíková Alena , Panigrahi Aswini K. , Uboldi Alessandro D. , Dalley Rachel A. , Handman Emanuela , Stuart Kenneth 
Published: September 2008

ABSTRACT

A mitochondrial inner membrane protein, designated MIX, seems to be essential for cell viability. The deletion of both alleles was not possible, and the deletion of a single allele led to a loss of virulence and aberrant mitochondrial segregation and cell division in
Leishmania major
. However, the mechanism by which MIX exerts its effect has not been determined. We show here that MIX is also expressed in the mitochondrion of
Trypanosoma brucei
, and using RNA interference, we found that its loss leads to a phenotype that is similar to that described for
Leishmania
. The loss of MIX also had a major effect on cytochrome
c
oxidase activity, on the mitochondrial membrane potential, and on the production of mitochondrial ATP by oxidative phosphorylation. Using a tandem affinity purification tag, we found that MIX is associated with a multiprotein complex that contains subunits of the mitochondrial cytochrome
c
oxidase complex (respiratory complex IV), the composition of which was characterized in detail. The specific function of MIX is unknown, but it appears to be important for the function of complex IV and for mitochondrial segregation and cell division in
T. brucei
.
A Signaling-Regulated, Short-Chain Dehydrogenase of
Stagonospora nodorum
Regulates Asexual Development
Authors: Tan Kar-Chun , Heazlewood Joshua L. , Millar A. Harvey , Thomson Gordon , Oliver Richard P. , Solomon Peter S. 
Published: September 2008

ABSTRACT

The fungus
Stagonospora nodorum
is a causal agent of leaf and glume blotch disease of wheat. It has been previously shown that inactivation of heterotrimeric G protein signaling in
Stagonospora nodorum
caused development defects and reduced pathogenicity [P. S. Solomon et al., Mol. Plant-Microbe Interact. 17:456-466, 2004]. In this study, we sought to identify targets of the signaling pathway that may have contributed to phenotypic defects of the signaling mutants. A comparative analysis of
Stagonospora nodorum
wild-type and Gα-defective mutant (
gna1
) intracellular proteomes was performed via two-dimensional polyacrylamide gel electrophoresis. Several proteins showed significantly altered abundances when comparing the two strains. One such protein, the short-chain dehydrogenase Sch1, was 18-fold less abundant in the
gna1
strain, implying that it is positively regulated by Gα signaling. Gene expression and transcriptional enhanced green fluorescent protein fusion analyses of
Sch1
indicates strong expression during asexual development. Mutant strains of
Stagonospora nodorum
lacking
Sch1
demonstrated poor growth on minimal media and exhibited a significant reduction in asexual sporulation on all growth media examined. Detailed histological experiments on
sch1
pycnidia revealed that the gene is required for the differentiation of the subparietal layers of asexual pycnidia resulting in a significant reduction in both pycnidiospore size and numbers.
Cytological Characterization of YpsB, a Novel Component of the
Bacillus subtilis
Divisome
Authors: Tavares José Roberto , de Souza Robson F. , Meira Guilherme Louzada Silva , Gueiros-Filho Frederico J. 
Published: September 2008

ABSTRACT

Cell division in bacteria is carried out by an elaborate molecular machine composed of more than a dozen proteins and known as the divisome. Here we describe the characterization of a new divisome protein in
Bacillus subtilis
called YpsB. Sequence comparisons and phylogentic analysis demonstrated that YpsB is a paralog of the division site selection protein DivIVA. YpsB is present in several gram-positive bacteria and likely originated from the duplication of a DivIVA-like gene in the last common ancestor of bacteria of the orders
Bacillales
and
Lactobacillales
. We used green fluorescent protein microscopy to determine that YpsB localizes to the divisome. Similarly to that for DivIVA, the recruitment of YpsB to the divisome requires late division proteins and occurs significantly after Z-ring formation. In contrast to DivIVA, however, YpsB is not retained at the newly formed cell poles after septation. Deletion analysis suggests that the N terminus of YpsB is required to target the protein to the divisome. The high similarity between the N termini of YpsB and DivIVA suggests that the same region is involved in the targeting of DivIVA. YpsB is not essential for septum formation and does not appear to play a role in septum positioning. However, a
ypsB
deletion has a synthetic effect when combined with a mutation in the cell division gene
ftsA
. Thus, we conclude that YpsB is a novel
B. subtilis
cell division protein whose function has diverged from that of its paralog DivIVA.
The Chromate-Inducible
chrBACF
Operon from the Transposable Element Tn
OtChr
Confers Resistance to Chromium(VI) and Superoxide
Authors: Branco Rita , Chung Ana Paula , Johnston Tatiana , Gurel Volkan , Morais Paula , Zhitkovich Anatoly 
Published: September 2008

ABSTRACT

Large-scale industrial use of chromium(VI) has resulted in widespread contamination with carcinogenic chromium(VI). The abilities of microorganisms to survive in these environments and to detoxify chromate require the presence of specific resistance systems. Here we report identification of the transposon-located (Tn
OtChr
) chromate resistance genes from the highly tolerant strain
Ochrobactrum tritici
5bvl1 surviving chromate concentrations of >50 mM. The 7,189-bp-long Tn
OtChr
of the mixed Tn
21
/Tn
3
transposon subfamily contains a group of
chrB
,
chrA
,
chrC
, and
chrF
genes situated between divergently transcribed resolvase and transposase genes. The
chrB
and
chrA
genes, but not
chrF
or
chrC
, were essential for establishment of high resistance in chromium-sensitive
O. tritici
. The
chr
promoter was strongly induced by chromate or dichromate, but it was completely unresponsive to Cr(III), oxidants, sulfate, or other oxyanions. Plasmid reporter experiments identified ChrB as a chromate-sensing regulator of
chr
expression. Induction of the
chr
operon suppressed accumulation of cellular Cr through the activity of a chromate efflux pump encoded by
chrA
. Expression of
chrB
,
chrC
, or
chrF
in an
Escherichia coli sodA sodB
double mutant restored its aerobic growth in minimal medium and conferred resistance to superoxide-generating agents menadione and paraquat. Nitroblue tetrazolium staining on native gels showed that ChrC protein had superoxide dismutase activity. Tn
OtChr
appears to represent a mobile genetic system for the distribution of the chromate-regulated resistance operon. The presence of three genes protecting against superoxide toxicity should provide an additional survival advantage to Tn
OtChr
-containing cells in the environments with multiple redox-active contaminants.
A Functional Phenylacetic Acid Catabolic Pathway Is Required for Full Pathogenicity of
Burkholderia cenocepacia
in the
Caenorhabditis elegans
Host Model
Authors: Law Robyn J. , Hamlin Jason N. R. , Sivro Aida , McCorrister Stuart J. , Cardama Georgina A. , Cardona Silvia T. 
Published: September 2008

ABSTRACT

Burkholderia cenocepacia
is a member of the
Burkholderia cepacia
complex, a group of metabolically versatile bacteria that have emerged as opportunistic pathogens in cystic fibrosis and immunocompromised patients. Previously a screen of transposon mutants in a rat pulmonary infection model identified an attenuated mutant with an insertion in
paaE
, a gene related to the phenylacetic acid (PA) catabolic pathway. In this study, we characterized gene clusters involved in the PA degradation pathway of
B. cenocepacia
K56-2 in relation to its pathogenicity in the
Caenorhabditis elegans
model of infection. We demonstrated that targeted-insertion mutagenesis of
paaA
and
paaE
, which encode part of the putative PA-coenzyme A (CoA) ring hydroxylation system,
paaZ
, coding for a putative ring opening enzyme, and
paaF
, encoding part of the putative beta-oxidation system, severely reduces growth on PA as a sole carbon source.
paaA
and
paaE
insertional mutants were attenuated for virulence, and expression of
paaE
in
trans
restored pathogenicity of the
paaE
mutant to wild-type levels. Interruption of
paaZ
and
paaF
slightly increased virulence. Using gene interference by ingested double-stranded RNA, we showed that the attenuated phenotype of the
paaA
and
paaE
mutants is dependent on a functional p38 mitogen-activated protein kinase pathway in
C. elegans
. Taken together, our results demonstrate that
B. cenocepacia
possesses a functional PA degradation pathway and that the putative PA-CoA ring hydroxylation system is required for full pathogenicity in
C. elegans
.
The Bile Response Repressor BreR Regulates Expression of the
Vibrio cholerae breAB
Efflux System Operon
Authors: Cerda-Maira Francisca A. , Ringelberg Carol S. , Taylor Ronald K. 
Published: September 2008

ABSTRACT

Enteric pathogens have developed several resistance mechanisms to survive the antimicrobial action of bile. We investigated the transcriptional profile of
Vibrio cholerae
O1 El Tor strain C6706 under virulence gene-inducing conditions in the presence and absence of bile. Microarray analysis revealed that the expression of 119 genes was affected by bile. The mRNA levels of genes encoding proteins involved in transport were increased in the presence of bile, whereas the mRNA levels of genes encoding proteins involved in pathogenesis and chemotaxis were decreased. This study identified genes encoding transcriptional regulators from the TetR family (
vexR
and
breR
) and multidrug efflux pumps from the resistance-nodulation-cell division superfamily (
vexB
and
vexD
[herein renamed
breB
]) that were induced in response to bile. Further analysis regarding
vexAB
and
breAB
expression in the presence of various antimicrobial compounds established that
vexAB
was induced in the presence of bile, sodium dodecyl sulfate, or novobiocin and that the induction of
breAB
was specific to bile. BreR is a direct repressor of the
breAB
promoter and is able to regulate its own expression, as demonstrated by transcriptional and electrophoretic mobility shift assays (EMSA). The expression of
breR
and
breAB
is induced in the presence of the bile salts cholate, deoxycholate, and chenodeoxycholate, and EMSA showed that deoxycholate is able to abolish the formation of BreR-P

breR

complexes. We propose that deoxycholate is able to interact with BreR and induce a conformational change that interferes with the DNA binding ability of BreR, resulting in
breAB
and
breR
expression. These results provide new insight into a transcriptional regulator and a transport system that likely play essential roles in the ability of
V. cholerae
to resist the action of bile in the host.
SpoT-Triggered Stringent Response Controls
usp
Gene Expression in
Pseudomonas aeruginosa
Authors: Boes Nelli , Schreiber Kerstin , Schobert Max 
Published: September 2008

ABSTRACT

The universal stress proteins (Usps) UspK (PA3309) and UspN (PA4352) of
Pseudomonas aeruginosa
are essential for surviving specific anaerobic energy stress conditions such as pyruvate fermentation and anaerobic stationary phase. Expression of the respective genes is under the control of the oxygen-sensing regulator Anr. In this study we investigated the regulation of
uspN
and three additional
P. aeruginosa usp
genes:
uspL
(PA1789),
uspM
(PA4328), and
uspO
(PA5027). Anr induces expression of these genes in response to anaerobic conditions. Using promoter-
lacZ
fusions, we showed that P

uspL

-
lacZ
, P

uspM

-
lacZ
, and P

uspO

-
lacZ
were also induced in stationary phase as described for P

uspN

-
lacZ
. However, stationary phase gene expression was abolished in the
P. aeruginosa
triple mutant Δ
anr
Δ
relA
Δ
spoT
. The
relA
and
spoT
genes encode the regulatory components of the stringent response. We determined pppGpp and ppGpp levels using a thin-layer chromatography approach and detected the accumulation of ppGpp in the wild type and the Δ
relA
mutant in stationary phase, indicating a SpoT-derived control of ppGpp accumulation. Additional investigation of stationary phase in LB medium revealed that alkaline pH values are involved in the regulatory process of ppGpp accumulation.
Lsr2 of
Mycobacterium
Represents a Novel Class of H-NS-Like Proteins
Authors: Gordon Blair R. G. , Imperial Robin , Wang Linru , Navarre William Wiley , Liu Jun 
Published: September 2008

ABSTRACT

Lsr2 is a small, basic protein present in
Mycobacterium
and related actinomycetes. Our previous in vitro biochemical studies showed that Lsr2 is a DNA-bridging protein, a property shared by H-NS-like proteins in gram-negative bacteria. Here we present in vivo evidence based on genetic complementation experiments that Lsr2 is a functional analog of H-NS, the first such protein identified in gram-positive bacteria. We show that
lsr2
can complement the phenotypes related to
hns
mutations in
Escherichia coli
, including β-glucoside utilization, mucoidy, motility, and hemolytic activity. We also show that Lsr2 binds specifically to H-NS-regulated genes and the repression of
hlyE
by Lsr2 can be partially eliminated by overexpression of
slyA
, suggesting that the molecular mechanisms of Lsr2 repression and depression are similar to those of H-NS. The functional equivalence of these two proteins is further supported by the ability of
hns
to complement the
lsr2
phenotype in
Mycobacterium smegmatis
. Taken together, our results demonstrate unequivocally that Lsr2 is an H-NS-like protein.
PqsE Functions Independently of PqsR-PseudomonasQuinolone Signal and Enhances therhlQuorum-Sensing System
Authors: Farrow John M. , Sund Zoe M. , Ellison Matthew L. , Wade Dana S. , Coleman James P. , Pesci Everett C. 
Published: September 2008

ABSTRACTPseudomonas aeruginosais an opportunistic pathogen that causes both acute and chronic infections in immunocompromised individuals. This gram-negative bacterium produces a battery of virulence factors that allow it to infect and survive in many different hostile environments. The control of many of these virulence factors falls under the influence of one of threeP. aeruginosacell-to-cell signaling systems. The focus of this study, the quinolone signaling system, functions through thePseudomonasquinolone signal (PQS), previously identified as 2-heptyl-3-hydroxy-4-quinolone. This signal binds to and activates the LysR-type transcriptional regulator PqsR (also known as MvfR), which in turn induces the expression of thepqsABCDEoperon. The first four genes of this operon are required for PQS synthesis, but the fifth gene,pqsE, is not. The function of thepqsEgene is not known, but it is required for the production of multiple PQS-controlled virulence factors and for virulence in multiple models of infection. In this report, we show that PqsE can activate PQS-controlled genes in the absence of PqsR and PQS. Our data also suggest that the regulatory activity of PqsE requires RhlR and indicate that apqsEmutant can be complemented for pyocyanin production by a large excess of exogenousN-butyryl homoserine lactone (C4-HSL). Finally, we show that PqsE enhances the ability ofEscherichia coliexpressing RhlR to respond to C4-HSL. Overall, our data lead us to conclude that PqsE functions as a regulator that is independent of PqsR and PQS but dependent on therhlquorum-sensing system.
Dual Role of FtsH in Regulating Lipopolysaccharide Biosynthesis in
Escherichia coli
Authors: Katz Chen , Ron Eliora Z. 
Published: September 2008

ABSTRACT

In
Escherichia coli
, FtsH (HflB) is a membrane-bound, ATP-dependent metalloendoprotease belonging to the AAA family (ATPases associated with diverse cellular activities). FtsH has a limited spectrum of known substrates, including the transcriptional activator σ
32
. FtsH is the only known
E. coli
protease that is essential, as it regulates the concentration of LpxC, which carries out the first committed step in the synthesis of lipid A. Here we identify a new FtsH substrate—3-deoxy-
d
-
manno
-octulosonate (KDO) transferase—which carries out the attachment of two KDO residues to the lipid A precursor (lipid IV
A
) to form the minimal essential structure of the lipopolysaccharide (LPS) (KDO
2
-lipid A). Thus, FtsH regulates the concentration of the lipid moiety of LPS (lipid A) as well as the sugar moiety (KDO-based core oligosaccharides), ensuring a balanced synthesis of LPS.
A Novel Mutator of
Escherichia coli
Carrying a Defect in the
dgt
Gene, Encoding a dGTP Triphosphohydrolase
Authors: Gawel Damian , Hamilton Michael D. , Schaaper Roel M. 
Published: September 2008

ABSTRACT

A novel mutator locus in
Escherichia coli
was identified from a collection of random transposon insertion mutants. Several mutators in this collection were found to have an insertion in the
dgt
gene, encoding a previously characterized dGTP triphosphohydrolase. The mutator activity of the
dgt
mutants displays an unusual specificity. Among the six possible base pair substitutions in a
lacZ
reversion system, the G·C→C·G transversion and A·T→G·C transition are strongly enhanced (10- to 50-fold), while a modest effect (two- to threefold) is also observed for the G·C→A·T transition. Interestingly, a two- to threefold reduction in mutant frequency (antimutator effect) is observed for the G·C→T·A transversion. In the absence of DNA mismatch repair (
mutL
) some of these effects are reduced or abolished, while other effects remain unchanged. Analysis of these effects, combined with the DNA sequence contexts in which the reversions take place, suggests that alterations of the dGTP pools as well as alterations in the level of some modified dNTP derivatives could affect the fidelity of in vivo DNA replication and, hence, account for the overall mutator effects.
Novel Proteins That Modulate Type IV Pilus Retraction Dynamics in
Pseudomonas aeruginosa
Authors: Asikyan Miranda L. , Kus Julianne V. , Burrows Lori L. 
Published: September 2008

ABSTRACT

Pseudomonas aeruginosa
uses type IV pili to colonize various materials and for surface-associated twitching motility. We previously identified five phylogenetically distinct alleles of
pilA
in
P. aeruginosa
, four of which occur in genetic cassettes with specific accessory genes (J. V. Kus, E. Tullis, D. G. Cvitkovitch, and L. L. Burrows, Microbiology 150:1315-1326, 2004). Each of the five pilin alleles, with and without its associated pilin accessory gene, was used to complement a group II PAO1
pilA
mutant. Expression of group I or IV
pilA
genes restored twitching motility to the same extent as the PAO1 group II pilin. In contrast, poor twitching resulted from complementation with group III or group V
pilA
genes but increased significantly when the cognate
tfpY
or
tfpZ
accessory genes were cointroduced. The enhanced motility was linked to an increase in recoverable surface pili and not to alterations in total pilin pools. Expression of the group III or V pilins in a PAO1
pilA-pilT
double mutant yielded large amounts of surface pili, regardless of the presence of the accessory genes. Therefore, poor piliation in the absence of the TfpY and TfpZ accessory proteins results from a net increase in PilT-mediated retraction. Similar phenotypes were observed for
tfpY
single and
tfpY-pilT
double knockout mutants of group III strain PA14. A PilA
V
-TfpY chimera produced few surface pili, showing that the accessory proteins are specific for their cognate pilin. The genetic linkage between specific pilin and accessory genes may be evolutionarily conserved because the accessory proteins increase pilus expression on the cell surface, thereby enhancing function.