bluuebunny/crossref_metadata_2025_split huggingface dataset:
The Developmentally Regulatedalb1Gene ofAspergillus fumigatus: Its Role in Modulation of Conidial Morphology and Virulence
Authors: Tsai Huei-Fung , Chang Yun C. , Washburn Ronald G. , Wheeler Michael H. , Kwon-Chung K. J. 
Published: December 2019

ABSTRACTAspergillus fumigatus, an important opportunistic pathogen which commonly affects neutropenic patients, produces conidia with a bluish-green color. We identified a gene,alb1, which is required for conidial pigmentation. Thealb1gene encodes a putative polyketide synthase, and disruption ofalb1resulted in an albino conidial phenotype. Expression ofalb1is developmentally regulated, and the 7-kb transcript is detected only during the conidiation stage. Thealb1mutation was found to block 1,3,6,8-tetrahydroxynaphthalene production, indicating thatalb1is involved in dihydroxynaphthalene-melanin biosynthesis. Scanning electron microscopy studies showed that thealb1disruptant exhibited a smooth conidial surface, whereas complementation of thealb1deletion restored the echinulate wild-type surface. Disruption ofalb1resulted in a significant increase in C3 binding on conidial surfaces, and the conidia of thealb1disruptant were ingested by human neutrophils at a higher rate than were those of the wild type. Thealb1-complemented strain producing bluish-green conidia exhibited inefficient C3 binding and neutrophil-mediated phagocytosis quantitatively similar to those of the wild type. Importantly, thealb1disruptant had a statistically significant loss of virulence compared to the wild-type andalb1-complemented strains in a murine model. These results suggest that disruption ofalb1causes pleiotropic effects on conidial morphology and fungal virulence.
Target Choice and Orientation Preference of the Insertion Sequence IS
903
Authors: Hu Wen-Yuan , Derbyshire Keith M. 
Published: December 2019

ABSTRACT

We have examined the targeting preference of the bacterial insertion element IS
903
by determining the sites of insertion of a large number of transposition events into the 55-kb conjugative plasmid pOX38. Despite the large target size, all the insertions were clustered in four small distinct regions associated with conjugal DNA transfer. Within these regions, many different sites were used for insertion; however, there were a few sites that IS
903
inserted into more than once. Alignment of the insertion sites showed that there was no consensus sequence within the 9-bp target duplication but that there were preferred sequences located symmetrically on either side of the target. This is consistent with target recognition by a dimer or multimer of transposase, with either sequence-specific or structure-specific interactions on both sides of the target. We show further that when one of these preferred regions was cloned into a second conjugative plasmid, pUB307, it was still a preferred target, implying that all the sequences necessary for target selection are contained within this DNA segment. Also, we observed a very strong preference for insertion in a single orientation in pUB307. We examined the possibility that either DNA replication from the origin of vegetative replication,
oriV
, or the origin of transfer,
oriT
, might determine this orientation effect. We find that reversing the direction of vegetative replication had no effect on the orientation of transposon insertions; however, reversing the direction of DNA transfer abolished the orientation effect. This supports the idea that conjugal DNA transfer imparts a polarity on the target that is sensed by the transposon.
Identification and Characterization of Two Quiescent Porin Genes,
nmpC
and
ompN
, in
Escherichia coli
B
E
Authors: Prilipov Alexej , Phale Prashant S. , Koebnik Ralf , Widmer Christine , Rosenbusch Jurg P. 
Published: December 2019

ABSTRACT

The genomic DNA of the B
E
strain of
Escherichia coli
has been scrutinized to detect porin genes that have not been identified so far. Southern blot analysis yielded two DNA segments which proved highly homologous to, yet distinct from, the
ompC
,
ompF
, and
phoE
porin genes. The two genes were cloned and sequenced. One of them, designated
ompN
, encodes a porin which, due to low levels of expression, has eluded prior identification. The functional properties (single-channel conductance) of the OmpN porin, purified to homogeneity, closely resemble those of the OmpC porin from
E. coli
K-12. The second DNA fragment detected corresponds to the
nmpC
gene, which, due to an insertion of an IS
1
element in its coding region, is not expressed in
E. coli
B
E
.
Genetic Recombination in
Bacillus subtilis
168: Effects of
recU
and
recS
Mutations on DNA Repair and Homologous Recombination
Authors: Fernández Silvia , Sorokin Alexei , Alonso Juan C. 
Published: December 2019

ABSTRACT

Bacillus subtilis
recombination-deficient mutants were constructed by inserting a selectable marker (
cat
gene) into the
yppB
and
ypbC
coding regions. The
yppB
:
cat
and
ypbC
:
cat
null alleles rendered cells sensitive to DNA-damaging agents, impaired plasmid transformation (25- and 100-fold), and moderately affected chromosomal transformation when present in an otherwise Rec
+
B. subtilis
strain. The
yppB
gene complemented the defect of the
recG40
strain.
yppB
and
ypbC
and their respective null alleles were termed “
recU
” and “
recU1
” (
recU
:
cat
) and “
recS
” and “
recS1
” (
recS
:
cat
), respectively. The
recU
and
recS
mutations were introduced into
rec
-deficient strains representative of the α (
recF
), β (
addA5 addB72
), γ (
recH342
), and ɛ (
recG40
) epistatic groups. The
recU
mutation did not modify the sensitivity of
recH
cells to DNA-damaging agents, but it did affect inter- and intramolecular recombination in
recH
cells. The
recS
mutation did not modify the sensitivity of
addAB
cells to DNA-damaging agents, and it marginally affected
recF
,
recH
, and
recU
cells. The
recS
mutation markedly reduced (about 250-fold) intermolecular recombination in
recH
cells, and there were reductions of 10- to 20-fold in
recF
,
addAB
, and
recU
cells. Intramolecular recombination was blocked in
recS recF
,
recS addAB
, and
recS recU
cells. RecU and RecS have no functional counterparts in
Escherichia coli
. Altogether, these data indicate that the
recU
and
recS
proteins are required for DNA repair and intramolecular recombination and that the
recF
(α epistatic group),
addAB
(β),
recH
(γ),
recU
(ɛ), and
recS
genes provide overlapping activities that compensate for the effects of single mutation. We tentatively placed
recS
within a new group, termed “ζ.”
tetA
(L) Mutants of a Tetracycline-Sensitive Strain of
Bacillus subtilis
with the Polynucleotide Phosphorylase Gene Deleted
Authors: Bechhofer David H. , Stasinopoulos Stan J. 
Published: December 2019

ABSTRACT

A
Bacillus subtilis
strain with the polynucleotide phosphorylase gene deleted was sensitive for growth in the presence of tetracycline. This strain was used to select for tetracycline-resistant mutants. A point mutation in the
tetA
(L) promoter and a spontaneously occurring
tetA
(L) gene copy number mutant were characterized.
A Periplasmic and Extracellular
c
-Type Cytochrome of
Geobacter sulfurreducens
Acts as a Ferric Iron Reductase and as an Electron Carrier to Other Acceptors or to Partner Bacteria
Authors: Seeliger Sabine , Cord-Ruwisch Ralf , Schink Bernhard 
Published: December 2019

ABSTRACT

An extracellular electron carrier excreted into the growth medium by cells of
Geobacter sulfurreducens
was identified as a
c
-type cytochrome. The cytochrome was found to be distributed in about equal amounts in the membrane fraction, the periplasmic space, and the surrounding medium during all phases of growth with acetate plus fumarate. It was isolated from periplasmic preparations and purified to homogeneity by cation-exchange chromatography, gel filtration, and hydrophobic interaction chromatography. The electrophoretically homogeneous cytochrome had a molecular mass of 9.57 ± 0.02 kDa and exhibited in its reduced state absorption maxima at wavelengths of 552, 522, and 419 nm. The midpoint redox potential determined by redox titration was −0.167 V. With respect to molecular mass, redox properties, and molecular features, this cytochrome exhibited its highest similarity to the cytochromes
c
of
Desulfovibrio salexigens
and
Desulfuromonas acetoxidans
. The
G. sulfurreducens
cytochrome
c
reduced ferrihydrite (Fe(OH)
3
), Fe(III) nitrilotriacetic acid, Fe(III) citrate, and manganese dioxide at high rates. Elemental sulfur, anthraquinone disulfonate, and humic acids were reduced more slowly.
G. sulfurreducens
reduced the cytochrome with acetate as an electron donor and oxidized it with fumarate.
Wolinella succinogenes
was able to reduce externally provided cytochrome
c
of
G. sulfurreducens
with molecular hydrogen or formate as an electron donor and oxidized it with fumarate or nitrate as an electron acceptor. A coculture could be established in which
G. sulfurreducens
reduced the cytochrome with acetate, and the reduced cytochrome was reoxidized by
W. succinogenes
in the presence of nitrate. We conclude that this cytochrome can act as iron(III) reductase for electron transfer to insoluble iron hydroxides or to sulfur, manganese dioxide, or other oxidized compounds, and it can transfer electrons to partner bacteria.
Temperature Regulation of Heat-Labile Enterotoxin (LT) Synthesis in
Escherichia coli
Is Mediated by an Interaction of H-NS Protein with the LT A-Subunit DNA
Authors: Trachman Julie D. , Maas Werner K. 
Published: December 2019

ABSTRACT

Protein and mRNA levels of heat-labile enterotoxin (LT) of
Escherichia coli
are highest at 37°C, and they decrease gradually as temperature is decreased. This temperature effect is eliminated in an Hns
−
mutant. Deletion of portions of DNA coding for the LT A subunit also results in an increase in LT expression at low temperatures, suggesting that the H-NS protein causes inhibition of transcription at low temperatures by interacting with the LT A-subunit DNA. The region that interacts with H-NS is referred to as the downstream regulatory element (DRE). Plasmids in an
hns
strain from which the DRE has been deleted still produce elevated levels of LT at 18°C, suggesting that intact DRE is not required for transcription from the LT promoter.
Opposing Roles of the
Staphylococcus aureus
Virulence Regulators, Agr and Sar, in Triton X-100- and Penicillin-Induced Autolysis
Authors: Fujimoto David F. , Bayles Kenneth W. 
Published: December 2019

ABSTRACT

The regulation of murein hydrolases is a critical aspect of peptidoglycan growth and metabolism. In the present study, we demonstrate that mutations within the
Staphylococcus aureus
virulence factor regulatory genes,
agr
and
sar
, affect autolysis, resulting in decreased and increased autolysis rates, respectively. Zymographic analyses of these mutant strains suggest that
agr
and
sar
exert their effects on autolysis, in part, by modulating murein hydrolase expression and/or activity.
Biosynthesis of Di-
myo
-Inositol-1,1′-Phosphate, a Novel Osmolyte in Hyperthermophilic Archaea
Authors: Chen Liangjing , Spiliotis Elias T. , Roberts Mary F. 
Published: December 2019

ABSTRACT

Biosynthesis of di-
myo
-inositol-1,1′-phosphate (DIP) is proposed to occur with
myo
-inositol and
myo
-inositol 1-phosphate (I-1-P) used as precursors. Activation of the I-1-P with CTP and condensation of the resultant CDP-inositol (CDP-I) with
myo
-inositol then generates DIP. The sole known biosynthetic pathway of inositol in all organisms is the conversion of
d
-glucose-6-phosphate to
myo
-inositol. This conversion requires two key enzymes:
l
-I-1-P synthase and I-1-P phosphatase. Enzymatic assays using
31
P nuclear magnetic resonance spectroscopy as well as a colorimetric assay for inorganic phosphate have confirmed the occurrence of
l
-I-1-P synthase and a moderately specific I-1-P phosphatase. The enzymatic reaction that couples CDP-I with
myo
-inositol to generate DIP has also been detected in
Methanococcus igneus
.
13
C labeling studies with [2,3-
13
C]pyruvate and [3-
13
C]pyruvate were used to examine this pathway in
M. igneus
. Label distribution in DIP was consistent with inositol units formed from glucose-6-phosphate, but the label in the glucose moiety was scrambled via transketolase and transaldolase activities of the pentose phosphate pathway.
Dentilisin Activity Affects the Organization of the Outer Sheath ofTreponema denticola
Authors: Ishihara Kazuyuki , Kuramitsu Howard K. , Miura Tadashi , Okuda Katsuji 
Published: December 2019

ABSTRACTProlyl-phenylalanine-specific serine protease (dentilisin) is a major extracellular protease produced byTreponema denticola. The gene,prtP, coding for the protease was recently cloned and sequenced (K. Ishihara, T. Miura, H. K. Kuramitsu, and K. Okuda, Infect. Immun. 64:5178–5186, 1996). In order to determine the role of this protease in the physiology and virulence ofT. denticola, a dentilisin-deficient mutant, K1, was constructed following electroporation with aprtP-inactivated DNA fragment. No chymotrypsin-like protease activity was detected in the dentilisin-deficient mutant. In addition, the high-molecular-mass oligomeric protein characteristic of the outer sheath of the organism decreased in the mutant. Furthermore, the hydrophobicity of the mutant was decreased, and coaggregation of the mutant withFusobacterium nucleatumwas enhanced compared to that of the wild-type organism. The results obtained with a mouse abscess model system indicated that the virulence of the mutant was attenuated relative to that of the wild-type organism. These results suggest that dentilisin activity plays a major role in the structural organization of the outer sheath ofT. denticola. The loss of dentilsin activity and the structural change in the outer sheath affect the pathogenicity ofT. denticola.
Chemical Structure of Lipid A Isolated fromFlavobacterium meningosepticumLipopolysaccharide
Authors: Kato Hitomi , Haishima Yuji , Iida Takatoshi , Tanaka Akira , Tanamoto Ken-ichi 
Published: December 2019

ABSTRACTThe chemical structure of the lipid A of the lipopolysaccharide component isolated fromFlavobacterium meningosepticumIFO 12535 was elucidated. Methylation and nuclear magnetic resonance analyses showed that two kinds of hydrophilic backbone exist in the free lipid A: a β (1→6)-linked 2-amino-2-deoxy-d-glucose, which is usually present in enterobacterial lipid A’s, and a 2-amino-6-O-(2,3-diamino-2,3-dideoxy-β-d-glucopyranosyl)-2-deoxy-d-glucose, in a molar ratio of 1.00:0.35. Both backbones were α-glycosidically phosphorylated in position 1, and the hydroxyl groups at positions 4, 4′, and 6′ were unsubstituted. Liquid secondary ion-mass spectrometry revealed a pseudomolecular ion atm/z1673 [M-H]−as a major monophosphoryl lipid A component carrying five acyl groups. Fatty acid analysis showed that the lipid A contained 1 mol each of amide-linked (R)-3-OH iC17:0, ester-linked (R)-3-OH iC15:0, amide-linked (R)-3-O-(iC15:0)-iC17:0, and both amide- and ester-linked (R)-3-OH C16:0. Fatty acid distribution analyses using several mass spectrometry determinations demonstrated that the former two constituents were distributed on positions 2 and 3 of the reducing terminal unit of the backbones and that the latter two were attached to the 2′ and 3′ positions in the nonreducing terminal residue.
Bacterial SOS Checkpoint Protein SulA Inhibits Polymerization of Purified FtsZ Cell Division Protein
Authors: Trusca Dorina , Scott Solomon , Thompson Chris , Bramhill David 
Published: December 2019

ABSTRACT

Cell division of
Escherichia coli
is inhibited when the SulA protein is induced in response to DNA damage as part of the SOS checkpoint control system. The SulA protein interacts with the tubulin-like FtsZ division protein. We investigated the effects of purified SulA upon FtsZ. SulA protein inhibits the polymerization and the GTPase activity of FtsZ, while point mutant SulA proteins show little effect on either of these FtsZ activities. SulA did not inhibit the polymerization of purified FtsZ2 mutant protein, which was originally isolated as insensitive to SulA. These studies define polymerization assays for FtsZ which respond to an authentic cellular regulator. The observations presented here support the notion that polymerization of FtsZ is central to its cellular role and that direct, reversible inhibition of FtsZ polymerization by SulA may account for division inhibition.
Both the
fipA
Gene of pKM101 and the
pifC
Gene of F Inhibit Conjugal Transfer of RP1 by an Effect on
traG
Authors: Santini Joanne M. , Stanisich Vilma A. 
Published: December 2019

ABSTRACT

The mechanisms by which gene products inhibit the conjugal transfer of IncP plasmids (e.g., RP1) have been little studied. We have isolated and characterized one such gene,
fipA
(624 nucleotides), from the
Sma
I (14.8 kb)-
Aat
II (15.6 kb) region of pKM101(IncN). This gene, which is also conserved in other IncN plasmids, is transcribed in an anticlockwise direction, probably as part of a transfer operon that includes
traHI
. The FipA protein (24 kDa) appears to be cytoplasmic and, when expressed from a multicopy plasmid, retards the growth of
Escherichia coli
WP2. The mode of action of
fipA
was compared with that of the apparently unrelated
pifC
gene from F(IncFI). Both genes inhibit the transfer of IncPα and IncPβ plasmids but to different degrees. They also inhibit the mobilization of RSF1010 (which requires the RP1 pilus genes and
traG
) but not of CloDF13 (which encodes a
traG
homolog). Evidence that
traG
was the specific target of inhibition was obtained in an artificial system in which cloned
traG
was used to enhance RSF1010 mobilization via the N pilus system. Such enhancement did not occur in the presence of
fipA
or
pifC
. The availability of an in vivo assay of PifC enabled us to show that F
pif
operon expression increased in cells carrying F′
lac
and
traG
, but only if the
traG
coding sequence was intact. This finding suggested that conjugal inhibition of RP1 was most likely due to a PifC-TraG protein interaction. On phenotypic grounds inhibition of
traG
by
fipA
is also likely to occur posttranscriptionally. Whether or not the selection of
traG
as the inhibition target is an evolutionary tactic to limit the spread of P plasmids, we anticipate that
fipA
and
pifC
will prove useful in further investigation of the conjugal roles of
traG
and its homologs.
A Molecular Switch Controlling Competence and Motility: Competence Regulatory Factors ComS, MecA, and ComK Control ς
D
-Dependent Gene Expression in
Bacillus subtilis
Authors: Liu Jiajian , Zuber Peter 
Published: December 2019

ABSTRACT

Bacillus subtilis
, like many bacteria, will choose among several response pathways when encountering a stressful environment. Among the processes activated under growth-restricting conditions are sporulation, establishment of motility, and competence development. Recent reports implicate ComK and MecA-ClpC as part of a system that regulates both motility and competence development. MecA, while negatively controlling competence by inhibiting ComK, stimulates ς
D
-dependent transcription of genes that function in motility and autolysin production. Both ComK-dependent and -independent pathways have been proposed for MecA’s role in the regulation of motility. Mutations in
mecA
reduce the transcription of
hag
. encoding flagellin, and are partially suppressed by
comK
in both medium promoting motility and medium promoting competence. Reduced ς
D
levels are observed in
mecA
mutants grown in competence medium, but no change in ς
D
concentration is detected in a
comK
mutant. The
comF
operon, transcription of which requires ComK, is located immediately upstream of the operon that contains the
flgM
gene, encoding the ς
D
-specific antisigma factor. An insertion mutation that disrupts the putative
comF-flgM
transcription unit confers a phenotype identical to that of the
comK
mutant with respect to
hag-lacZ
expression. Expression of a
flgM-lacZ
operon fusion is reduced in both
sigD
and
comK
mutant cells but is abolished in the
sigD comK
double mutant. Reverse transcription-PCR examination of the
comF-flgM
transcript indicates that readthrough from
comF
into the
flgM
operon is dependent on ComK. ComK negatively controls the transcription of
hag
by stimulating the transcription of
comF-flgM
, thereby increasing the production of the FlgM antisigma factor that inhibits ς
D
activity. There likely exists another
comK
-independent mechanism of
hag
transcription that requires
mecA
and possibly affects the ς
D
concentration in cells undergoing competence development.
Involvement of the
gapA
- and
epd
(
gapB
)-Encoded Dehydrogenases in Pyridoxal 5′-Phosphate Coenzyme Biosynthesis in
Escherichia coli
K-12
Authors: Yang Yong , Zhao Genshi , Man Tsz-Kwong , Winkler Malcolm E. 
Published: December 2019

ABSTRACT

We show that
epd
(
gapB
) mutants lacking an erythrose 4-phosphate (E4P) dehydrogenase are impaired for growth on some media and contain less pyridoxal 5′-phosphate (PLP) and pyridoxamine 5′-phosphate (PMP) than their
epd
+
parent. In contrast to a previous report, we found that
gapA epd
double mutants lacking the glyceraldehyde 3-phosphate and E4P dehydrogenases are auxotrophic for pyridoxine. These results implicate the GapA and Epd dehydrogenases in de novo PLP and PMP coenzyme biosynthesis.
Importance of Glutathione for Growth and Survival of
Escherichia coli
Cells: Detoxification of Methylglyoxal and Maintenance of Intracellular K
+
Authors: Ferguson G. P. , Booth I. R. 
Published: December 2019

ABSTRACT

The role of the tripeptide glutathione in the growth and survival of
Escherichia coli
cells has been investigated. Glutathione-deficient mutants leak potassium and have a reduced cytoplasmic pH. These mutants are more sensitive to methylglyoxal than the parent strain, indicating that in the absence of glutathione-dependent detoxification, acidification of the cytoplasm cannot fully protect cells. However, increasing the intracellular pH of the glutathione-deficient strain resulted in enhanced sensitivity to methylglyoxal. This suggests that acidification of the cytoplasm can provide some protection to
E. coli
cells in the absence of glutathione. In the presence of the Kdp system, glutathione-deficient mutants are highly sensitive to methylglyoxal. This is due to the higher intracellular pH in these cells. In the absence of methylglyoxal, the presence of the Kdp system in a glutathione-deficient strain also leads to an extended lag upon dilution into fresh medium. These data highlight the importance of glutathione for the regulation of the K
+
pool and survival of exposure to methylglyoxal.
The
carB
Gene Encoding the Large Subunit of Carbamoylphosphate Synthetase from
Lactococcus lactis
Is Transcribed Monocistronically
Authors: Martinussen Jan , Hammer Karin 
Published: December 2019

ABSTRACT

The biosynthesis of carbamoylphosphate is catalyzed by the heterodimeric enzyme carbamoylphosphate synthetase. The genes encoding the two subunits of this enzyme in procaryotes are normally transcribed as an operon, but the gene encoding the large subunit (
carB
) in
Lactococcus lactis
is shown to be transcribed as an isolated unit. Carbamoylphosphate is a precursor in the biosynthesis of both pyrimidine nucleotides and arginine. By mutant analysis,
L. lactis
is shown to possess only one
carB
gene; the same gene product is thus required for both biosynthetic pathways. Furthermore, arginine may satisfy the requirement for carbamoylphosphate in pyrimidine biosynthesis through degradation by means of the arginine deiminase pathway. The expression of the
carB
gene is subject to regulation at the level of transcription by pyrimidines, most probably by an attenuator mechanism. Upstream of the
carB
gene, an open reading frame showing a high degree of similarity to those of glutathione peroxidases from other organisms was identified.