bluuebunny/crossref_metadata_2025_split huggingface dataset:
Identification and Application of Plasmids Suitable for Transfer of Foreign DNA to Members of the Genus
Gordonia
Authors: Arenskötter Matthias , Baumeister Dirk , Kalscheuer Rainer , Steinbüchel Alexander 
Published: August 2003

ABSTRACT

Gene transfer systems for
Gordonia polyisoprenivorans
strains VH2 and Y2K based on electroporation and conjugation, respectively, were established. Several parameters were optimized, resulting in transformation efficiencies of >4 × 10
5
CFU/μg of plasmid DNA. In contrast to most previously described electroporation protocols, the highest efficiencies were obtained by applying a heat shock after the intrinsic electroporation. Under these conditions, transfer and autonomous replication of plasmid pNC9503 was also demonstrated to proceed in
G. alkanivorans
DSM44187,
G. nitida
DSM44499
T
,
G. rubropertincta
DSM43197
T
,
G. rubropertincta
DSM46038, and
G. terrae
DSM43249
T
. Conjugational plasmid DNA transfer to
G. polyisoprenivorans
resulted in transfer frequencies of up to 5 × 10
−6
of the recipient cells. Recombinant strains capable of polyhydroxyalkanoate synthesis from alkanes were constructed.
Description of a “Phoenix” Phenomenon in the Growth of
Campylobacter jejuni
at Temperatures Close to the Minimum for Growth
Authors: Kelly A. F. , Martínez-Rodriguez A. , Bovill R. A. , Mackey B. M. 
Published: August 2003

ABSTRACT

When
Campylobacter jejuni
cultures that had been grown in broth at 39°C were subcultured into fresh medium at 30°C, there was a transient period of growth followed by a decline in viable-cell numbers before growth resumed once more. We propose that this complex behavior is the net effect of the growth of inoculum cells followed by a loss of viability due to oxidative stress and the subsequent emergence of a spontaneously arising mutant population that takes over the culture.
Analysis of the
argK-tox
Gene Cluster in Nontoxigenic Strains of
Pseudomonas syringae
pv. phaseolicola
Authors: González Ana Isabel , Pérez de la Vega Marcelino , Ruiz María Luisa , Polanco Carlos 
Published: August 2003

ABSTRACT

The analysis of 46 isolates obtained directly from different and distant common bean fields from the northwestern part of Spain revealed that they do not produce phaseolotoxin. The isolates were classified as race 5, and their analysis revealed that they do not carry the
argK-tox
gene cluster involved in the biosynthesis of the phaseolotoxin.
Increased Exopolysaccharide Production in
Lactococcus lactis
due to Increased Levels of Expression of the NIZO B40
eps
Gene Cluster
Authors: Boels Ingeborg C. , van Kranenburg Richard , Kanning Marja W. , Chong Barrie Fong , de Vos Willem M. , Kleerebezem Michiel 
Published: August 2003

ABSTRACT

Exopolysaccharides (EPS) play an important role in the rheology and texture of fermented food products. This is the first report demonstrating that homologous overexpression of a complete
eps
gene cluster in
Lactococcus lactis
leads to increased EPS production levels. A ninefold-elevated EPS plasmid copy number led to an almost threefold increase in the
eps
expression level, resulting in an almost fourfold increase in the NIZO B40 EPS production level. It was previously reported that increased EPS precursor levels did not influence NIZO B40 EPS production levels. However, the present results indicate that the maximal NIZO B40 EPS production level is limited by the activity level of the expression products of the
eps
gene cluster rather than by the level of EPS precursors.
Mannitol Is Required for Stress Tolerance inAspergillus nigerConidiospores
Authors: Ruijter George J. G. , Bax Maarten , Patel Hema , Flitter Simon J. , van de Vondervoort Peter J. I. , de Vries Ronald P. , vanKuyk Patricia A. , Visser Jaap 
Published: August 2003

ABSTRACTd-Mannitol is the predominant carbon compound in conidiospores of the filamentous fungusAspergillus nigerand makes up 10 to 15% of the dry weight. A number of physiological functions have been ascribed to mannitol, including serving as a reserve carbon source, as an antioxidant, and to store reducing power. In this study, we cloned and characterized theA. niger mpdAgene, which encodes mannitol 1-phosphate dehydrogenase (MPD), the first enzyme in the mannitol biosynthesis pathway. ThempdApromoter contains putative binding sites for the development-specific transcription factors BRLA and ABAA. Furthermore, increased expression ofmpdAin sporulating mycelium suggests that mannitol biosynthesis is, to a certain extent, developmentally regulated inA. niger. Inactivation ofmpdAabolished mannitol biosynthesis in growing mycelium and reduced the mannitol level in conidiospores to 30% that in the wild type, indicating that MPD and mannitol 1-phosphate phosphatase form the major metabolic pathway for mannitol biosynthesis inA. niger. The viability of spores after prolonged storage and germination kinetics were normal in anmpdAnull mutant, indicating that mannitol does not play an essential role as a reserve carbon source inA. nigerconidia. However, conidiospores of a ΔmpdAstrain were extremely sensitive to a variety of stress conditions, including high temperature, oxidative stress and, to a lesser extent, freezing and lyophilization. Since mannitol supplied in the medium during sporulation repaired this deficiency, mannitol appears to be essential for the protection ofA. nigerspores against cell damage under these stress conditions.
Diverged Binding Specificity of Rim101p, the
Candida albicans
Ortholog of PacC
Authors: Ramón Ana M. , Fonzi William A. 
Published: August 2003

ABSTRACT

The biology of
Candida albicans
, including dimorphism and virulence, is significantly influenced by environmental pH. The response to ambient pH includes the pH-conditional expression of several genes, which is directly or indirectly regulated by Rim101p. Rim101p is homologous to PacC, a transcription factor that regulates pH-conditional gene expression in
Aspergillus nidulans
. PacC binds 5′-GCCARG-3′ sequences upstream of pH-responsive genes and either activates or represses transcription. The absence of
pacC
consensus binding sites upstream of
PHR1
, a
RIM101
-dependent, alkaline pH-induced gene of
C. albicans
, suggested either that
PHR1
is indirectly regulated by Rim101p or that the binding specificity of Rim101p is different. In vitro binding studies demonstrated that Rim101p strongly bound two regions upstream of
PHR1
that were only weakly bound by PacC. Deletion analysis and site-specific mutagenesis demonstrated that both sites were functionally significant, mutation of either site reduced
RIM101
-dependent induction, and expression was abolished in the double mutant. Furthermore, oligonucleotides containing these sites conferred pH-conditional expression when inserted upstream of a reporter gene. The consensus sequence of these sites, 5′-CCAAGAAA-3′, was identical to the binding recognition sequence identified by in vitro selection of Rim101p binding oligonucleotides from a random pool. The functional significance of this binding sequence was reinforced by its observed presence upstream of a number of newly identified pH-conditional genes. We conclude that Rim101p acts as a transcription factor and directly regulates pH-conditional gene expression but has a binding specificity different from that of PacC.
PAK
Paradox:
Paramecium
Appears To Have More K
+
-Channel Genes than Humans
Authors: Haynes W. John , Ling Kit-Yin , Saimi Yoshiro , Kung Ching 
Published: August 2003

ABSTRACT

K
+
-selective ion channels (K
+
channels) have been found in bacteria, archaea, eucarya, and viruses. In
Paramecium
and other ciliates, K
+
currents play an essential role in cilia-based motility. We have retrieved and sequenced seven closely related
Paramecium
K
+
-channel gene (
PAK
) sequences by using previously reported fragments. An additional eight unique K
+
-channel sequences were retrieved from an indexed library recently used in a pilot genome sequencing project. Alignments of these protein translations indicate that while these 15 genes have diverged at different times, they all maintain many characteristics associated with just one subclass of metazoan K
+
channels (CNG/ERG type). Our results indicate that most of the genes are expressed, because all predicted frameshifts and several gaps in the homolog alignments contain
Paramecium
intron sequences deleted from reverse transcription-PCR products. Some of the variations in the 15 genomic nucleotide sequences involve an absence of introns, even between very closely related sequences, suggesting a potential occurrence of reverse transcription in the past. Extrapolation from the available genome sequence indicates that
Paramecium
harbors as many as several hundred of this one type of K
+
-channel gene. This quantity is far more numerous than those of K
+
-channel genes of all types known in any metazoan (e.g., ∼80 in humans, ∼30 in flies, and ∼15 in
Arabidopsis
). In an effort to understand this plurality, we discuss several possible reasons for their maintenance, including variations in expression levels in response to changes in the freshwater environment, like that seen with other major plasma membrane proteins in
Paramecium
.
Asexual Development Is Increased in
Neurospora crassa
cat
-
3
-Null Mutant Strains
Authors: Michán Shaday , Lledías Fernando , Hansberg Wilhelm 
Published: August 2003

ABSTRACT

We use asexual development of
Neurospora crassa
as a model system with which to determine the causes of cell differentiation. Air exposure of a mycelial mat induces hyphal adhesion, and adherent hyphae grow aerial hyphae that, in turn, form conidia. Previous work indicated the development of a hyperoxidant state at the start of these morphogenetic transitions and a large increase in catalase activity during conidiation. Catalase 3 (CAT-3) increases at the end of exponential growth and is induced by different stress conditions. Here we analyzed the effects of
cat
-
3
-null strains on growth and asexual development. The lack of CAT-3 was not compensated by other catalases, even under oxidative stress conditions, and
cat
-

3
RIP

colonies were sensitive to H
2
O
2
, indicating that wild-type (Wt) resistance to external H
2
O
2
was due to CAT-3.
cat
-

3
RIP

colonies grown in the dark produced high levels of carotenes as a consequence of oxidative stress. Light exacerbated oxidative stress and further increased carotene synthesis. In the
cat
-

3
RIP

mutant strain, increased aeration in liquid cultures led to increased hyphal adhesion and protein oxidation. Compared to the Wt, the
cat
-

3
RIP

mutant strain produced six times more aerial hyphae and conidia in air-exposed mycelial mats, as a result of longer and more densely packed aerial hyphae. Protein oxidation in colonies was threefold higher and showed more aerial hyphae and conidia in mutant strains than did the Wt. Results indicate that oxidative stress due to lack of CAT-3 induces carotene synthesis, hyphal adhesion, and more aerial hyphae and conidia.
POS5
Gene of
Saccharomyces cerevisiae
Encodes a Mitochondrial NADH Kinase Required for Stability of Mitochondrial DNA
Authors: Strand Micheline K. , Stuart Gregory R. , Longley Matthew J. , Graziewicz Maria A. , Dominick Olivia C. , Copeland William C. 
Published: August 2003

ABSTRACT

In a search for nuclear genes that affect mutagenesis of mitochondrial DNA in
Saccharomyces cerevisiae
, an ATP-NAD (NADH) kinase, encoded by
POS5
, that functions exclusively in mitochondria was identified. The
POS5
gene product was overproduced in
Escherichia coli
and purified without a mitochondrial targeting sequence. A direct biochemical assay demonstrated that the
POS5
gene product utilizes ATP to phosphorylate both NADH and NAD
+
, with a twofold preference for NADH. Disruption of
POS5
increased minus-one frameshift mutations in mitochondrial DNA 50-fold, as measured by the

arg8
m

reversion assay, with no increase in nuclear mutations. Also, a dramatic increase in petite colony formation and slow growth on glycerol or limited glucose were observed.
POS5
was previously described as a gene required for resistance to hydrogen peroxide. Consistent with a role in the mitochondrial response to oxidative stress, a
pos5
deletion exhibited a 28-fold increase in oxidative damage to mitochondrial proteins and hypersensitivity to exogenous copper. Furthermore, disruption of
POS5
induced mitochondrial biogenesis as a response to mitochondrial dysfunction. Thus, the
POS5
NADH kinase is required for mitochondrial DNA stability with a critical role in detoxification of reactive oxygen species. These results predict a role for NADH kinase in human mitochondrial diseases.
The Effects of the Asian Crisis of 1997 on Emergent Markets Through a Critical Phenomena Model
Authors: Figueroa M. G. , Mariani M. C. , Ferraro M. B. 
Published: August 2003

In the present work we have analyzed the financial Asian crisis of 1997, and its consequences on emerging markets. We have done so by means of a phase transition model originally presented by A. Johansen and D. Sornette [1].We have analyzed the crashes on leading indices of Hong Kong (HSI), Turkey (XU100), Mexico (MMX), Brazil (Bovespa) and Argentina (Merval).With the exception of Argentina's index, we were able to obtain optimum values for the critical date, corresponding to the most probable date of the crash.
Late-onset noninfectious pulmonary complications after allogeneic stem cell transplantation are significantly associated with chronic graft-versus-host disease and with the graft-versus-leukemia effect
Authors: Sakaida Emiko , Nakaseko Chiaki , Harima Akane , Yokota Akira , Cho Ryuko , Saito Yasushi , Nishimura Miki 
Published: August 2003

AbstractLate-onset noninfectious pulmonary complications (LONIPCs) occurring beyond 3 months after allogeneic stem cell transplantation (allo-SCT) have become recognized as life-threatening complications, and they reduce the recipient's quality of life. However, the pathogenesis and optimal treatment for LONIPCs are still unclear. In this study, we retrospectively analyzed the incidence and outcome of LONIPCs among allo-SCT recipients. Between October 1993 and September 2001, 96 patients underwent allo-SCT and 76 patients who survived and were free of disease for more than 3 months after SCT were enrolled. Among the 76 patients, 18 patients (23.7%) developed LONIPCs at a median interval of 227 days after allo-SCT (range, 91-1105 days). The patients with LONIPCs were subclassified into those with bronchiolitis obliterans (BO) (6 patients), with interstitial pneumonia (IP) (11 patients), or with both BO and IP (1 patient). The presence of extensive chronic graft-versus-host disease (GVHD) was significantly associated with the development of LONIPCs (P = .0008). Liver or skin involvement in chronic GVHD was not associated, but sicca syndrome was significantly associated with the development of LONIPCs (P &lt; .0001). Most of the IP patients (58.3%) responded well to immunosuppressive treatment, while BO patients did not respond to the therapy. Eight of the 18 patients with LONIPCs died. The major cause of death was respiratory failure (62.5%). The relapse rate of primary malignant disease in the LONIPC patients was significantly lower than that of non-LONIPC patients (1 of 17 [5.9%] versus 16 of 52 [30.8%]; P = .0387). These results indicate that the development of LONIPCs was strongly associated with chronic GVHD and especially with sicca syndrome and the graft-versus-leukemia (GVL) effect. (Blood. 2003;102:4236-4242)
Chemokine stimulation of human peripheral blood T lymphocytes induces rapid dephosphorylation of ERM proteins, which facilitates loss of microvilli and polarization
Authors: Brown Martin J. , Nijhara Ruchika , Hallam John A. , Gignac Michelle , Yamada Kenneth M. , Erlandsen Stanley L. , Delon Jérôme , Kruhlak Michael , Shaw Stephen 
Published: August 2003

AbstractLymphocyte microvilli mediate initial rolling-adhesion along endothelium but are lost during transmigration from circulation to tissue. However, the mechanism for resorption of lymphocyte microvilli remains unexplored. We show that chemokine stimulation of human peripheral blood T (PBT) cells is sufficient to induce rapid resorption of microvilli. Microvilli in other cells are regulated by ezrin/radixin/moesin (ERM) proteins, which link the plasma membrane to the cortical F-actin cytoskeleton; maintenance of these linkages requires ERM activation, reflected by phosphorylation at a specific carboxy-terminal threonine residue. Carboxyphosphorylated-ERM (cpERM) proteins in resting PBT cells show a punctate peripheral distribution consistent with localization to microvilli. cpERM dephosphorylation begins within seconds of stimulation by chemokines (stromal derived factor 1α [SDF-1α] or secondary lymphoid tissue cytokine), and ERM proteins lose their punctate distribution with kinetics paralleling the loss of microvilli. The cpERM proteins are preferentially associated with the cytoskeleton at rest and this association is lost with chemokine-induced dephosphorylation. Transfection studies show that a dominant-negative ERM construct destroys microvilli, whereas a construct mimicking cpERM facilitates formation of microvilli, retards chemokine-induced loss of microvilli, and markedly impairs chemokine-induced polarization. Thus, chemokine induces rapid dephosphorylation and inactivation of cpERM, which may in turn facilitate 2 aspects of cytoskeletal reorganization involved in lymphocyte recruitment: loss of microvilli and polarization.
Phenotypic correction of a mouse model of hemophilia A using AAV2 vectors encoding the heavy and light chains of FVIII
Authors: Scallan Ciaran D. , Liu Tongyao , Parker Amy E. , Patarroyo-White Susannah L. , Chen Haifeng , Jiang Haiyan , Vargas Joseph , Nagy Dea , Powell Sharon K. , Wright J. Fraser , Sarkar Rita , Kazazian Haig H. , McClelland Alan , Couto Linda B. 
Published: August 2003

AbstractUsing separate adeno-associated viral 2 (AAV2) vectors to deliver the heavy and light chains of factor VIII (FVIII) we have overcome the packaging limitations of AAV, achieving phenotypic correction of hemophilia A in mice. AAV vectors were constructed that use a liver-specific promoter and the cDNA sequences of either the human or canine heavy and light chains of FVIII. After intraportal vein injection of these vectors in hemophilia-A mice, therapeutic to superphysiologic levels of active FVIII were achieved in plasma in a dose-dependent manner. Phenotypic correction of the bleeding diathesis was demonstrated by survival of all treated mice after tail clipping. Biochemical analysis demonstrated lower levels of heavy-chain (25- to 100-fold) compared with light-chain protein in the plasma of treated animals. Differences in gene transfer and transcription did not account for the differences in protein expression. We hypothesize that improvements in FVIII activity could be achieved by improvements in FVIII heavy-chain expression. This work demonstrates that cotransduction of liver with AAV vectors expressing the heavy and light chains of FVIII corrects hemophilia A in vivo, providing an alternative approach to the use of a single vector. This strategy may potentially be useful for other large therapeutic proteins that contain functionally distinct domains.
T-cell interaction with ICAM-1/ICAM-2 double-deficient brain endothelium in vitro: the cytoplasmic tail of endothelial ICAM-1 is necessary for transendothelial migration of T cells
Authors: Lyck Ruth , Reiss Yvonne , Gerwin Nicole , Greenwood John , Adamson Peter , Engelhardt Britta 
Published: August 2003

AbstractEndothelial intercellular adhesion molecule 1 (ICAM-1) and ICAM-2 are both involved in lymphocyte extravasation during immunosurveillance and inflammation. To define their exact role during T-cell extravasation, we used mouse T cells and ICAM-1-/-ICAM-2-/- brain endothelioma cells. ICAM-1-/-ICAM-2-/- brain endothelioma cells did not support transendothelial migration (TEM) of T cells in vitro. Re-expression of different ICAM-1 mutants in the ICAM-1-/-ICAM-2-/- endothelioma line bEndI1/2.1 or in the ICAM-1-/- endothelioma line bEndI1.1 demonstrated that the extracellular domain of ICAM-1 suffices to support T-cell adhesion while the presence of the cytoplasmic tail was strictly required for TEM. Surprisingly, tyrosine phosphorylation of endothelial ICAM-1 was not necessary for TEM of T cells or for Rho guanosine triphosphatase (RhoGTPase) activation. Furthermore, cytoplasmic deletion mutants of ICAM-1 were unable to mediate RhoGTPase activation. Thus, our data demonstrate that the cytoplasmic tail of endothelial ICAM-1—independently from tyrosine phosphorylation—is essential for supporting TEM of T lymphocytes, while Rho signaling is involved in endothelial cells. (Blood. 2003;102:3675-3683)
Human dendritic cells express neuronal Eph receptor tyrosine kinases: role of EphA2 in regulating adhesion to fibronectin
Authors: de Saint-Vis Blandine , Bouchet Caroline , Gautier Grégory , Valladeau Jenny , Caux Christophe , Garrone Pierre 
Published: August 2003

AbstractEph receptor tyrosine kinases and their ligands, the ephrins, have been primarily described in the nervous system for their roles in axon guidance, development, and cell intermingling. Here we address whether Eph receptors may also regulate dendritic cell (DC) trafficking. Reverse transcription-polymerase chain reaction (RT-PCR) analysis showed that DCs derived from CD34+ progenitors, but not from monocytes, expressed several receptors, in particular EphA2, EphA4, EphA7, EphB1, and EphB3 mRNA. EphB3 was specifically expressed by Langerhans cells, and EphA2 and EphA7 were expressed by both Langerhans- and interstitial-type DCs. EphA and EphB protein expression on DCs generated in vitro was confirmed by staining with ephrin-A3-Fc and ephrin-B3-Fc fusion proteins that bind to different Eph members, in particular EphA2 and EphB3. Immunostaining with anti-EphA2 antibodies demonstrated the expression of EphA2 by immature DCs and by skin Langerhans cells isolated ex vivo. Interestingly, ephrin expression was detected in epidermal keratinocytes and also in DCs. Adhesion of CD34+-derived DCs to fibronectin, but not to poly-l-lysine, was increased in the presence of ephrin-A3-Fc, a ligand of EphA2, through a β1 integrin activation pathway. As such, EphA2/ephrin-A3 interactions may play a role in the localization and network of Langerhans cells in the epithelium and in the regulation of their trafficking. (Blood. 2003;102:4431-4440)