bluuebunny/crossref_metadata_2025_split huggingface dataset:
Ecophysiology of “
Halarsenatibacter silvermanii
” Strain SLAS-1
T
, gen. nov., sp. nov., a Facultative Chemoautotrophic Arsenate Respirer from Salt-Saturated Searles Lake, California
Authors: Blum Jodi Switzer , Han Sukkyun , Lanoil Brian , Saltikov Chad , Witte Brian , Tabita F. Robert , Langley Sean , Beveridge Terry J. , Jahnke Linda , Oremland Ronald S. 
Published: February 2009

ABSTRACT

Searles Lake occupies a closed basin harboring salt-saturated, alkaline brines that have exceptionally high concentrations of arsenic oxyanions. Strain SLAS-1
T
was previously isolated from Searles Lake (R. S. Oremland, T. R. Kulp, J. Switzer Blum, S. E. Hoeft, S. Baesman, L. G. Miller, and J. F. Stolz, Science 308:1305-1308, 2005). We now describe this extremophile with regard to its substrate affinities, its unusual mode of motility, sequenced
arrABD
gene cluster, cell envelope lipids, and its phylogenetic alignment within the order
Halanaerobacteriales
, assigning it the name “
Halarsenatibacter silvermanii
” strain SLAS-1
T
. We also report on the substrate dynamics of an anaerobic enrichment culture obtained from Searles Lake that grows under conditions of salt saturation and whose members include a novel sulfate reducer of the order
Desulfovibriales
, the archaeon
Halorhabdus utahensis
, as well as a close homolog of strain SLAS-1
T
.
Rhizobia from Lanzarote, the Canary Islands, That Nodulate
Phaseolus vulgaris
Have Characteristics in Common with
Sinorhizobium meliloti
Isolates from Mainland Spain
Authors: Zurdo-Piñeiro José Luis , García-Fraile Paula , Rivas Raúl , Peix Alvaro , León-Barrios Milagros , Willems Anne , Mateos Pedro Francisco , Martínez-Molina Eustoquio , Velázquez Encarna , van Berkum Peter 
Published: February 2009

ABSTRACT

The stable, low-molecular-weight (LMW) RNA fractions of several rhizobial isolates of
Phaseolus vulgaris
grown in the soil of Lanzarote, an island of the Canary Islands, were identical to a less-common pattern found within
Sinorhizobium meliloti
(assigned to group II) obtained from nodules of alfalfa and alfalfa-related legumes grown in northern Spain. The
P. vulgaris
isolates and the group II LMW RNA
S. meliloti
isolates also were distinguishable in that both had two conserved inserts of 20 and 46 bp in the 16S-23S internal transcribed spacer region that were not present in other strains of
S. meliloti
. The isolates from
P. vulgaris
nodulated bean but not
Medicago sativa
, while those recovered from
Medicago
,
Melilotus
, and
Trigonella
spp. nodulated both host legumes. The bean isolates also were distinguished from those of
Medicago
,
Melilotus
, and
Trigonella
spp. by
nodC
sequence analysis. The
nodC
sequences of the bean isolates were most similar to those reported for
S. meliloti
bv. mediterranense and
Sinorhizobium fredii
bv. mediterranense (GenBank accession numbers DQ333891 and AF217267, respectively). None of the evidence placed the bean isolates from Lanzarote in the genus
Rhizobium
, which perhaps is inconsistent with seed-borne transmission of
Rhizobium etli
from the Americas to the Canaries as an explanation for the presence of bean-nodulating rhizobia in soils of Lanzarote.
p21-Activated Kinases Cla4 and Ste20 Regulate Vacuole Inheritance in
Saccharomyces cerevisiae
Authors: Bartholomew Clinton R. , Hardy Christopher F. J. 
Published: February 2009

ABSTRACT

Each time
Saccharomyces cerevisiae
cells divide they ensure that both the mother and daughter cell inherit a vacuole by actively transporting a portion of the vacuole into the bud. As the mother cell begins budding, a tubular and vesicular segregation structure forms that is transported into the bud by the myosin V motor Myo2, which is bound to the vacuole-specific myosin receptor, Vac17 (41, 59, 70, 79). Upon arriving in the bud the segregation structure is resolved to found the daughter vacuole. The mechanism that regulates segregation structure resolution in a spatially dependent manner is unknown. In addition to resolving the segregation structure, Vac17 is degraded specifically in the bud to provide directionality to vacuole inheritance. It has been proposed that bud-specific degradation of Vac17 is promoted by proteins localized to or activated solely in the bud (77). The p21-activated kinases (PAKs) Cla4 and Ste20 are localized to and activated in the bud. Here we report that Cla4 is localized to the segregation structure just prior to segregation structure resolution, and cells lacking PAK function fail to resolve the segregation structure. Overexpression of either Cla4 or Ste20 inhibited vacuole inheritance and this inhibition was suppressed by the expression of nondegradable
VAC17
. Finally, PAK activity was required for Vac17 degradation in late M phase and
CLA4
overexpression promoted Vac17 degradation. We propose that Cla4 and Ste20 are bud-specific proteins that play roles in both segregation structure resolution and the degradation of Vac17.
Degradation of
Saccharomyces cerevisiae
Transcription Factor Gcn4 Requires a C-Terminal Nuclear Localization Signal in the Cyclin Pcl5
Authors: Streckfuss-Bömeke Katrin , Schulze Florian , Herzog Britta , Scholz Eva , Braus Gerhard H. 
Published: February 2009

ABSTRACT

Pcl5 is a
Saccharomyces cerevisiae
cyclin that directs the phosphorylation of the general amino acid control transcriptional activator Gcn4 by the cyclin-dependent kinase (CDK) Pho85. Phosphorylation of Gcn4 by Pho85/Pcl5 initiates its degradation via the ubiquitin/proteasome system and is regulated by the availability of amino acids. In this study, we show that Pcl5 is a nuclear protein and that artificial dislocation of Pcl5 into the cytoplasm prevents the degradation of Gcn4. Nuclear localization of Pcl5 depends on the β-importin Kap95 and does not require Pho85, Gcn4, or the CDK inhibitor Pho81. Pcl5 nuclear import is independent on the availability of amino acids and is mediated by sequences in its C-terminal domain. The nuclear localization signal is distinct from other functional domains of Pcl5. This is corroborated by a C-terminally truncated Pcl5 variant, which carries the N-terminal nuclear domain of Pho80. This hybrid is still able to fulfill Pcl5 function, whereas Pho80, which is another Pho85 interacting cyclin, does not mediate Gcn4 degradation.
Efficient Gene Replacements in
Toxoplasma gondii
Strains Deficient for Nonhomologous End Joining
Authors: Fox Barbara A. , Ristuccia Jessica G. , Gigley Jason P. , Bzik David J. 
Published: February 2009

ABSTRACT

A high frequency of nonhomologous recombination has hampered gene targeting approaches in the model apicomplexan parasite
Toxoplasma gondii
. To address whether the nonhomologous end-joining (NHEJ) DNA repair pathway could be disrupted in this obligate intracellular parasite, putative KU proteins were identified and a predicted
KU80
gene was deleted. The efficiency of gene targeting via double-crossover homologous recombination at several genetic loci was found to be greater than 97% of the total transformants in
KU80
knockouts. Gene replacement efficiency was markedly increased (300- to 400-fold) in
KU80
knockouts compared to wild-type strains. Target DNA flanks of only ∼500 bp were found to be sufficient for efficient gene replacements in
KU80
knockouts.
KU80
knockouts stably retained a normal growth rate in vitro and the high virulence phenotype of type I strains but exhibited an increased sensitivity to double-strand DNA breaks induced by treatment with phleomycin or γ-irradiation. Collectively, these results revealed that a significant KU-dependent NHEJ DNA repair pathway is present in
Toxoplasma gondii
. Integration essentially occurs only at the homologous targeted sites in the
KU80
knockout background, making this genetic background an efficient host for gene targeting to speed postgenome functional analysis and genetic dissection of parasite biology.
Tagging of Endogenous Genes in aToxoplasma gondiiStrain Lacking Ku80
Authors: Huynh My-Hang , Carruthers Vern B. 
Published: February 2009

ABSTRACTAs with other organisms with a completed genome sequence, opportunities for performing large-scale studies, such as expression and localization, onToxoplasma gondiiare now much more feasible. We present a system for tagging genes endogenously with yellow fluorescent protein (YFP) in a Δku80strain. Ku80 is involved in DNA strand repair and nonhomologous DNA end joining; previous studies in other organisms have shown that in its absence, random integration is eliminated, allowing the insertion of constructs with homologous sequences into the proper loci. We generated a vector consisting of YFP and a dihydrofolate reductase-thymidylate synthase selectable marker. The YFP is preceded by a ligation-independent cloning (LIC) cassette, which allows the insertion of PCR products containing complementary LIC sequences. We demonstrated that the Δku80strain is more effective and efficient in integrating the YFP-tagged constructs into the correct locus than wild-type strain RH. We then selected several hypothetical proteins that were identified by a proteomic screen of excreted-secreted antigens and that displayed microarray expression profiles similar to known micronemal proteins, with the thought that these could potentially be new proteins with roles in cell invasion. We localized these hypothetical proteins by YFP fluorescence and showed expression by immunoblotting. Our findings demonstrate that the combination of the Δku80strain and the pYFP.LIC constructs reduces both the time and cost required to determine localization of a new gene of interest. This should allow the opportunity for performing larger-scale studies of novelT. gondiigenes.
Subspecies IIIa and IIIb Salmonellae Are Defective for Colonization of Murine Models of Salmonellosis Compared to
Salmonella enterica
subsp. I Serovar Typhimurium
Authors: Katribe Erin , Bogomolnaya Lydia M. , Wingert Heather , Andrews-Polymenis Helene 
Published: February 2009

ABSTRACT

Non-subspecies I salmonellae are commensals of cold-blooded vertebrates and cause sporadic disease in mammals. The reasons why non-subspecies I salmonellae do not circulate in populations of warm-blooded vertebrates, but instead only cause occasional disease in this niche, are unknown. We examined the ability of
Salmonella enterica
subsp. IIIa (subsp.
arizonae
) and subsp. IIIb (subsp.
diarizonae
) isolates to grow competitively with subspecies I (serovar Typhimurium) ATCC 14028 in vitro, to colonize
Salmonella
-sensitive BALB/c mice, and to persist in the intestine of
Salmonella
-resistant CBA/J mice in competitive infections. Subspecies IIIa had severely reduced intestinal colonization, intestinal persistence, and systemic spread in mice. Subspecies IIIa is nonmotile on swarming agar and thus may also have reduced motility under viscous conditions in vivo. Surprisingly, subspecies IIIb colonizes the intestinal tract of BALB/c mice normally yet does not spread systemically. Subspecies IIIb colonization of the intestine of CBA/J mice is reduced late in infection. In order to understand why these isolates do not colonize systemic sites, we determined that subspecies IIIa and IIIb are not internalized well and do not replicate in J774-A.1 murine macrophages, despite normal adherence to these cells. We further show that selected effectors of both type III secretion systems 1 and 2 are secreted by subspecies IIIa and IIIb in vitro but that each of these isolates secretes a different combination of effectors. We outline the phenotypic differences between these subspecies and subspecies I and provide a possible explanation for the inability of these strains to spread systemically in murine models.
An
Aeromonas caviae
Genomic Island Is Required for both O-Antigen Lipopolysaccharide Biosynthesis and Flagellin Glycosylation
Authors: Tabei S. Mohammed B. , Hitchen Paul G. , Day-Williams Michaela J. , Merino Susana , Vart Richard , Pang Poh-Choo , Horsburgh Gavin J. , Viches Silvia , Wilhelms Markus , Tomás Juan M. , Dell Anne , Shaw Jonathan G. 
Published: February 2009

ABSTRACT

Aeromonas caviae
Sch3N possesses a small genomic island that is involved in both flagellin glycosylation and lipopolysaccharide (LPS) O-antigen biosynthesis. This island appears to have been laterally acquired as it is flanked by insertion element-like sequences and has a much lower G+C content than the average aeromonad G+C content. Most of the gene products encoded by the island are orthologues of proteins that have been shown to be involved in pseudaminic acid biosynthesis and flagellin glycosylation in both
Campylobacter jejuni
and
Helicobacter pylori
. Two of the genes,
lst
and
lsg
, are LPS specific as mutation of them results in the loss of only a band for the LPS O-antigen. Lsg encodes a putative Wzx flippase, and mutation of Lsg affects only LPS; this finding supports the notion that flagellin glycosylation occurs within the cell before the flagellins are exported and assembled and not at the surface once the sugar has been exported. The proteins encoded by
flmA
,
flmB
,
neuA
,
flmD
, and
neuB
are thought to make up a pseudaminic acid biosynthetic pathway, and mutation of any of these genes resulted in the loss of motility, flagellar expression, and a band for the LPS O-antigen. Furthermore, pseudaminic acid was shown to be present on both flagellin subunits that make up the polar flagellum filament, to be present in the LPS O-antigen of the
A. caviae
wild-type strain, and to be absent from the
A. caviae flmD
mutant strain.
Characterization of a
Mesorhizobium loti
α-Type Carbonic Anhydrase and Its Role in Symbiotic Nitrogen Fixation
Authors: Kalloniati Chrysanthi , Tsikou Daniela , Lampiri Vasiliki , Fotelli Mariangela N. , Rennenberg Heinz , Chatzipavlidis Iordanis , Fasseas Costas , Katinakis Panagiotis , Flemetakis Emmanouil 
Published: February 2009

ABSTRACT

Carbonic anhydrase (CA) (EC 4.2.1.1) is a widespread enzyme catalyzing the reversible hydration of CO
2
to bicarbonate, a reaction that participates in many biochemical and physiological processes.
Mesorhizobium loti
, the microsymbiont of the model legume
Lotus japonicus
, possesses on the symbiosis island a gene (msi040) encoding an α-type CA homologue, annotated as CAA1. In the present work, the
CAA1
open reading frame from
M. loti
strain R7A was cloned, expressed, and biochemically characterized, and it was proven to be an active α-CA. The biochemical and physiological roles of the
CAA1
gene in free-living and symbiotic rhizobia were examined by using an
M. loti
R7A disruption mutant strain. Our analysis revealed that
CAA1
is expressed in both nitrogen-fixing bacteroids and free-living bacteria during growth in batch cultures, where gene expression was induced by increased medium pH.
L. japonicus
plants inoculated with the
CAA1
mutant strain showed no differences in top-plant traits and nutritional status but consistently formed a higher number of nodules exhibiting higher fresh weight, N content, nitrogenase activity, and δ
13
C abundance. Based on these results, we propose that although
CAA1
is not essential for nodule development and symbiotic nitrogen fixation, it may participate in an auxiliary mechanism that buffers the bacteroid periplasm, creating an environment favorable for NH
3
protonation, thus facilitating its diffusion and transport to the plant. In addition, changes in the nodule δ
13
C abundance suggest the recycling of at least part of the HCO
3
−
produced by CAA1.
Regulation ofigaAand the Rcs System by the MviA Response Regulator inSalmonella enterica
Authors: García-Calderón Clara B. , Casadesús Josep , Ramos-Morales Francisco 
Published: February 2009

ABSTRACTIgaA is a membrane protein that prevents overactivation of the Rcs regulatory system in enteric bacteria. Here we provide evidence thatigaAis the first gene in a σ70-dependent operon ofSalmonella entericaserovar Typhimurium that also includesyrfG,yrfH, andyrfI. We also show that the Lon protease and the MviA response regulator participate in regulation of theigaAoperon. Our results indicate that MviA regulatesigaAtranscription in an RpoS-dependent manner, but the results also suggest that MviA may regulate RcsB activation in an RpoS- and IgaA-independent manner.
Nanoscale Structural and Mechanical Properties of Nontypeable
Haemophilus influenzae
Biofilms
Authors: Arce Fernando Terán , Carlson Ross , Monds James , Veeh Richard , Hu Fen Z. , Stewart Philip S. , Lal Ratnesh , Ehrlich Garth D. , Avci Recep 
Published: February 2009

ABSTRACT

Nontypeable
Haemophilus influenzae
(NTHI) bacteria are commensals in the human nasopharynx, as well as pathogens associated with a spectrum of acute and chronic infections. Two important factors that influence NTHI pathogenicity are their ability to adhere to human tissue and their ability to form biofilms. Extracellular polymeric substances (EPS) and bacterial appendages such as pili critically influence cell adhesion and intercellular cohesion during biofilm formation. Structural components in the outer cell membrane, such as lipopolysaccharides, also play a fundamental role in infection of the host organism. In spite of their importance, these pathogenic factors are not yet well characterized at the nanoscale. Here, atomic force microscopy (AFM) was used in aqueous environments to visualize structural details, including probable Hif-type pili, of live NTHI bacteria at the early stages of biofilm formation. Using single-molecule AFM-based spectroscopy, the molecular elasticities of lipooligosaccharides present on NTHI cell surfaces were analyzed and compared between two strains (PittEE and PittGG) with very different pathogenicity profiles. Furthermore, the stiffness of single cells of both strains was measured and subsequently their turgor pressure was estimated.
X-Ray Crystallographic and Mutational Studies of Fluoroacetate Dehalogenase from
Burkholderia
sp. Strain FA1
Authors: Jitsumori Keiji , Omi Rie , Kurihara Tatsuo , Kurata Atsushi , Mihara Hisaaki , Miyahara Ikuko , Hirotsu Ken , Esaki Nobuyoshi 
Published: February 2009

ABSTRACT

Fluoroacetate dehalogenase catalyzes the hydrolytic defluorination of fluoroacetate to produce glycolate. The enzyme is unique in that it catalyzes the cleavage of a carbon-fluorine bond of an aliphatic compound: the bond energy of the carbon-fluorine bond is among the highest found in natural products. The enzyme also acts on chloroacetate, although much less efficiently. We here determined the X-ray crystal structure of the enzyme from
Burkholderia
sp. strain FA1 as the first experimentally determined three-dimensional structure of fluoroacetate dehalogenase. The enzyme belongs to the α/β hydrolase superfamily and exists as a homodimer. Each subunit consists of core and cap domains. The catalytic triad, Asp104-His271-Asp128, of which Asp104 serves as the catalytic nucleophile, was found in the core domain at the domain interface. The active site was composed of Phe34, Asp104, Arg105, Arg108, Asp128, His271, and Phe272 of the core domain and Tyr147, His149, Trp150, and Tyr212 of the cap domain. An electron density peak corresponding to a chloride ion was found in the vicinity of the N
ε1
atom of Trp150 and the N
ε2
atom of His149, suggesting that these are the halide ion acceptors. Site-directed replacement of each of the active-site residues, except for Trp150, by Ala caused the total loss of the activity toward fluoroacetate and chloroacetate, whereas the replacement of Trp150 caused the loss of the activity only toward fluoroacetate. An interaction between Trp150 and the fluorine atom is probably an absolute requirement for the reduction of the activation energy for the cleavage of the carbon-fluorine bond.
Functional Genomics ofEnterococcus faecalis: Multiple Novel Genetic Determinants for Biofilm Formation in the Core Genome
Authors: Ballering Katie S. , Kristich Christopher J. , Grindle Suzanne M. , Oromendia Ana , Beattie David T. , Dunny Gary M. 
Published: February 2009

ABSTRACTThe ability ofEnterococcus faecalisto form robust biofilms on host tissues and on abiotic surfaces such as catheters likely plays a major role in the pathogenesis of opportunistic antibiotic-resistantE. faecalisinfections and in the transfer of antibiotic resistance genes. We have carried out a comprehensive analysis of genetic determinants of biofilm formation in the core genome ofE. faecalis. Here we describe 68 genetic loci predicted to be involved in biofilm formation that were identified byrecombinaseinvivoexpressiontechnology (RIVET); most of these genes have not been studied previously. Differential expression of a number of these determinants during biofilm growth was confirmed by quantitative reverse transcription-PCR, and genetic complementation studies verified a role in biofilm formation for several candidate genes. Of particular interest was genetic locus EF1809, predicted to encode a regulatory protein of the GntR family. We isolated 14 independent nonsibling clones containing the putative promoter region for this gene in the RIVET screen; EF1809 also showed the largest increase in expression during biofilm growth of any of the genes tested. Since an in-frame deletion of EF1809 resulted in a severe biofilm defect that could be complemented by the cloned wild-type gene, we have designated EF1809ebrA(enterococcalbiofilmregulator). Most of the novel genetic loci identified in our studies are highly conserved in gram-positive bacterial pathogens and may thus constitute a pool of uncharacterized genes involved in biofilm formation that may be useful targets for drug discovery.
Role of FimW, FimY, and FimZ in Regulating the Expression of Type I Fimbriae in
Salmonella enterica
Serovar Typhimurium
Authors: Saini Supreet , Pearl Jeffrey A. , Rao Christopher V. 
Published: February 2009

ABSTRACT

Type I fimbriae in
Salmonella enterica
serovar Typhimurium are surface appendages that facilitate binding to eukaryotic cells. Expression of the
fim
gene cluster is known to be regulated by three proteins—FimW, FimY, and FimZ—and a tRNA encoded by
fimU
. In this work, we investigated how these proteins and tRNA coordinately regulate
fim
gene expression. Our results indicate that FimY and FimZ independently activate the P

fimA

promoter which controls the expression of the
fim
structural genes. FimY and FimZ were also found to strongly activate each other's expression and weakly activate their own expression. FimW was found to negatively regulate
fim
gene expression by repressing transcription from the P

fimY

promoter, independent of FimY or FimZ. Moreover, FimW and FimY interact within a negative feedback loop, as FimY was found to activate the P

fimW

promoter. In the case of
fimU
, the expression of this gene was not found to be regulated by FimW, FimY, or FimZ. We also explored the effect of
fim
gene expression on
Salmonella
pathogenicity island 1 (SPI1). Our results indicate that FimZ alone is able to enhance the expression of
hilE
, a known repressor of SPI1 gene expression. Based on our results, we were able to propose an integrated model for the
fim
gene circuit. As this model involves a combination of positive and negative feedback, we hypothesized that the response of this circuit may be bistable and thus a possible mechanism for phase variation. However, we found that the response was continuous and not bistable.
ClpXP Degrades SsrA-Tagged Proteins inStreptococcus pneumoniae
Authors: Ahlawat Sarita , Morrison Donald A. 
Published: February 2009

ABSTRACTBacterial proteins that are abnormally truncated due to incomplete mRNA or the presence of rare codons are extended by an SsrA tag during ribosome rescue in atrans-translation process important for maintaining protein quality. InEscherichia coli, the SsrA-tagged proteins become the target of the Tsp, Lon, FtsH, ClpXP, and ClpAP proteases. Here we show that degradation of model SsrA-tagged proteins inStreptococcus pneumoniaedepends primarily or exclusively on ClpXP in vivo. In addition, we show theE. coliSsrA tag is also a target ofS. pneumoniaeClpXP in vivo, even though the N-terminal portions of the tags differ significantly between the two species, suggesting there may be no adaptor protein for SsrA inS. pneumoniae.