bluuebunny/crossref_metadata_2025_split huggingface dataset:
Ecological Constraints Limit the Fitness of Fungal Hybrids in the
Heterobasidion annosum
Species Complex
Authors: Garbelotto Matteo , Gonthier Paolo , Nicolotti Giovanni 
Published: July 2007

ABSTRACT

The ability of two closely related species to maintain species boundaries in spite of retained interfertility between them is a documented driving force of speciation. Experimental evidence to support possible interspecific postzygotic isolation mechanisms for organisms belonging to the kingdom Fungi is still missing. Here we report on the outcome of a series of controlled comparative inoculation experiments of parental wild genotypes and F
1
hybrid genotypes between closely related and interfertile taxa within the
Heterobasidion annosum
fungal species complex. Results indicated that these fungal hybrids are not genetically unfit but can fare as well as parental genotypes when inoculated on substrates favorable to both parents. However, when placed in substrates favoring one of the parents, hybrids are less competitive than the parental genotypes specialized on that substrate. Furthermore, in some but not all fungus × plant combinations, a clear asymmetry in fitness was observed between hybrids carrying identical nuclear genomes but different cytoplasms. This work provides some of the first experimental evidence of ecologically driven postzygotic reinforcement of isolation between closely related fungal species characterized by marked host specificity. Host specialization is one of the most striking traits of a large number of symbiotic and parasitic fungi; thus, we suggest the ecological mechanism proven here to reinforce isolation among
Heterobasidion
spp. may be generally valid for host-specialized fungi. The validity of this generalization is supported by the low number of known fungal hybrids and by their distinctive feature of being found in substrates different from those colonized by parental species.
Advanced Environmental Surveillance and Molecular Analyses Indicate Separate Importations Rather than Endemic Circulation of Wild Type 1 Poliovirus in Gaza District in 2002
Authors: Manor Y. , Blomqvist S. , Sofer D. , Alfandari J. , Halmut T. , Abramovitz B. , Mendelson E. , Shulman L. M. 
Published: July 2007

ABSTRACT
An improved sewage surveillance algorithm (sample acquisition, processing, and molecular analysis) for wild and vaccine-derived polioviruses was developed and validated. It was based on plaque isolation with sensitive and high-throughput methods. The molecular analysis included sequencing; a comparison of the type, rate, and distribution of nucleotide substitutions with a profile for outbreaks evolving from a single progenitor; and phylogenetic analysis for relative similarity. The analyses revealed that two environmental wild type 1 isolates from the Gaza district in 2002 were imported separately, most likely from Egyptian southern governorates, and were not linked by endemic circulation. These findings illustrate the continuous spreading potential of wild-type poliovirus and underscore the value of extensive environmental surveillance employing advanced molecular analysis to monitor wild poliovirus in poliomyelitis-free regions.
A Cell Number-Counting Factor Regulates Levels of a Novel Protein, SslA, as Part of a Group Size Regulation Mechanism in
Dictyostelium
Authors: Gao Tong , Roisin-Bouffay Celine , Hatton R. Diane , Tang Lei , Brock Debra A. , DeShazo Tiffany , Olson Laura , Hong Wan-Pyo , Jang Wonhee , Canseco Elvia , Bakthavatsalam Deenadayalan , Gomer Richard H. 
Published: July 2007

ABSTRACT

Developing
Dictyostelium
cells form aggregation streams that break into groups of ∼2 × 10
4
cells. The breakup and subsequent group size are regulated by a secreted multisubunit counting factor (CF). To elucidate how CF regulates group size, we isolated second-site suppressors of
smlA
−
, a transformant that forms small groups due to oversecretion of CF.
smlA
−
sslA1
(
CR11
) cells form roughly wild-type-size groups due to an insertion in the beginning of the coding region of
sslA1
, one of two highly similar genes encoding a novel protein. The insertion increases levels of SslA. In wild-type cells, the
sslA1
(
CR11
) mutation forms abnormally large groups. Reducing SslA levels by antisense causes the formation of smaller groups. The
sslA
(
CR11
) mutation does not affect the extracellular accumulation of CF activity or the CF components countin and CF50, suggesting that SslA does not regulate CF secretion. However, CF represses levels of SslA. Wild-type cells starved in the presence of
smlA
−
cells, recombinant countin, or recombinant CF50 form smaller groups, whereas
sslA1
(
CR11
) cells appear to be insensitive to the presence of
smlA
−
cells, countin, or CF50, suggesting that the
sslA1
(
CR11
) insertion affects CF signal transduction. We previously found that CF reduces intracellular glucose levels. s
slA
(
CR11
) does not significantly affect glucose levels, while glucose increases SslA levels. Together, the data suggest that SslA is a novel protein involved in part of a signal transduction pathway regulating group size.
A
Toxoplasma gondii
Leucine-Rich Repeat Protein Binds Phosphatase Type 1 Protein and Negatively Regulates Its Activity
Authors: Daher Wassim , Oria Gabrielle , Fauquenoy Sylvain , Cailliau Katia , Browaeys Edith , Tomavo Stanislas , Khalife Jamal 
Published: July 2007

ABSTRACT

We have characterized the
Toxoplasma gondii
protein phosphatase type 1 (TgPP1) and a potential regulatory binding protein belonging to the leucine-rich repeat protein family, designated TgLRR1. TgLRR1 is capable of binding to TgPP1 to inhibit its activity and to override a G
2
/M cell cycle checkpoint in
Xenopus
oocytes. In the parasite, TgLRR1 mRNA and protein are both highly expressed in the rapidly replicating and virulent tachyzoites, while only low levels are detected in the slowly dividing and quiescent bradyzoites. TgPP1 mRNA and protein levels are equally abundant in tachyzoites and bradyzoites. Affinity pull down and immunoprecipitation experiments reveal that the TgLRR1-TgPP1 interaction takes place in the nuclear subcompartment of tachyzoites. These results are consistent with those of localization studies using both indirect immunofluorescence with specific polyclonal antibody and transient transfection of
T. gondii
vector expressing TgLRR1 and TgPP1. The inability to obtain stable transgenic tachyzoites suggested that overexpression of TgLRR1 and TgPP1 may impair the parasite's growth. Together with the activation of
Xenopus
oocyte meiosis reinitiation, these data indicate that TgLRR1 protein could play a role in the regulation of the
T. gondii
cell cycle through the modulation of phosphatase activity.
Transcriptional and Physiological Responses of
Bradyrhizobium japonicum
to Desiccation-Induced Stress
Authors: Cytryn Eddie J. , Sangurdekar Dipen P. , Streeter John G. , Franck William L. , Chang Woo-suk , Stacey Gary , Emerich David W. , Joshi Trupti , Xu Dong , Sadowsky Michael J. 
Published: July 2007

ABSTRACT

The growth and persistence of rhizobia and bradyrhizobia in soils are negatively impacted by drought conditions. In this study, we used genome-wide transcriptional analyses to obtain a comprehensive understanding of the response of
Bradyrhizobium japonicum
to drought. Desiccation of cells resulted in the differential expression of 15 to 20% of the 8,480
B. japonicum
open reading frames, with considerable differentiation between early (after 4 h) and late (after 24 and 72 h) expressed genes. While 225 genes were universally up-regulated at all three incubation times in response to desiccation, an additional 43 and 403 up-regulated genes were common to the 4/24- and 24/72-h incubation times, respectively. Desiccating conditions resulted in the significant induction (>2.0-fold) of the trehalose-6-phosphate synthetase (
otsA
), trehalose-6-phosphate phosphatase (
otsB
), and trehalose synthase (
treS
) genes, which encode two of the three trehalose synthesis pathways found in
B. japonicum
. Gene induction was correlated with an elevated intracellular concentration of trehalose and increased activity of trehalose-6-phosphate synthetase, collectively supporting the hypothesis that this disaccharide plays a prominent and important role in promoting desiccation tolerance in
B. japonicum
. Microarray data also indicated that σ
54
- and σ
24
-associated transcriptional regulators and genes encoding isocitrate lyase, oxidative stress responses, the synthesis and transport of exopolysaccharides, heat shock response proteins, enzymes for the modification and repair of nucleic acids, and the synthesis of pili and flagella are also involved in the response of
B. japonicum
to desiccation. Polyethylene glycol-generated osmotic stress induced significantly fewer genes than those transcriptionally activated by desiccation. However, 67 genes were commonly induced under both conditions. Taken together, these results suggest that
B. japonicum
directly responds to desiccation by adapting to changes imparted by reduced water activity, such as the synthesis of trehalose and polysaccharides and, secondarily, by the induction of a wide variety of proteins involved in protection of the cell membrane, repair of DNA damage, stability and integrity of proteins, and oxidative stress responses.
Anaerobic Regulation of
Shigella flexneri
Virulence: ArcA Regulates
fur
and Iron Acquisition Genes
Authors: Boulette Megan L. , Payne Shelley M. 
Published: July 2007

ABSTRACT

Invasion and plaque formation in epithelial monolayers are routinely used to assess the virulence of
Shigella flexneri
, a causative agent of dysentery. A modified plaque assay was developed to identify factors contributing to the virulence of
S. flexneri
under the anaerobic conditions present in the colon. This assay demonstrated the importance of the ferrous iron transport system Feo, as well as the global transcription factors Fur, ArcA, and Fnr, for
Shigella
plaque formation in anoxic environments. Transcriptional analyses of
S. flexneri
iron transport genes indicated that anaerobic conditions activated
feoABC
while repressing genes encoding two other iron transport systems, the ABC transporter Sit and the Iuc/Iut aerobactin siderophore synthesis and transport system. The anaerobic transcription factors ArcA and Fnr activated expression of
feoABC
, while ArcA repressed
iucABCD iutA
. Transcription of
fur
, encoding the iron-responsive transcriptional repressor of bacterial iron acquisition, was also repressed anaerobically in an ArcA-dependent manner.
Hydroxyatrazine
N
-Ethylaminohydrolase (AtzB): an Amidohydrolase Superfamily Enzyme Catalyzing Deamination and Dechlorination
Authors: Seffernick Jennifer L. , Aleem Asma , Osborne Jeffrey P. , Johnson Gilbert , Sadowsky Michael J. , Wackett Lawrence P. 
Published: July 2007

ABSTRACT

Hydroxyatrazine [2-(
N
-ethylamino)-4-hydroxy-6-(
N
-isopropylamino)-1,3,5-triazine]
N
-ethylaminohydrolase (AtzB) is the sole enzyme known to catalyze the hydrolytic conversion of hydroxyatrazine to
N
-isopropylammelide. AtzB, therefore, serves as the point of intersection of multiple
s
-triazine biodegradative pathways and is completely essential for microbial growth on
s
-triazine herbicides. Here,
atzB
was cloned from
Pseudomonas
sp. strain ADP and its product was purified to homogeneity and characterized. AtzB was found to be dimeric, with subunit and holoenzyme molecular masses of 52 kDa and 105 kDa, respectively. The
k

cat

and
K

m

of AtzB with hydroxyatrazine as a substrate were 3 s
−1
and 20 μM, respectively. Purified AtzB had a 1:1 zinc-to-subunit stoichiometry. Sequence analysis revealed that AtzB contained the conserved mononuclear amidohydrolase superfamily active-site residues His74, His76, His245, Glu248, His280, and Asp331. An intensive in vitro investigation into the substrate specificity of AtzB revealed that 20 of the 51 compounds tested were substrates for AtzB; this allowed for the identification of specific substrate structural features required for catalysis. Substrates required a monohydroxylated
s
-triazine ring with a minimum of one primary or secondary amine substituent and either a chloride or amine leaving group. AtzB catalyzed both deamination and dechlorination reactions with rates within a range of one order of magnitude. This differs from AtzA and TrzN, which do not catalyze deamination reactions, and AtzC, which is not known to catalyze dechlorination reactions.
Alterations in the Two Globular Domains or in the Connecting α-Helix of Bacterial Ribosomal Protein L9 Induces +1 Frameshifts
Authors: Leipuviene Ramune , Björk Glenn R. 
Published: July 2007

ABSTRACT

The ribosomal 50S subunit protein L9, encoded by the gene
rplI
, is an elongated protein with an α-helix connecting the N- and C-terminal globular domains. We isolated
rplI
mutants that suppress the +1 frameshift mutation
hisC3072
in
Salmonella enterica
serovar Typhimurium. These mutants have amino acid substitutions in the N-terminal domain (G24D) or in the C-terminal domain (I94S, A102D, G126V, and F132S) of L9. In addition, different one-base deletions in
rplI
altered either the final portion of the C terminus or removed the C-terminal domain with or without the connecting α-helix. An alanine-to-proline substitution at position 59 (A59P), which breaks the α-helix between the globular domains, induced +1 frameshifting, suggesting that the geometrical relationship between the N and C domains is important to maintain the reading frame. Except for the alterations G126V in the C terminus and A59P in the connecting α-helix, our results confirm earlier results obtained by using the phage T4 gene
60
-based system to monitor bypassing. The way
rplI
mutations suppress various frameshift mutations suggests that bypassing of many codons from several takeoff and landing sites occurred instead of a specific frameshift forward at overlapping codons.
Characterization of the Osmoprotectant Transporter OpuC from
Pseudomonas syringae
and Demonstration that Cystathionine-β-Synthase Domains Are Required for Its Osmoregulatory Function
Authors: Chen Chiliang , Beattie Gwyn A. 
Published: July 2007

ABSTRACT

The plant pathogen
Pseudomonas syringae
may cope with osmotic stress on plants, in part, by importing osmoprotective compounds. In this study, we found that
P. syringae
pv. tomato strain DC3000 was distinct from most bacterial species in deriving greater osmoprotection from exogenous choline than from glycine betaine. This superior osmoprotection was correlated with a higher capacity for uptake of choline than for uptake of glycine betaine. Of four putative osmoregulatory ABC transporters in DC3000, one, designated OpuC, functioned as the primary or sole transporter for glycine betaine and as one of multiple transporters for choline under high osmolarity. Surprisingly, the homolog of the well-characterized ProU transporter from
Escherichia coli
and
Salmonella enterica
serovar Typhimurium did not function in osmoprotection. The
P. syringae
pv. tomato OpuC transporter was more closely related to the
Bacillus subtilis
and
Listeria monocytogenes
OpuC transporters than to known osmoprotectant transporters in gram-negative bacteria based on sequence similarity and genetic arrangement. The
P. syringae
pv. tomato OpuC transporter had a high affinity for glycine betaine, a low affinity for choline, and a broad substrate specificity that included acetylcholine, carnitine, and proline betaine. Tandem cystathionine-β-synthase (CBS) domains in the ATP-binding component of OpuC were required for transporter function. The presence of these CBS domains was correlated with osmoregulatory function among the putative transporters examined in DC3000 and was found to be predictive of functional osmoregulatory transporters in other pseudomonads. These results provide the first functional evaluation of an osmoprotectant transporter in a
Pseudomonas
species and demonstrate the usefulness of the CBS domains as predictors of osmoregulatory activity.
Salicylic Acid, Yersiniabactin, and Pyoverdin Production by the Model PhytopathogenPseudomonas syringaepv. tomato DC3000: Synthesis, Regulation, and Impact on Tomato andArabidopsisHost Plants
Authors: Jones Alexander M. , Lindow Steven E. , Wildermuth Mary C. 
Published: July 2007

ABSTRACTA genetically tractable model plant pathosystem,Pseudomonas syringaepv. tomato DC3000 on tomato andArabidopsis thalianahosts, was used to investigate the role of salicylic acid (SA) and iron acquisition via siderophores in bacterial virulence. Pathogen-induced SA accumulation mediates defense in these plants, and DC3000 contains the genes required for the synthesis of SA, the SA-incorporated siderophore yersiniabactin (Ybt), and the fluorescent siderophore pyoverdin (Pvd). We found that DC3000 synthesizes SA, Ybt, and Pvd under iron-limiting conditions in culture. Synthesis of SA and Ybt by DC3000 requirespchA, an isochorismate synthase gene in the Ybt genomic cluster, and exogenous SA can restore Ybt production by thepchAmutant. Ybt was also produced by DC3000 in planta, suggesting that Ybt plays a role in DC3000 pathogenesis. However, thepchAmutant did not exhibit any growth defect or altered virulence in plants. This lack of phenotype was not attributable to plant-produced SA restoring Ybt production, as thepchAmutant grew similarly to DC3000 in anArabidopsisSA biosynthetic mutant, and in planta Ybt was not detected inpchA-infected wild-type plants. In culture, no growth defect was observed for thepchAmutant versus DC3000 for any condition tested. Instead, enhanced growth of thepchAmutant was observed under stringent iron limitation and additional stresses. This suggests that SA and Ybt production by DC3000 is costly and that Pvd is sufficient for iron acquisition. Further exploration of the comparative synthesis and utility of Ybt versus Pvd production by DC3000 found siderophore-dependent amplification ofybtgene expression to be absent, suggesting that Ybt may play a yet unknown role in DC3000 pathogenesis.
Expression and Association of Group IV Nitrogenase NifD and NifH Homologs in the Non-Nitrogen-Fixing ArchaeonMethanocaldococcus jannaschii
Authors: Staples Christopher R. , Lahiri Surobhi , Raymond Jason , Von Herbulis Lindsay , Mukhophadhyay Biswarup , Blankenship Robert E. 
Published: July 2007

ABSTRACTUsing genomic analysis, researchers previously identified genes coding for proteins homologous to the structural proteins of nitrogenase (J. Raymond, J. L. Siefert, C. R. Staples, and R. E. Blankenship, Mol. Biol. Evol. 21:541-554, 2004). The expression and association of NifD and NifH nitrogenase homologs (named NflD and NflH for “Nif-like” D and H, respectively) have been detected in a non-nitrogen-fixing hyperthermophilic methanogen,Methanocaldococcus jannaschii. These homologs are expressed constitutively and do not appear to be directly involved with nitrogen metabolism or detoxification of compounds such as cyanide or azide. The NflH and NflD proteins were found to interact with each other, as determined by bacterial two-hybrid studies. Upon immunoisolation, NflD and NflH copurified, along with three other proteins whose functions are as yet uncharacterized. The apparent presence of genes coding for NflH and NflD in all known methanogens, their constitutive expression, and their high sequence similarity to the NifH and NifD proteins or the BchL and BchN/BchB proteins suggest that NflH and NflD participate in an indispensable and fundamental function(s) in methanogens.
Mutational Analysis of Active Site Residues Essential for Sensing of Organic Hydroperoxides by
Bacillus subtilis
OhrR
Authors: Soonsanga Sumarin , Fuangthong Mayuree , Helmann John D. 
Published: July 2007

ABSTRACT

Bacillus subtilis
OhrR is the prototype for the one-Cys family of organic peroxide-sensing regulatory proteins. Mutational analyses indicate that the high sensitivity of the active site cysteine (C15) to peroxidation requires three Tyr residues. Y29 and Y40 from the opposing subunit of the functional dimer hydrogen bond with the reactive Cys thiolate, and substitutions at these positions reduce or eliminate the ability of OhrR to respond to organic peroxides. Y19 is also critical for peroxide sensing, and the Ala substitution mutant (OhrR Y19A) is less susceptible to oxidation at the active site C15 in vivo. The Y19A protein also displays decreased sensitivity to peroxide-mediated oxidation in vitro. Y19 is in van der Waals contact with two residues critical for protein function, F16 and R23. The latter residue makes critical contact with the DNA backbone in the OhrR-operator complex. These results indicate that the high sensitivity of the OhrR C15 residue to oxidation requires interactions with the opposed Tyr residues. Oxidative modification of C15 likely disrupts the C15-Y29′-Y40′ hydrogen bond network and thereby initiates conformational changes that reduce the ability of OhrR to bind to its operator site.
Docking of the Periplasmic FecB Binding Protein to the FecCD Transmembrane Proteins in the Ferric Citrate Transport System ofEscherichia coli
Authors: Braun Volkmar , Herrmann Christina 
Published: July 2007

ABSTRACTCitrate-mediated iron transport across the cytoplasmic membrane is catalyzed by an ABC transporter that consists of the periplasmic binding protein FecB, the transmembrane proteins FecC and FecD, and the ATPase FecE. Salt bridges between glutamate residues of the binding protein and arginine residues of the transmembrane proteins are predicted to mediate the positioning of the substrate-loaded binding protein on the transmembrane protein, based on the crystal structures of the ABC transporter for vitamin B12, consisting of the BtuF binding protein and the BtuCD transmembrane proteins (E. L. Borths et al., Proc. Natl. Acad. Sci. USA 99:16642-16647, 2002). Here, we examined the role of the residues predicted to be involved in salt-bridge formation between FecB and FecCD by substituting these residues with alanine, cysteine, arginine, and glutamate and by analyzing the citrate-mediated iron transport of the mutants. Replacement of E93 in FecB with alanine [FecB(E93A)], cysteine, or arginine nearly abolished citrate-mediated iron transport. Mutation FecB(E222R) nearly eliminated transport, and FecB(E222A) and FecB(E222C) strongly reduced transport. FecD(R54C) and FecD(R51E) abolished transport, whereas other R-to-C mutations in putative interaction sites between FecCD and FecB substantially reduced transport. The introduced cysteine residues in FecB and FecCD also served to examine the formation of disulfide bridges in place of salt bridges between the binding protein and the transmembrane proteins. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis results suggest cross-linking of FecB(E93C) to FecD(R54C) and FecB(E222C) to FecC(R60C). The data are consistent with the proposal that FecB(E93) is contained in the region that binds to FecD and FecB(E222) in the region that binds to FecC.