bluuebunny/crossref_metadata_2025 huggingface dataset:
An Expression-Driven Approach to the Prediction of Carbohydrate Transport and Utilization Regulons in theHyperthermophilic BacteriumThermotoga maritima
Authors: Conners Shannon B. , Montero Clemente I. , Comfort Donald A. , Shockley Keith R. , Johnson Matthew R. , Chhabra Swapnil R. , Kelly Robert M.
Published: October 2005

Comprehensive analysis of genome-wide expression patterns during growth of the hyperthermophilic bacteriumThermotoga maritimaon 14 monosaccharide and polysaccharide substrates was undertaken with the goal of proposing carbohydrate specificities for transport systems and putative transcriptional regulators. Saccharide-induced regulons were predicted through the complementary use of comparative genomics, mixed-model analysis of genome-wide microarray expression data, and examination of upstream sequence patterns. The results indicate thatT. maritimarelies extensively on ABC transporters for carbohydrate uptake, many of which are likely controlled by local regulators responsive to either the transport substrate or a key metabolic degradation product. Roles in uptake of specific carbohydrates were suggested for members of the expanded Opp/Dpp family of ABC transporters. In this family, phylogenetic relationships among transport systems revealed patterns of possible duplication and divergence as a strategy for the evolution of new uptake capabilities. The presence of GC-rich hairpin sequences between substrate-binding proteins and other components of Opp/Dpp family transporters offers a possible explanation for differential regulation of transporter subunit genes. Numerous improvements toT. maritimagenome annotations were proposed, including the identification of ABC transport systems originally annotated as oligopeptide transporters as candidate transporters for rhamnose, xylose, β-xylan, andβ -glucans and identification of genes likely to encode proteins missing from current annotations of the pentose phosphate pathway. Beyond the information obtained forT. maritima, the present study illustrates how expression-based strategies can be used for improving genome annotation in other microorganisms, especially those for which genetic systems are unavailable.
A Mycothiol Synthase Mutant of
Mycobacterium smegmatis
Produces Novel Thiols and Has an Altered Thiol Redox Status
Authors: Newton Gerald L. , Ta Philong , Fahey Robert C.
Published: October 2005

Mycobacteria and other actinomycetes do not produce glutathione but make mycothiol (MSH; AcCys-GlcN-Ins) that has functions similar to those of glutathione and is essential for growth of
Mycobacterium tuberculosis
. Mycothiol synthase (MshD) catalyzes N acetylation of Cys-GlcN-Ins to produce MSH in
Mycobacterium smegmatis
mc
2
155, and Cys-GlcN-Ins is maintained at a low level. The mycothiol synthase mutant, the
mshD
::Tn
5
mutant, produces high levels of Cys-GlcN-Ins along with two novel thiols,
N
-formyl-Cys-GlcN-Ins and
N
-succinyl-Cys-GlcN-Ins, and a small amount of MSH. The nonenzymatic reaction of acyl-coenzyme A (CoA) with Cys-GlcN-Ins to produce acyl-Cys-GlcN-Ins is a facile reaction under physiologic conditions, with succinyl-CoA being an order of magnitude more reactive than acetyl-CoA. The uncatalyzed reaction rates are adequate to account for the observed production of
N
-succinyl-Cys-GlcN-Ins and MSH under physiologic conditions. It was shown that the
N
-acyl-Cys-GlcN-Ins compounds are maintained in a substantially reduced state in the mutant but that Cys-GlcN-Ins exists in disulfide forms at 5 to 40% at different stages of growth. MSH was able to facilitate reduction of
N
-succinyl-Cys-GlcN-Ins disulfide through thiol-disulfide exchange, but
N
-formyl-Cys-GlcN-Ins was ineffective. The oxidized state of Cys-GlcN-Ins in cells appears to result from a high susceptibility to autoxidation and a low capacity of the cell to reduce its disulfide forms. The mutant exhibited no enhanced sensitivity to hydrogen peroxide,
tert
-butyl hydroperoxide, or cumene hydroperoxide relative to the parent strain, suggesting that the most abundant thiol,
N
-formyl-Cys-GlcN-Ins, functions as a substitute for MSH.
Genetic and Functional Characterization of theEscherichia coliBarA-UvrY Two-Component System: Point Mutations in the HAMP Linker of the BarA Sensor Give a Dominant-Negative Phenotype
Authors: Tomenius Henrik , Pernestig Anna-Karin , Méndez-Catalá Claudia F. , Georgellis Dimitris , Normark Staffan , Melefors Öjar
Published: October 2005

The BarA-UvrY two-component system family is strongly associated with virulence but is poorly understood at the molecular level. During our attempts to complement abarAdeletion mutant, we consistently generated various mutated BarA proteins. We reasoned that characterization of the mutants would help us to better understand the signal transduction mechanism in tripartite sensors. This was aided by the demonstrated ability to activate the UvrY regulator with acetyl phosphate independently of the BarA sensor. Many of the mutated BarA proteins had poor complementation activity but could counteract the activity of the wild-type sensor in a dominant-negative fashion. These proteins carried point mutations in or near the recently identified HAMP linker, previously implicated in signal transduction between the periplasm and cytoplasm. This created sensor proteins with an impaired kinase activity and a net dephosphorylating activity. Using further site-directed mutagenesis of a HAMP linker-mutated protein, we could demonstrate that the phosphoaccepting aspartate 718 and histidine 861 are crucial for the dephosphorylating activity. Additional analysis of the HAMP linker-mutated BarA sensors demonstrated that a dephosphorylating activity can operate via phosphotransfer within a tripartite sensor dimer in vivo. This also means that a tripartite sensor can be arranged as a dimer even in the dephosphorylating mode.
Gene Cloning and Molecular Characterization of an Extracellular Poly(l-Lactic Acid) Depolymerase fromAmycolatopsissp. Strain K104-1
Authors: Matsuda Emiko , Abe Naoki , Tamakawa Hideyuki , Kaneko Jun , Kamio Yoshiyuki
Published: October 2005

We have isolated a polylactide or poly(l-lactic acid) (PLA)-degrading bacterium,Amycolatopsissp. strain K104-1, and purified PLA depolymerase (PLD) from the culture fluid of the bacterium. Here, we cloned and expressed thepldgene encoding PLD inStreptomyces lividans1326 and characterized a recombinant PLD (rPLD) preparation. We also describe the processing mechanism from nascent PLD to mature PLD. Thepldgene encodes PLD as a 24,225-Da polypeptide consisting of 238 amino acids. Biochemical and Western immunoblot analyses of PLD and its precursors revealed that PLD is synthesized as a precursor (prepro-type), requiring proteolytic cleavage of the N-terminal 35-amino-acid extension including the 26-amino-acid signal sequence and 9-residue prosequence to generate the mature enzyme of 20,904 Da. The cleavage of the prosequence was found to be autocatalytic. PLD showed about 45% similarity to many eukaryotic serine proteases. In addition, three amino acid residues, H57, D102, and S195(chymotrypsin numbering), which are implicated in forming the catalytic triad necessary for cleavage of amide bond of substrates in eukaryotic serine proteases, were conserved in PLD as residues H74, D111, and S197. The G193residue (chymotrypsin numbering), which is implicated in forming an oxyanion hole with residue S195and forms an important hydrogen bond for interaction with the carbonyl group of the scissile peptide bond, was also conserved in PLD. The functional analysis of the PLD mutants H74A, D111A, and S197A revealed that residues H74, D111, and S197are important for the depolymerase and caseinolytic activities of PLD and for cleavage of the prosequence from pro-type PLD to form the mature one. The PLD preparation had elastase activity which was not inhibited by 1 mM elastatinal, which is 10 times higher than needed for complete inhibition of porcine pancreatic elastase. The rPLD preparation degraded PLA with an average molecular mass of 220 kDa into lactic acid dimers through lactic acid oligomers and finally into lactic acid. The PLD preparation bound to high polymers of 3-hydoxybutyrate, ε-caprolacton, and butylene succinate as well as PLA, but it degraded only PLA.
Rhamnolipids Modulate Swarming Motility Patterns ofPseudomonas aeruginosa
Authors: Caiazza Nicky C. , Shanks Robert M. Q. , O'Toole G. A.
Published: October 2005

Pseudomonas aeruginosais capable of twitching, swimming, and swarming motility. The latter form of translocation occurs on semisolid surfaces, requires functional flagella and biosurfactant production, and results in complex motility patterns. From the point of inoculation, bacteria migrate as defined groups, referred to as tendrils, moving in a coordinated manner capable of sensing and responding to other groups of cells. We were able to show thatP. aeruginosaproduces extracellular factors capable of modulating tendril movement, and genetic analysis revealed that modulation of these movements was dependent on rhamnolipid biosynthesis. AnrhlBmutant (deficient in mono- and dirhamnolipid production) and anrhlCmutant (deficient in dirhamnolipid production) exhibited altered swarming patterns characterized by irregularly shaped tendrils. In addition, agar supplemented with rhamnolipid-containing spent supernatant inhibited wild-type (WT) swarming, whereas agar supplemented with spent supernatant from mutants that do not make rhamnolipids had no effect on WTP. aeruginosaswarming. Addition of purified rhamnolipids to swarming medium also inhibited swarming motility of the WT strain. We also show that asadBmutant does not sense and/or respond to other groups of swarming cells and this mutant was capable of swarming on media supplemented with rhamnolipid-containing spent supernatant or purified rhamnolipids. The abilities to produce and respond to rhamnolipids in the context of group behavior are discussed.
Role of the Nfo (YqfS) and ExoA Apurinic/Apyrimidinic Endonucleases in Protecting
Bacillus subtilis
Spores from DNA Damage
Authors: Salas-Pacheco José M. , Setlow Barbara , Setlow Peter , Pedraza-Reyes Mario
Published: October 2005

The
Bacillus subtilis
enzymes ExoA and Nfo (originally termed YqfS) are endonucleases that can repair apurinic/apyrimidinic (AP) sites and strand breaks in DNA. We have analyzed how the lack of ExoA and Nfo affects the resistance of growing cells and dormant spores of
B. subtilis
to a variety of treatments, some of which generate AP sites and DNA strand breaks. The lack of ExoA and Nfo sensitized spores (termed α
−
β
−
) lacking the majority of their DNA-protective α/β-type small, acid-soluble spore proteins (SASP) to wet heat. However, the lack of these enzymes had no effect on the wet-heat resistance of spores that retained α/β-type SASP. The lack of either ExoA or Nfo sensitized wild-type spores to dry heat, but loss of both proteins was necessary to sensitize α
−
β
−
spores to dry heat. The lack of ExoA and Nfo also sensitized α
−
β
−
, but not wild-type, spores to desiccation. In contrast, loss of ExoA and Nfo did not sensitize growing cells or wild-type or α
−
β
−
spores to hydrogen peroxide or
t
-butylhydroperoxide. Loss of ExoA and Nfo also did not increase the spontaneous mutation frequency of growing cells.
exoA
expression took place not only in growing cells, but also in the forespore compartment of the sporulating cell. These results, together with those from previous work, suggest that ExoA and Nfo are additional factors that protect
B. subtilis
spores from DNA damage accumulated during spore dormancy.
Expression Dynamics of Arsenic Respiration and Detoxification in
Shewanella
sp. Strain ANA-3
Authors: Saltikov Chad W. , Wildman Richard A. , Newman Dianne K.
Published: October 2005

Because arsenate [As(V)] reduction by bacteria can significantly enhance arsenic mobility in the environment, it is important to be able to predict when this activity will occur. Currently, two bacterial systems are known that specifically reduce As(V), namely, a respiratory system (encoded by the
arr
genes) and a detoxification system (encoded by the
ars
genes). Here we analyze the conditions under which these two systems are expressed in
Shewanella
sp. strain ANA-3. The
ars
system is expressed under both aerobic and anaerobic conditions, whereas the
arr
system is only expressed anaerobically and is repressed by oxygen and nitrate. When cells are grown on As(V), the
arr
system is maximally induced during exponential growth, with peak expression of the
ars
system occurring at the beginning of stationary phase. Both the
arr
and
ars
systems are specifically induced by arsenite [As(III)], but the
arr
system is activated by a concentration of As(III) that is 1,000 times lower than that required for the
arsC
system (≤100 nM versus ≤100 μM, respectively). A double mutant was constructed that does not reduce As(V) under any growth conditions. In this strain background, As(V) is capable of inducing the
arr
system at low micromolar concentrations, but it does not induce the
ars
system. Collectively, these results demonstrate that the two As(V) reductase systems in ANA-3 respond to different amounts and types of inorganic arsenic.
Bile Salts Modulate Expression of the CmeABC Multidrug Efflux Pump inCampylobacter jejuni
Authors: Lin Jun , Cagliero Cédric , Guo Baoqing , Barton Yi-Wen , Maurel Marie-Christine , Payot Sophie , Zhang Qijing
Published: October 2005

CmeABC, a multidrug efflux pump, is involved in the resistance ofCampylobacter jejunito a broad spectrum of antimicrobial agents and is essential forCampylobactercolonization in animal intestine by mediating bile resistance. Previously, we have shown that expression of this efflux pump is under the control of a transcriptional repressor named CmeR. Inactivation of CmeR or mutation in thecmeABCpromoter (PcmeABC) region derepressescmeABC, leading to overexpression of this efflux pump. However, it is unknown if the expression ofcmeABCcan be conditionally induced by the substrates it extrudes. In this study, we examined the expression ofcmeABCin the presence of various antimicrobial compounds. Although the majority of the antimicrobials tested did not affect the expression ofcmeABC, bile salts drastically elevated the expression of this efflux operon. The induction was observed with both conjugated and unconjugated bile salts and was in a dose- and time-dependent manner. Experiments using surface plasmon resonance demonstrated that bile salts inhibited the binding of CmeR to PcmeABC, suggesting that bile compounds are inducing ligands of CmeR. The interaction between bile salts and CmeR likely triggers conformational changes in CmeR, resulting in reduced binding affinity of CmeR to PcmeABC. Bile did not affect the transcription ofcmeR, indicating that altered expression ofcmeRis not a factor in bile-induced overexpression ofcmeABC. In addition to the CmeR-dependent induction, some bile salts (e.g., taurocholate) also activated the expression ofcmeABCby a CmeR-independent pathway. Consistent with the elevated production of CmeABC, the presence of bile salts in culture media resulted in increased resistance ofCampylobacterto multiple antimicrobials. These findings reveal a new mechanism that modulates the expression ofcmeABCand further support the notion that bile resistance is a natural function of CmeABC.
CpsE from Type 2Streptococcus pneumoniaeCatalyzes the Reversible Addition of Glucose-1-Phosphate to a Polyprenyl Phosphate Acceptor, Initiating Type 2 Capsule Repeat Unit Formation
Authors: Cartee Robert T. , Forsee W. Thomas , Bender Matthew H. , Ambrose Karita D. , Yother Janet
Published: October 2005

The majority of the 90 capsule types made by the gram-positive pathogenStreptococcus pneumoniaeare assembled by a block-type mechanism similar to that utilized by the Wzy-dependent O antigens and capsules of gram-negative bacteria. In this mechanism, initiation of repeat unit formation occurs by the transfer of a sugar to a lipid acceptor. InS. pneumoniae, this step is catalyzed by CpsE, a protein conserved among the majority of capsule types. Membranes fromS. pneumoniaetype 2 strain D39 andEscherichia colicontaining recombinant Cps2E catalyzed incorporation of [14C]Glc from UDP-[14C]Glc into a lipid fraction in a Cps2E-dependent manner. The Cps2E-dependent glycolipid product from both membranes was sensitive to mild acid hydrolysis, suggesting that Cps2E was catalyzing the formation of a polyprenyl pyrophosphate Glc. Addition of exogenous polyprenyl phosphates ranging in size from 35 to 105 carbons to D39 andE. colimembranes stimulated Cps2E activity. The stimulation was due, in part, to utilization of the exogenous polyprenyl phosphates as an acceptor. The glycolipid product synthesized in the absence of exogenous polyprenyl phosphates comigrated with a 60-carbon polyprenyl pyrophosphate Glc. When 10 or 100 μM UMP was added to reaction mixtures containing D39 membranes, Cps2E activity was inhibited 40% and 80%, respectively. UMP, which acted as a competitive inhibitor of UDP-Glc, also stimulated Cps2E to catalyze the reverse reaction, with synthesis of UDP-Glc from the polyprenyl pyrophosphate Glc. These data indicated that Cps2E was catalyzing the addition of Glc-1-P to a polyprenyl phosphate acceptor, likely undecaprenyl phosphate.
Regulation of thepspAVirulence Factor and EssentialpcsBMurein Biosynthetic Genes by the Phosphorylated VicR (YycF) Response Regulator inStreptococcus pneumoniae
Authors: Ng Wai-Leung , Tsui Ho-Ching Tiffany , Winkler Malcolm E.
Published: October 2005

The VicRK (YycFG) two-component regulatory system (TCS) is required for virulence of the human respiratory pathogenStreptococcus pneumoniae(pneumococcus). The VicR (YycF) response regulator (RR) is essential through its positive regulation ofpcsB, which encodes an extracellular protein that mediates murein biosynthesis. To determine other genes that are regulated by VicR, we performed microarray analyses on a unique ΔvicRdeletion mutant, which was constructed by uncoupling regulation ofpcsB. Results from these microarray experiments support the idea that the VicR RR exerts strong positive regulation on the transcription of a set of genes encoding important surface proteins, including the PspA virulence factor, two proteins (Spr0096 and Spr1875) containing LysM peptidoglycan-binding domains, and a putative membrane protein (Spr0709) of unknown function. To demonstrate direct regulation, we performed band shift and footprinting experiments using purified unphosphorylated VicR and phosphorylated VicR-P, which was prepared by reaction with acetyl phosphate. VicR and VicR-P bound to regions upstream ofpcsB,pspA,spr0096,spr1875, andspr0709.Phosphorylation of VicR to VicR-P increased the apparent strength and changed the nature of binding to these regions. DNase I footprinting of VicR and VicR-P bound to regions upstream ofpcsB,pspA,spr0096, andspr1875showed protection of extended regions containing a degenerate sequence related to a previously proposed consensus. These combined approaches did not support autoregulation of thevicRKXoperon or substantive direct regulation of fatty acid biosynthesis by VicR or VicR-P. However, the ΔvicRmutant required fatty acids in some conditions, which supports the notion that the VicRK TCS may mediate membrane integrity as well as murein biosynthesis and virulence factor expression inS. pneumoniae.
Nocturnal Production of Endospores in Natural Populations ofEpulopiscium-Like Surgeonfish Symbionts
Authors: Flint Joseph F. , Drzymalski Dan , Montgomery W. Linn , Southam Gordon , Angert Esther R.
Published: October 2005

Prior studies have described a morphologically diverse group of intestinal microorganisms associated with surgeonfish. Despite their diversity of form, 16S rRNA gene surveys and fluorescent in situ hybridizations indicate that these bacteria are low-G+C gram-positive bacteria related toEpulopisciumspp. Many of these bacteria exhibit an unusual mode of reproduction, developing multiple offspring intracellularly. Previous reports have suggested that someEpulopiscium-like symbionts produce dormant or phase-bright intracellular offspring. Close relatives ofEpulopiscium, such asMetabacterium polysporaandClostridium lentocellum, are endospore-forming bacteria, which raises the possibility that the phase-bright offspring are endospores. Structural evidence and the presence of dipicolinic acid demonstrate that phase-bright offspring ofEpulopiscium-like bacteria are true endospores. In addition, endospores are formed as part of the normal daily life cycle of these bacteria. In the populations studied, mature endospores were seen only at night and the majority of cells in a given population produced one or two endospores per mother cell. Phylogenetic analyses confirmed the close relationship between the endospore-forming surgeonfish symbionts characterized here and previously describedEpulopisciumspp. The broad distribution of endospore formation among theEpulopisciumphylogenetic group raises the possibility that sporulation is a characteristic of the group. We speculate that spore formation inEpulopiscium-like symbionts may be important for dispersal and may also enhance survival in the changing conditions of the fish intestinal tract.
Mutations in HlyD, Part of the Type 1 Translocator for Hemolysin Secretion, Affect the Folding of the Secreted Toxin
Authors: Pimenta A. L. , Racher K. , Jamieson L. , Blight M. A. , Holland I. B.
Published: October 2005

HlyD, a member of the membrane fusion protein family, is essential for the secretion of the RTX hemolytic toxin HlyA from
Escherichia coli
. Random point mutations affecting HlyA secretion were obtained, distributed in most periplasmic regions of the HlyD molecule. Analysis of the secretion phenotypes of different mutants allowed the identification of regions in HlyD involved in different steps of HlyA translocation. Four mutants, V349-I, T85-I, V334-I and L165-Q, were conditionally defective, a phenotype shown to be linked to the presence of inhibitory concentrations of Ca
2+
in extracellular medium. Hly mutant T85-I was defective at an early stage in secretion, while mutants V334-I and L165-Q appeared to accumulate HlyA in the cell envelope, indicating a block at an intermediate step. Mutants V349-I, V334-I, and L165-Q were only partially defective in secretion, allowing significant levels of HlyA to be transported, but in the case of V349-I and L165-Q the HlyA molecules secreted showed greatly reduced hemolytic activity. Hemolysin molecules secreted from V349-I and V334-I are defective in normal folding and can be reactivated in vitro to the same levels as HlyA secreted from the wild-type translocator. Both V349-I and V334-I mutations mapped to the C-terminal lipoyl repeat motif, involved in the switching from the helical hairpin to the extended form of HlyD during assembly of the functional transport channel. These results suggest that HlyD is an integral component of the transport pathway, whose integrity is essential for the final folding of secreted HlyA into its active form.
Metabolic and Evolutionary Relationships among
Pyrococcus
Species: Genetic Exchange within a Hydrothermal Vent Environment
Authors: Hamilton-Brehm Scott D. , Schut Gerrit J. , Adams Michael W. W.
Published: October 2005

Pyrococcus furiosus
and
Pyrococcus woesei
grow optimally at temperatures near 100°C and were isolated from the same shallow marine volcanic vent system. Hybridization of genomic DNA from
P. woesei
to a DNA microarray containing all 2,065 open reading frames (ORFs) annotated in the
P. furiosus
genome, in combination with PCR analysis, indicated that homologs of 105 ORFs present in
P. furiosus
are absent from the uncharacterized genome of
P. woesei
. Pulsed-field electrophoresis indicated that the sizes of the two genomes are comparable, and the results were consistent with the hypothesis that
P. woesei
lacks the 105 ORFs found in
P. furiosus
. The missing ORFs are present in
P. furiosus
mainly in clusters. These clusters include one cluster (Mal I, PF1737 to PF1751) involved in maltose metabolism and another cluster (PF0691 to PF0695) whose products are thought to remove toxic reactive nitrogen species. Accordingly, it was found that
P. woesei
, in contrast to
P. furiosus
, is unable to utilize maltose as a carbon source for growth, and the growth of
P. woesei
on starch was inhibited by addition of a nitric oxide generator. In
P. furiosus
the ORF clusters not present in
P. woesei
are bracketed by or are in the vicinity of insertion sequences or long clusters of tandem repeats (LCTRs). While the role of LCTRs in lateral gene transfer is not known, the Mal I cluster in
P. furiosus
is a composite transposon that undergoes replicative transposition. The same locus in
P. woesei
lacks any evidence of insertion activity, indicating that
P. woesei
is a sister or even the parent of
P. furiosus. P. woesei
may have acquired by lateral gene transfer more than 100 ORFs from other organisms living in the same thermophilic environment to produce the type strain of
P. furiosus
.
QscR-Mediated Transcriptional Activation of Serine Cycle Genes in
Methylobacterium extorquens
AM1
Authors: Kalyuzhnaya Marina G. , Lidstrom Mary E.
Published: October 2005

QscR, a LysR-type regulator, is the major regulator of assimilatory C
1
metabolism in
Methylobacterium extorquens
AM1. It has been shown to interact with the promoters of the two operons that encode the majority of the serine cycle enzymes (
sga-hpr-mtdA-fch
for the
qsc
1 operon and
mtkA-mtkB-ppc-mclA
for the
qsc
2 operon), as well as with the promoter of
glyA
and its own promoter. To obtain further insights into the mechanisms of this regulation, we mapped transcriptional start sites for the
qsc1
and
qsc2
operons and for
glyA
via primer extension analysis. We also identified the specific binding sites for QscR upstream of the
qsc1
and
qsc2
operons and
glyA
by DNase I footprinting. The QscR protected areas were located at nucleotides −216 to −165, nucleotides −59 to −26, and nucleotides −72 to −39 within the promoter-regulatory regions upstream of transcriptional starts of, respectively,
qsc1
,
qsc2
and
glyA
. To examine the nature of the metabolic signal that may influence QscR-mediated regulation of the serine cycle genes,
Pqsc1
::
xylE
translational fusions were constructed and expression of XylE monitored in the wild-type strain, as well as in knockout mutants defective in a variety of methylotrophy functions. The data from these experiments pointed toward formyl-H
4
F being a coinducer of QscR and possibly the major signal in the regulation of the serine cycle in
M. extorquens
AM1. The ability of formyl-H
4
F to enhance the binding of QscR to a specific region upstream of one of the serine cycle operons was demonstrated in gel retardation experiments.
Chlamydial GroEL Autoregulates Its Own Expression through Direct Interactions with the HrcA Repressor Protein
Authors: Wilson Adam C. , Wu Christine C. , Yates John R. , Tan Ming
Published: October 2005

In the pathogenic bacterium
Chlamydia trachomatis
, a transcriptional repressor, HrcA, regulates the major heat shock operons,
dnaK
and
groE
. Cellular stress causes a transient increase in transcription of these heat shock operons through relief of HrcA-mediated repression, but the pathway leading to derepression is unclear. Elevated temperature alone is not sufficient, and it is hypothesized that additional chlamydial factors play a role. We used DNA affinity chromatography to purify proteins that interact with HrcA in vivo and identified a higher-order complex consisting of HrcA, GroEL, and GroES. This endogenous HrcA complex migrated differently than recombinant HrcA, but the complex could be disrupted, releasing native HrcA that resembled recombinant HrcA. In in vitro assays, GroEL increased the ability of HrcA to bind to the CIRCE operator and to repress transcription. Other chlamydial heat shock proteins, including the two additional GroEL paralogs present in all chlamydial species, did not modulate HrcA activity.
Nouvelle méthode de suivi au champ de la sénescence des feuilles de maïs
Authors: Combe Laurette
Published: October 2005

A new method was developed to describe and quantify leaf senescence in a field-grown maize canopy with the purpose of future crop modelling. Leaf shape was analysed using the outlines of over 500 adult leaves from plants grown under diverse culture conditions. Leaf width (λ) taken at regular distances from the ligule (L) allowed the development of leaf-shape equations. These are third-degree polynomials. Thus, four leaf measurements are needed for characterizing leaf shape: maximum width (λm), total length (Lt), width at ligule level (λo), and distance from ligule to the point of maximum width (Lx). Assuming y = (λ/λm) and x = (L/Lt), the shape equation can be written as y = (1  x)(ax2 + bx + yo). Coefficients a and b were estimated from (λo/λm) = yo and (Lx/Lt) = xm. The leaf area between the ligule and any point on the midrib can be obtained by analytically integrating the leaf shape equation. In the present work, degreening was used as the indicator of senescence. Thus, once the shape equation was been defined, easily taken measurements of “green colour” allowed me to calculate the remaining green area by leaf. The vertical profile of the green area on the whole plant could then be described. The time course of green leaf area in a field-grown maize canopy was analysed from silking up to harvest. The farther the leaves were from the ear and the lower they were in the plant, the earlier they lost their green colour. The smallest leaves fully changed colour in 1 week, while this process took 2 weeks for the largest ones. A whole plant lost its green colour in about 5 weeks. Drought caused earlier but not faster leaf degreening. Low plant densities delayed discoloration of the leaves close to the ear, even under drought conditions. The start of senescence in leaves 716 was almost simultaneous in all crop conditions, but it could not be determined whether this was due to a threshold effect of the source/sink ratio or to another factor. This method was easy to set up for field studies with maize, but it could be also used for studies on other graminaceous species and some broad-leaved species. It could be extended to describe the physiological functioning over the leaf length and across the vertical profile of the whole plant. Key words: leaf shape, leaf area, leaf senescence, leaf color, maize, plant density, drought.
The relationships among lodging, stem anatomy, degree of lignification, and resistance to mycosphaerella blight in field pea (Pisum sativum)
Authors: Banniza S , Hashemi P , Warkentin T D , Vandenberg A , Davis A R
Published: October 2005

The relationships among lodging, stem anatomy, degree of stem lignification, and resistance to mycosphaerella blight (Mycosphaerella pinodes (Berk. & Blox.) Vestergr.) in field pea (Pisum sativum L.) were investigated in field experiments in Saskatchewan from 2000 to 2002. Disease severity, lodging resistance, plant height, internode length and diameter, and the cellulose, lignin, and fibre contents of stems were measured from 9 (2000) and 20 (2001, 2002) cultivars of field pea. Significant differences were observed in resistance to mycosphaerella blight and lodging among cultivars. In all years, ‘AC Tamor’ was among those cultivars with the highest disease ratings and lodging scores, whereas ‘CDC Striker’, ‘Carneval’, and ‘Integra’ were among those with the lowest ratings. Disease severity was positively correlated with lodging scores of plants. Lodging was negatively correlated with the proportions of supportive tissue (formerly parenchyma cells in the pith periphery, which have differentiated to sclerenchyma) and xylem in stem sections, whereas disease ratings were negatively correlated with xylem. Lignin and fibre contents in pea stems were negatively correlated with both lodging and mycosphaerella blight severity. Cellulose content was also negatively correlated with lodging scores. This study establishes an anatomical and biochemical basis for further research into pea stem lodging and disease resistance. Key words: cellulose, fibre, lignin, sclerenchyma, supportive tissue, xylem.
Postglacial record of diatom assemblage changes related to climate in an alpine lake in the northern Rocky Mountains, Canada
Authors: Karst-Riddoch Tammy L , Pisaric Michael FJ , Youngblut Donald K , Smol John P
Published: October 2005

We relate changes in diatom assemblages to Holocene climate since ca. 10 300 cal. years BP at a small (4 ha), alkaline, alpine tundra lake in the Rocky Mountains of northeast British Columbia. A complex of small benthic Fragilaria-type taxa generally dominated the fossil diatom communities reflecting shallow, alkaline, and likely low light conditions resulting from minerogenic turbidity or extensive ice cover throughout the Holocene. Shifts in diatom assemblages occurred with changes in local vegetation, inferred from pollen and stomate records, and shifts in charcoal-inferred fire frequency (e.g., lowering of tree line and reduced fire frequency (approx. 7500 cal. years BP) and establishment of modern alpine tundra (approx. 3500 cal. years BP). Importantly, striking changes in diatom communities (e.g., during the mid-Holocene (approx. 5500 to approx. 3500 cal. years BP) and Medieval Warm Period (approx. AD 800 to approx. AD 1250)) occurred without evidence of vegetation shifts. During these times, diverse periphytic diatoms dominated suggesting improved benthic habitat availability with longer growing seasons and probably reduced alkalinity and turbidity with warmer- and wetter-than-present climates. Diatom analysis from this alpine lake provides a sensitive record of climate-related limnological responses, but also refines our previous understanding of Holocene climate changes in the northern Rocky Mountains inferred from terrestrial-based paleoecological records.Key words: alkalinity, alpine lakes, climate change, diatoms, Holocene, paleolimnology.
Structural characteristics of rootfungal interactions for five ericaceous species in eastern Canada
Authors: Massicotte H B , Melville L H , Peterson R L
Published: October 2005

A combination of light microscopy (including differential interference contrast) and laser scanning confocal microscopy was used to document the colonization patterns of epidermal cells and details of intracellular hyphal complexes of five native ericaceous hosts: Vaccinium oxycoccos L. (bog cranberry), Ledum groenlandicum Oeder. (Labrador tea), Vaccinium myrtilloides L. (velvet-leaf blueberry), Kalmia angustifolia L. (sheep laurel), and Gaultheria procumbens L. (wintergreen). Colonization patterns, hyphal complex morphology, and the structure of thick-walled epidermal cells varied considerably among hosts. Multiple hyphal connections were observed between adjacent epidermal cells, indicating that one fungal entry point may result in the colonization of more than one epidermal cell. Further field observations combined with fungal isolations from field-collected plants, identification, and reinoculation studies of other species in the large Ericaceae family are required to determine the full range of structural details in ericoid mycorrhizas.Key words: ericoid mycorrhizas, hair roots, intracellular hyphal complexes, confocal microscopy.
Mutations of corolla symmetry affect carpel and stamen development inHelianthus annuus
Authors: Berti Fabio , Fambrini Marco , Turi Maurizio , Bertini Daniele , Pugliesi Claudio
Published: October 2005

The inflorescence of sunflower (Helianthus annuus L.) is heterogamous with zygomorphic ray flowers located in the outermost whorl of the head and actinomorphic disk flowers arrayed in arcs radiating from the center of the head. Two mutants with altered corolla symmetry have been described. The Chrysanthemoides (Chry) mutant is characterized by a shift from the polysymmetric corolla of disk flowers into a monosymmetric ray-like corolla. The tubular ray flower (turf) mutant is characterized by a shift from the zygomorphic corolla of ray flowers into a nearly actinomorphic tubular-like corolla. We performed a genetic analysis of turf, showing that a single nuclear recessive gene controls the trait. Furthermore, we characterized in detail the morphological floral features of Chry and turf, demonstrating that both mutations also affect the development of stamens and carpels. Most disk flowers found in the peripheral whorls of Chry heads showed drastic reduction in stamen length, as well as absence of ovules, and developed an unbranched style. By contrast, tubular-like ray flowers of turf achieved the ability to differentiate both fertile stamens and ovules. Homeotic transformations were also identified in the tubular-like ray flowers of turf, affecting both filaments and anthers that displayed petaloid-like traits. Our results point to a primary role for TURF and CHRY in the programming of the corolla symmetry and suggest a key interaction of both genes with floral organ identity genes.Key words: Helianthus annuus, floral symmetry, floral mutants, actinomorphic flowers, zygomorphic flowers.
Age, stand density, and tree size as factors in root and basal grafting of lodgepole pine
Authors: Fraser Erin C , Lieffers Victor J , Landhäusser Simon M
Published: October 2005

This study investigated stand factors associated with the rate of root graft formation in lodgepole pine stands. Forty plot areas, each containing 10 trees, were excavated in pure, even-aged pine stands in western Alberta. Exposed root systems were examined for grafts and various stand measurements were recorded at each plot. Results indicate that the number of grafts per square metre is controlled by plot tree density and tree diameter. Also, the percentage of grafted trees increased with both increasing tree age and decreasing distance between trees. Grafts also appear to form relatively early in stand development; the majority of grafts in the present study had formed by the time roots were 20 years old and 50 mm in diameter. These results suggest that grafting is a common occurrence in lodgepole pine stands where trees are <109 cm apart, which translates to a density of approximately 8500 stems/ha (based on even tree distribution). However, even clumps within relatively low density stands are likely to be grafted from a relatively early stage of development.Key words: Pinus contorta, graft formation, stand dynamics.
Cytokinin accumulation and an altered ethylene response mediate the pleiotropic phenotype of the pea nodulation mutant R50 (sym16)
Authors: Ferguson Brett J , Wiebe Ericka M , Neil Emery R J , Guinel Frédérique C
Published: October 2005

R50 (sym16), a pleiotropic mutant of Pisum sativum L., is short, has thickened internodes and roots, and has a reduced number of lateral roots and nodules. Its low nodule phenotype can be restored with the application of ethylene inhibitors; furthermore, it can be mimicked by applying cytokinins (CKs) to the roots of the parent line ‘Sparkle’. Here, we report on the etiolation phenotypes of R50 and ‘Sparkle’, and on the interactive roles of ethylene and CKs in these lines. R50 displayed an altered etiolation phenotype, as it was shorter and thicker, and had more developed leaves than dark-grown ‘Sparkle’. Shoot morphological differences induced by exogenous ethylene or CKs were found to be less severe for R50. Ethylene inhibitor application induced root and shoot elongation and encouraged apical hook opening in both etiolated lines. Liquid chromatography - tandem mass spectrometry analysis indicated that CK concentrations in R50 were higher than in ‘Sparkle’, particularly in mature shoots where the levels were maintained at elevated concentrations. These differences indicate a reduction in the CK catabolism of R50. The accumulation of CKs can be directly related to several traits of R50, with the reduced number of nodules and altered shoot ethylene response being likely indirect effects. Key words: cytokinin, de-etiolation, ethylene, etiolation, pea, nodulation mutant.
How bird droppings can affect the vegetation composition of ombrotrophic bogs
Authors: Tomassen Hilde B.M. , Smolders Alfons J.P. , Lamers Leon P.M. , Roelofs Jan G.M.
Published: October 2005

Ombrotrophic bogs depend on nutrient input from the atmosphere, so bird droppings may be a significant nutrient source. We studied the influx of N, P, and K by bird droppings, and the effects of this nutrient influx on the vegetation composition and development of Irish ombrotrophic bogs. Bird droppings significantly increased the influx of N, K, and particularly P. Concentrations of N, P, and K in the peat and vegetation were much higher at sites with bird droppings than at reference sites without bird droppings. Pinus sylvestris L. trees were able to grow much better at the sites with bird droppings, and growth was severely limited by P and K at the reference sites. Sites with bird droppings allowed atypical species to grow on ombrotrophic bogs. This change in vegetation composition alters the structure of the ombrotrophic vegetation locally, thereby affecting the atmospheric nutrient input and interception of precipitation.Key words: nitrogen, nutrient limitation, potassium, phosphorus, Pinus sylvestris, soil CO2 evolution, Sphagnum.
Presence of β-glucosidase (bgl1) gene in Phaeosphaeria nodorum and Phaeosphaeria avenaria f.sp. triticea
Authors: Reszka E , Chung K R , Tekauz A , Malkus A , Arseniuk E , Krupinsky J M , Tsang H , Ueng P P
Published: October 2005

Phaeosphaeria avenaria f.sp. avenaria (Paa), the causal agent of stagonospora leaf blotch in oats, produces a glycosyl hydrolase family 3 enzyme, β-glucosidase, which is responsible for detoxification of steroidal avenacosides in oat leaves, but is not essential for pathogenicity. For a comparative genetic relatedness study, a Paa-like β-glucosidase gene (bgl1) was PCR-amplified from Phaeosphaeria nodorum and P. avenaria f.sp. triticea, and Phaeosphaeria spp. from dallis grass (Paspalum dilatatum) (isolate S-93-48) and rye (Secale cereale) (isolate Sn48-1). Different sizes of bgl1 gene coding sequences ranging from 3018 to 3023 bp were determined. The bgl1 gene structure in these Phaeosphaeria species was identical to that of Paa and contained four exons and three introns. Nucleotide variations occurring in introns 1 and 2 of the bgl1 gene divided wheat-biotype P. nodorum into four groups. Two 12-bp-long direct sequence repeats (5'-TCA/G ACT GGT TT/CA/G) were found in the promoter region of the bgl1 gene in Phaeosphaeria species; only one repeat was present in the two P. avenaria f.sp. triticea isolates ATCC26370 and ATCC26377 (Pat2) from foxtail barley (Hordeum jubatum) and some homothallic P. avenaria f.sp. triticea isolates (Pat1). With sequence similarities in the noncoding internal transcribed spacer region of nuclear rDNA, the partial gpd gene fragment containing the intron 4, and the full-length bgl1 gene, five Phaeosphaeria isolates (5413, 1919WRS, 1920WRS, 1921WRS, Sa37-2) from oat (Avena sativa) were molecularly determined to be Paa. Two oat isolates (Sa38-1 and Sa39-2) from Poland appeared to be Pat1. The results suggest that classification of two Phaeosphaeria avenaria formae speciales based on host specificity should be re-evaluated.Key words: wheat, oat, Phaeosphaeria, β-glucosidase gene.
Chloroplast haplotype diversity patterns inPackera pauciflora(Asteraceae) are affected by geographical isolation, hybridization, and breeding system
Authors: Bain J F , Golden J L
Published: October 2005

Packera pauciflora (Pursh) Löve & Löve is a wide ranging alpine or subalpine species that is disjunct in three regions of North America: the northern Rocky mountains, QuebecLabrador, and the Sierra Nevada of California. It is one of two Packera species known to be self-fertile. We assessed intrapopulational chloroplast haplotype diversity (n) and haplotype nucleotide diversity ([Formula: see text]) and found it was lower and population structure (Φst), higher than all but one of the five other Packera species examined to date. In addition, distinct differences exist between populations from California and those from the Rocky mountains and eastern Canada. Our results suggest that geographical isolation, hybridization, and breeding system have all played a role in the formation of haplotype diversity patterns, and that, in spite of the wide disjunctions, the species is monophyletic.Key words: Asteraceae, cpDNA, haplotypes, Packera, phylogeography, self-fertility.
Geology and UPb geochronology of the Neoarchean Snare River terrane: tracking evolving tectonic regimes and crustal growth mechanisms
Authors: Bennett Venessa , Jackson Valerie A , Rivers Toby , Relf Carolyn , Horan Pat , Tubrett Mike
Published: October 2005

UPb zircon crystallization ages determined by isotope dilution  thermal ionization mass spectrometry (IDTIMS) and laser ablation microprobe  inductively coupled plasma  mass spectrometry (LAMICPMS) for 13 intrusive units in the Neoarchean Snare River terrane (SRT) provide tight constraints on the timing of crust formation and orogenic evolution. Seven metaluminous plutons were emplaced over ~80 Ma from ca. 2674 to 2589 Ma, whereas six peraluminous bodies were emplaced in a ~15 Ma interval from ca. 2598 to 2585 Ma. A detrital zircon study yielded an age spectrum with peaks correlative with known magmatic events in the Slave Province, with the ca. 2635 Ma age of the youngest detrital zircon population providing a maximum estimate for the onset of sedimentation. This age contrasts with evidence for pre-2635 Ma sedimentation elsewhere in the SRT, indicating that sedimentation was protracted and diachronous. Evolution of the SRT can be subdivided into four stages: (i) 26742635 Ma  formation of a metaluminous protoarc in a tonalitetrondhjemitegranodiorite (TTG)  granitegreenstone tectonic regime (TR1) and coeval with early turbidite sedimentation; (ii) 26352608 Ma  continued turbidite sedimentation, D1/M1 juxtaposition of turbidites and protoarc lithologies prior to ~2608 Ma, and metaluminous granitoid plutonism; (iii) 26082597 Ma  onset of TR2, collision of Snare protoarc with Central Slave Basement Complex, D2/M2 crustal thickening and mid-crustal granulite-facies metamorphism, sychronous with metaluminous and peraluminous plutonism; and (iv) 25972586 Ma  orogenic collapse, D3/M3 mid-crustal uplift, granulite-facies metamorphism, and waning metaluminous and peraluminous plutonism. The distribution of igneous rocks yields an "orogenic stratigraphy" with an older upper crust underlain by a younger synorogenic mid-crust. These data can be used to provide constraints for the interpretation of the Slave  Northern Cordillera Lithospheric Evolution (SNORCLE) Lithoprobe transect.