bluuebunny/crossref_metadata_2025_split huggingface dataset:
Molecular analysis of the Pleistocene history of Saxifraga oppositifolia in the Alps
Authors: Holderegger R. , Stehlik I. , Abbott R. J. 
Published: March 2003

AbstractA recent circumpolar survey of chloroplast DNA (cpDNA) haplotypes identified Pleistocene glacial refugia for the Arctic‐Alpine Saxifraga oppositifolia in the Arctic and, potentially, at more southern latitudes. However, evidence for glacial refugia within the ice sheet covering northern Europe during the last glacial period was not detected either with cpDNA or in another study of S. oppositifolia that surveyed random amplified polymorphic DNA (RAPD) variation. If any genotypes survived in such refugia, they must have been swamped by massive postglacial immigration of periglacial genotypes. The present study tested whether it is possible to reconstruct the Pleistocene history of S. oppositifolia in the European Alps using molecular methods. Restriction fragment length polymorphism (RFLP) analysis of cpDNA of S. oppositifolia, partly sampled from potential nunatak areas, detected two common European haplotypes throughout the Alps, while three populations harboured two additional, rare haplotypes. RAPD analysis confirmed the results of former studies on S. oppositifolia; high within, but low among population genetic variation and no particular geographical patterning. Some Alpine populations were not perfectly nested in this common gene pool and contained private RAPD markers, high molecular variance or rare cpDNA haplotypes, indicating that the species could possibly have survived on ice‐free mountain tops (nunataks) in some parts of the Alps during the last glaciation. However, the overall lack of a geographical genetic pattern suggests that there was massive immigration of cpDNA and RAPD genotypes by seed and pollen flow during postglacial times. Thus, the glacial history of S. oppositifolia in the Alps appears to resemble closely that suggested previously for the species in northern Europe.
Genetic differentiation of the dengue vector, Aedes aegypti (Ho Chi Minh City, Vietnam) using microsatellite markers
Authors: Huber K. , Le Loan L. , Hoang T. H. , Ravel S. , Rodhain F. , Failloux A.‐B. 
Published: March 2003

AbstractDengue haemorrhagic fever emerged in the 1950s and has become a major public health concern in most Asian countries. In Vietnam, little is known about the intraspecific variation of the vector and its consequences on vectorial capacity. Here we report the use of microsatellite markers to differentiate Aedes aegypti populations in Ho Chi Minh City, a typical, overcrowded Asian city. Six microsatellite loci, with 5–14 alleles per locus, were scored in 20 mosquito samples collected in 1998 in Ho Chi Minh City. We found substantial differentiation among Ae. aegypti populations from the outskirts, whereas populations from the centre of the city showed less differentiation. These results are consistent with the hypothesis that populations of Ae. aegypti in central Ho Chi Minh City are panmictic because there are abundant larval breeding sites and an abundance of humans for adults to feed upon. In contrast, populations on the outskirts become differentiated largely through the processes of genetic drift because larval breeding sites are not as abundant. These findings implicate human activities associated with urbanization, as factors shaping the genetic structure of Ae. aegypti populations.
Post‐glacial colonization of northwestern North America by the forest‐associated American marten (Martes americana, Mammalia: Carnivora: Mustelidae)
Authors: Stone Karen D. , Flynn Rodney W. , Cook Joseph A. 
Published: March 2003

AbstractPhylogeographic patterns were used to assess intraspecific diversification of American martens (Martes americana). Within martens, two morphological groups (americanaandcaurina) have been recognized, though the level of distinction between them has been debated. We examined mitochondrial cytochromebgene haplotypes from 680 martens to explore the colonization history of the Pacific Northwest and found two clades that correspond to the morphological groups. The widespreadamericanaclade extends from interior Alaska south to Montana and eastward to Newfoundland and New England (i.e. northwestern, north‐central and northeastern North America). Thecaurinaclade occurs in western North America, minimally extending from Admiralty Island (southeastern Alaska) south to Oregon and Wyoming. Our data indicated two colonization events for the Pacific Northwest (one by members of each clade) and were consistent with the persistence of populations throughout past glacial periods in eastern and western refugia. Due to vegetational and geological history following the past deglaciation, we hypothesize that martens of thecaurinaclade spread along the North Pacific Coast, and into southeastern Alaska, earlier than martens of theamericanaclade. Mismatch distributions for theamericanaclade were indicative of populations that recently experienced demographic expansion, while mismatch distributions for thecaurinaclade suggested that populations were at equilibrium. These clades are reciprocally monophyletic and distinctive (interclade divergence ranged from 2.5 to 3.0% (uncorrectedp), whereas, intraclade divergence was < 0.7%), and two regions of sympatry have been identified. Genetic signatures of past admixture in hybrid zones may have been extinguished during subsequent glacial periods when ranges contracted. This recurrent pattern of relatively restricted western, or Pacific coastal, lineages and more widespread eastern, or interior continental, lineages exists across broad taxonomic groups and suggests a shared biogeographical history.
Contrasting patterns of divergence in quantitative traits and neutral DNA markers: analysis of clinal variation
Authors: Storz Jay F. 
Published: March 2003

AbstractClinal variation in quantitative traits is often attributed to the effects of spatially varying selection. However, identical patterns can be produced by the interplay between purely stochastic processes (i.e. drift in combination with spatially restricted gene flow). One means of distinguishing between adaptive and nonadaptive causes of geographical variation is to compare relative levels of between‐population divergence in quantitative traits and neutral DNA markers. Such comparisons can be used to test whether levels of trait divergence attributable to additive genetic effects (as measured byQST) exceed null expectations based on the level of divergence at neutral marker loci (as measured byFST). The purpose of this study was to use an approach based on ‘QSTvs.FST’ contrasts to test for evidence of diversifying selection on body size of an Indian fruit bat,Cynopterus sphinx(Chiroptera: Pteropodidae). Specifically, relative levels of between‐population divergence in body size and microsatellite DNA markers were compared to assess whether the observed pattern of clinal size variation could be explained by a neutral model of isolation by distance.QSTfor body size was calculated using unbiased estimators of within‐ and between‐population variance of principal component scores. The association between body size variation and geographical/environmental distance was tested using pairwise and partial matrix correspondence tests (MCTs). Independent variables (representing causal hypotheses) were constructed as between‐locality distance matrices. The effects of neutral genetic divergence were assessed by including a matrix of pairwiseFSTas an independent variable. Partial MCTs revealed highly significant associations between phenotypic divergence (QST) and both geographical and environmental distance, even when the effects of neutral genetic divergence (FST) were partialled out. Results of the tests confirmed that migration–drift equilibrium is not a sufficient explanation for the latitudinal pattern of clinal size variation inC. sphinx. The geographical patterning of pairwiseQSTis most likely attributable to spatially varying selection and/or the direct influence of latitudinally ordered environmental effects.
Generation of a non‐sporulating strain of Streptomyces coelicolor A3(2) by the manipulation of a developmentally controlled ftsZ promoter
Authors: Flärdh Klas , Leibovitz Emmanuelle , Buttner Mark J. , Chater Keith F. 
Published: March 2003

The differentiation of Streptomyces aerial hyphae into chains of unigenomic spores occurs through the synchronous formation of multiple FtsZ rings, leading to sporulation septa. We show here that developmental control of ftsZ transcription is required for sporulation in Streptomyces coelicolor A3(2). Three putative ftsZ promoters were detected in the ftsQ–ftsZ intergenic region. In addition, some readthrough from upstream promoter(s) contributed to ftsZ transcription. S1 nuclease protection assays and transcriptional fusions of the ftsZ promoter region to the egfp gene (for green fluorescent protein) provided evidence that ftsZ2p is a developmentally controlled promoter that is specifically upregulated in sporulating aerial hyphae. This upregulation required all the six early regulatory sporulation genes that were tested: whiA, B, G, H, I and J. The DNA sequence of the promoter indicated that it is not part of the developmental regulon that is controlled by the RNA polymerase sigma factor σWhiG. A strain in which the ftsZ2p promoter was inactivated grew normally during vegetative growth and formed aerial mycelium, but was deficient in sporulation septation. Thus, ftsZ2p was dispensable for vegetative growth, but was required for the strain to make sufficient FtsZ to support developmentally controlled multiple cell divisions in aerial hyphae.
The Escherichia coli histone‐like protein HU regulates rpoS translation
Authors: Balandina Anna , Claret Laurent , Hengge‐Aronis Regine , Rouviere‐Yaniv Josette 
Published: March 2003

Escherichia coli HU protein is a major component of the bacterial nucleoid. HU stabilizes higher order nucleoprotein complexes and belongs to a family of DNA architectural proteins. Here, we report that HU is required for efficient expression of the sigma S subunit of RNA polymerase. This rpoS‐encoded alternative σS factor induces a number of genes implicated in cell survival in stationary phase and in multiple stress resistance. By analysis of rpoS–lacZ fusions and by pulse‐chase experiments, we show that the efficiency of rpoS translation is reduced in cells lacking HU, whereas neither rpoS transcription nor protein stability is affected by HU. Gel mobility shift assays show that HU is able to bind specifically an RNA fragment containing the translational initiation region of rpoS mRNA 1000‐fold more strongly than double‐stranded DNA. Together with the in vivo data, this finding strongly suggests that, by binding to rpoS mRNA, HU directly stimulates rpoS translation. We demonstrate here that HU, an abundant DNA‐binding, histone‐like protein, is able specifically to recognize an RNA molecule and therefore play a role in post‐transcriptional regulation.
A maize‐specifically expressed gene cluster in Ustilago maydis
Authors: Basse Christoph W. , Kolb Sebastian , Kahmann Regine 
Published: March 2003

SummaryThe corn pathogen Ustilago maydis requires its host plant maize for development
and completion of its sexual cycle. We have identified the fungal mig2–1
gene as being specifically expressed during this biotrophic stage. Intriguingly,
mig2–1 is part of a gene cluster comprising five highly homologous
and similarly regulated genes designated mig2–1 to mig2–5.
Deletion analysis of the mig2–1 promoter provides evidence for negative
and positive regulation. The predicted polypeptides of all five genes lack significant
homologies to known genes but have characteristic N‐terminal secretion sequences.
The secretion signals of mig2–1 and mig2–5 were shown
to be functional, and secretion of a full length Mig2–1‐eGFP fusion protein
to the extracellular space was demonstrated. The central domains of the Mig2 proteins
are highly variable whereas the C‐termini are strongly conserved and share a characteristic
pattern of eight cysteine residues. The mig2 gene cluster was conserved in
a wide collection of U. maydis strains. Interestingly, some U. maydis
isolates from South America had lost the mig2–4 gene as a result of
a homologous recombination event. Furthermore, the related Ustilago scitaminea
strain, which is pathogenic on sugar cane, appears to lack the mig2 cluster.
We describe a model of how the mig2 cluster might have evolved and discuss its possible role in governing host interaction.
An ABC‐type, high‐affinity urea permease identified in cyanobacteria
Authors: Valladares Ana , Montesinos María Luz , Herrero Antonia , Flores Enrique 
Published: March 2003

SummaryUrea is an important nitrogen source for many microorganisms, but urea active transporters have not been characterized at a molecular level in any bacterium. Cells of Synechocystis sp. PCC 6803 and Anabaena sp. PCC 7120 exhibited the capacity to take up [14C]‐urea from low‐concentration (<1 μM) urea solutions. The Ks of Anabaena cells for urea was about 0.11 μM, and the observed uptake activity involved the transport and metabolism of urea. In contrast to urease, which was constitutively ex‐pressed, expression of the high‐affinity urea uptake activity was subjected to nitrogen control. In an Anabaena ureG (urease–) mutant, a concentrative, active transport of urea could be demonstrated. We found that a mutant of open reading frame (ORF) sll0374 from the Synechocystis genomic sequence lacked urea transport activity. This ORF encoded a conserved component of an ABC‐type transporter, but it is not clustered together with any other possible transporter‐encoding gene. An Anabaena homologue of sll0374, urtE, was isolated and found to be part of a cluster of genes, urtABCDE, putatively encoding all the elements of an ABC‐type permease. Although the longest transcript that we could detect only covered urtABC, the impairment of urea transport by inactivation of urtA, urtB or urtE suggested that the whole gene cluster is expressed producing the urea permease. Expression was induced under nitrogen‐limiting conditions, and a complex promoter regulated by the cyanobacterial global nitrogen control transcription factor NtcA was found upstream from urtA. Our work adds urea to the known substrates of the versatile class of ABC‐type transporters and suggests the involvement of a transporter of this superfamily in urea scavenging by some bacteria in natural environments.
Evaluation of a nutrient starvation model of Mycobacterium tuberculosis persistence by gene and protein expression profiling
Authors: Betts Joanna C. , Lukey Pauline T. , Robb Linda C. , McAdam Ruth A. , Duncan Ken 
Published: March 2003

SummaryThe search for new TB drugs that rapidly and effectively sterilize the tissues and are thus able to shorten the duration of chemotherapy from the current 6 months has been hampered by a lack of understanding of the metabolism of the bacterium when in a ‘persistent’ or latent form. Little is known about the condition in which the bacilli survive, although laboratory models have shown that Mycobacterium tuberculosis can exist in a non‐growing, drug‐resistant state that may mimic persistence in vivo. Using nutrient starvation, we have established a model in which M. tuberculosis arrests growth, decreases its respiration rate and is resistant to isoniazid, rifampicin and metronidazole. We have used microarray and proteome analysis to investigate the response of M. tuberculosis to nutrient starvation. Proteome analysis of 6‐week‐starved cultures revealed the induction of several proteins. Microarray analysis enabled us to monitor gene expression during adaptation to nutrient starvation and confirmed the changes seen at the protein level. This has provided evidence for slowdown of the transcription apparatus, energy metabolism, lipid biosynthesis and cell division in addition to induction of the stringent response and several other genes that may play a role in maintaining long‐term survival within the host. Thus, we have generated a model with which we can search for agents active against persistent M. tuberculosis and revealed a number of potential targets expressed under these conditions.
c‐Src/Lyn kinases activate Helicobacter pylori CagA through tyrosine phosphorylation of the EPIYA motifs
Authors: Stein Markus , Bagnoli Fabio , Halenbeck Robert , Rappuoli Rino , Fantl Wendy J. , Covacci Antonello 
Published: March 2003

SummaryThe human pathogen Helicobacter pylori colonizes the mucous layer of the

stomach. During parasitic infection, freely swimming bacteria adhere to the gastric

epithelial cells and trigger intracellular signalling pathways. This process requires

the translocation of the effector protein CagA into the host cell through a specialized

type IV secretion system encoded in the cag pathogenicity island. Following transfer, CagA is phosphorylated on tyrosine re‐sidues by a host cell kinase. Here, we describe how the tyrosine phosphorylation of CagA is restricted to a previously identified repeated sequence called D1. This sequence is located in the C‐terminal half of the protein and contains the five‐amino‐acid motif EPIYA, which is amplified by duplications in a large fraction of clinical isolates. Tyrosine phosphorylation of CagA is essential for the activation process that leads to dramatic changes in the morphology of cells growing in culture. In addition, we observed that two members of the src kinases family, c‐Src and Lyn, account for most of the CagA‐specific kinase activity in host cell lysates. Thus, CagA translocation followed by tyrosine phosphorylation at the EPIYA motifs promotes a growth factor‐like response with intense cytoskeletal rearrangements, cell elongation effects and increased cellular motility.
afsS is a target of AfsR, a transcriptional factor with ATPase activity that globally controls secondary metabolism in Streptomyces coelicolor A3(2)
Authors: Lee Ping‐Chin , Umeyama Takashi , Horinouchi Sueharu 
Published: March 2003

SummaryAfsR is a pleiotropic, global regulator that controls the production of actinorhodin, undecylprodigiosin and calcium‐dependent antibiotic in Streptomyces coelicolor
A3(2). AfsR, with 993 amino acids, is phosphorylated on serine and threonine residues
by a protein serine/threonine kinase AfsK and contains an OmpR‐like DNA‐binding fold
at its N‐terminal portion and A‐ and B‐type nucleotide‐binding motifs in the middle
of the protein. The DNA‐binding domain, in‐dependently of the nucleotide‐binding
domain, contributed the binding of AfsR to the upstream region of afsS that
locates immediately 3′ to afsR and encodes a 63‐amino‐acid protein.
No transcription of afsS in the ΔafsR background and restoration
of afsS transcription by afsR on a plasmid in the same genetic background
indicated that afsR served as a transcriptional activator for afsS.
Interestingly, the AfsR binding site overlapped the promoter of afsS, as determined
by DNase I protection assay and high‐resolution S1 nuclease mapping. The nucleotide‐binding
domain contributed distinct ATPase and GTPase activity. The phosphorylation of AfsR
by AfsK greatly enhanced the DNA‐binding activity and modulated the ATPase activity.
The DNA‐binding ability of AfsR was independent of the ATPase activity. However,
the ATPase activity was essential for transcriptional activation of afsS, probably because the energy available from ATP hydrolysis is required for the isomerization of the closed complex between AfsR and RNA polymerase to a transcriptionally competent open complex. Thus, AfsR turns out to be a unique transcriptional factor, in that it is modular, in which DNA‐binding and ATPase activities are physically separable, and the two functions are modulated by phosphorylation on serine and threonine residues.
Molecular characterization of phenyllactate dehydratase and its initiator from Clostridium sporogenes
Authors: Dickert Sandra , Pierik Antonio J. , Buckel Wolfgang 
Published: March 2003

SummaryThe heterotrimeric phenyllactate dehydratase from Clostridium sporogenes,
FldABC, catalyses the reversible dehydration of (R)‐phenyllactate to (E)‐cinnamate
in two steps: (i) CoA‐transfer from the cofactor cinnamoyl‐CoA to phenyllactate to
yield phenyllactyl‐CoA and the product cinnamate mediated by FldA, a (R)‐phenyllactate
CoA‐transferase; followed by (ii) dehydration of phenyllactyl‐CoA to cinnamoyl‐CoA
mediated by heterodimeric FldBC, a phenyllactyl‐CoA dehydratase. Phenyllactate dehydratase
requires initiation by ATP, MgCl2 and a reducing agent such as dithionite
mediated by an extremely oxygen‐sensitive initiator protein (FldI) present in the
cell‐free extract. All four genes coding for these proteins were cloned and shown
to be clustered in the order fldAIBC, which shares over 95% sequence
identity of nucleotide and protein levels with a gene cluster detected in the genome
of the closely related Clostridium botulinum Hall strain A. FldA shows sequence
similarities to a new family of CoA‐transferases, which apparently do not form covalent
enzyme CoA‐ester intermediates. An N‐terminal Strep II‐Tag containing enzymatically
active FldI was overproduced and purified from Escherichia coli. FldI was
characterized as a homodimeric protein, which contains one [4Fe‐4S]1+/2+
cluster with an electron spin S= 3/2 in the reduced form. The amino
acid sequence as well as the chemical and EPR‐properties of the pure protein are
very similar to those of component A of 2‐hydroxyglutaryl‐CoA dehydratase from Acidaminococcus
fermentans (HgdC), which was able to replace FldI in the activation of phenyllactate
dehydratase. Only in the oxidized state, FldI and component A exhibit significant
ATPase activity, which appears to be essential for unidirectional electron transfer.
Both subunits of phenyllactyl‐CoA dehydratase (FldBC) show significant sequence similarities
to both subunits of 2‐hydroxyglutaryl‐CoA dehydratase (HgdAB). The fldAIBC
gene cluster resembles the hadAIBC gene cluster in the genome of Clostridium
difficile and the hadABC,I genes in C. botulinum. The four subunits
of these deduced 2‐hydroxyacid dehydratases (65–81%
amino acid sequence identity between the had genes) probably code for a 2‐hydroxyisocaproate
dehydratase involved in leucine fermentation. This enzyme could be the target for
metronidazole in the treatment of pseudomembranous enterocolitis caused by C.
difficile.