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S Clegg

Publications and source records attributed to S Clegg.

At least 19 recordsLinked to original sources
PubMed2006

Divergence in mating signals correlates with ecological variation in the migratory songbird, Swainson's thrush (Catharus ustulatus).

Song divergence among populations of passerine birds is recognized as a potentially important premating isolation mechanism and may represent the first step in speciation. Because song divergence may be influenced by an array of acoustic, ecological, and genetic factors, the study of its origin requires a multifaceted approach. Here we describe the relationship between acoustic, neutral genetic and ecological variation in five populations of the Swainson's thrush: two from coastal temperate rainforest habitat representing the 'russet-backed' subspecies group, two from inland coniferous forest habitat representing the 'olive-backed' subspecies group, and one mixed locality that resides within a contact zone between the two groups. Song in the five populations is analysed using a multivariate analysis of spectral and temporal measurements, population genetic structure is assessed using an analysis of five microsatellite loci and ecological differences between populations are quantified using an analysis of climatic parameters. Matrix correspondence tests are used to distinguish between the potential for drift and selection in driving song divergence. No significant correlation was found between acoustic and genetic distance suggesting that song divergence cannot be explained by drift alone. A significant correlation between ecological and acoustic distance after accounting for genetic distance, suggests a potential role for ecological selection on divergence in spectral and temporal components of Swainson's thrush song.

Acoustics↗
PubMed2005

The complex nature of constitutional de novo apparently balanced translocations in patients presenting with abnormal phenotypes.

OBJECTIVE: To describe the systematic analysis of constitutional de novo apparently balanced translocations in patients presenting with abnormal phenotypes, characterise the structural chromosome rearrangements, map the translocation breakpoints, and report detectable genomic imbalances. METHODS: DNA microarrays were used with a resolution of 1 Mb for the detailed genome-wide analysis of the patients. Array CGH was used to screen for genomic imbalance and array painting to map chromosome breakpoints rapidly. These two methods facilitate rapid analysis of translocation breakpoints and screening for cryptic chromosome imbalance. Breakpoints of rearrangements were further refined (to the level of spanning clones) using fluorescence in situ hybridisation where appropriate. RESULTS: Unexpected additional complexity or genome imbalance was found in six of 10 patients studied. The patients could be grouped according to the general nature of the karyotype rearrangement as follows: (A) three cases with complex multiple rearrangements including deletions, inversions, and insertions at or near one or both breakpoints; (B) three cases in which, while the translocations appeared to be balanced, microarray analysis identified previously unrecognised imbalance on chromosomes unrelated to the translocation; (C) four cases in which the translocation breakpoints appeared simple and balanced at the resolution used. CONCLUSIONS: This high level of unexpected rearrangement complexity, if generally confirmed in the study of further patients, will have an impact on current diagnostic investigations of this type and provides an argument for the more widespread adoption of microarray analysis or other high resolution genome-wide screens for chromosome imbalance and rearrangement.

Cell Line↗
PubMed2004

The DNA sequence and comparative analysis of human chromosome 10.

The finished sequence of human chromosome 10 comprises a total of 131,666,441 base pairs. It represents 99.4% of the euchromatic DNA and includes one megabase of heterochromatic sequence within the pericentromeric region of the short and long arm of the chromosome. Sequence annotation revealed 1,357 genes, of which 816 are protein coding, and 430 are pseudogenes. We observed widespread occurrence of overlapping coding genes (either strand) and identified 67 antisense transcripts. Our analysis suggests that both inter- and intrachromosomal segmental duplications have impacted on the gene count on chromosome 10. Multispecies comparative analysis indicated that we can readily annotate the protein-coding genes with current resources. We estimate that over 95% of all coding exons were identified in this study. Assessment of single base changes between the human chromosome 10 and chimpanzee sequence revealed nonsense mutations in only 21 coding genes with respect to the human sequence.

Animals↗
PubMed2004

DNA sequence and analysis of human chromosome 9.

Chromosome 9 is highly structurally polymorphic. It contains the largest autosomal block of heterochromatin, which is heteromorphic in 6-8% of humans, whereas pericentric inversions occur in more than 1% of the population. The finished euchromatic sequence of chromosome 9 comprises 109,044,351 base pairs and represents >99.6% of the region. Analysis of the sequence reveals many intra- and interchromosomal duplications, including segmental duplications adjacent to both the centromere and the large heterochromatic block. We have annotated 1,149 genes, including genes implicated in male-to-female sex reversal, cancer and neurodegenerative disease, and 426 pseudogenes. The chromosome contains the largest interferon gene cluster in the human genome. There is also a region of exceptionally high gene and G + C content including genes paralogous to those in the major histocompatibility complex. We have also detected recently duplicated genes that exhibit different rates of sequence divergence, presumably reflecting natural selection.

Base Composition↗
PubMed2003

The DNA sequence and analysis of human chromosome 6.

Chromosome 6 is a metacentric chromosome that constitutes about 6% of the human genome. The finished sequence comprises 166,880,988 base pairs, representing the largest chromosome sequenced so far. The entire sequence has been subjected to high-quality manual annotation, resulting in the evidence-supported identification of 1,557 genes and 633 pseudogenes. Here we report that at least 96% of the protein-coding genes have been identified, as assessed by multi-species comparative sequence analysis, and provide evidence for the presence of further, otherwise unsupported exons/genes. Among these are genes directly implicated in cancer, schizophrenia, autoimmunity and many other diseases. Chromosome 6 harbours the largest transfer RNA gene cluster in the genome; we show that this cluster co-localizes with a region of high transcriptional activity. Within the essential immune loci of the major histocompatibility complex, we find HLA-B to be the most polymorphic gene on chromosome 6 and in the human genome.

Animals↗
PubMed2003

Array painting: a method for the rapid analysis of aberrant chromosomes using DNA microarrays.

OBJECTIVE: The authors describe a method, termed array painting, which allows the rapid, high resolution analysis of the content and breakpoints of aberrant chromosomes. METHODS: Array painting is similar in concept to reverse chromosome painting and involves the hybridisation of probes generated by PCR of small numbers of flow sorted chromosomes on large insert genomic clone DNA microarrays. RESULTS: and CONCLUSIONS: By analysing patients with cytogenetically balanced chromosome rearrangements, the authors show the effectiveness of array painting as a method to map breakpoints prior to cloning and sequencing chromosome rearrangements.

Adult↗
PubMed2001

Control of FimY translation and type 1 fimbrial production by the arginine tRNA encoded by fimU in Salmonella enterica serovar Typhimurium.

Expression of type 1 fimbriae in Salmonella enterica serovar Typhimurium undergoes phase variation or alteration between a fimbriate and a non-fimbriate phenotype. This variation is known to be dependent upon environmental conditions in vitro and is thought to be a complex process involving regulation by a number of proteins. The regulatory genes located within the fim cluster include fimZ, fimY and fimW. A fourth gene of the cluster, fimU, encodes a tRNA molecule specific for rare arginine codons. We have shown previously that fimU affects the expression of S. typhimurium type 1 fimbriae, and that fimU is functionally related to the Escherichia coli gene argU. A high frequency of rare arginine codons was found within the three fim regulatory genes, and five of these codons were clustered within fimY alone. To investigate the affects of fimU on FimY production, a FimY fusion with the E. coli maltose-binding protein was constructed and expressed in an E. coli argU background. Western blots of extracts from the argU mutant and parental strain indicated that production of FimY was significantly reduced in the absence of a functional tRNAArg(UCU). FimY production in this mutant could be restored to high levels when fimU was introduced on a plasmid, and also when three rare arginine codons, located within the first 14 positions within fimY, were exchanged for major arginine codons. A Tn10 insertion from a Salmonella enteritidis fimU mutant was transduced into S. typhimurium, and this strain was analysed for the expression of type 1 fimbriae. The resulting S. typhimurium fimU mutant was found to be non-fimbriate under all conditions tested and could be complemented by the introduction of fimU alone on a plasmid. In addition, this mutant could be complemented by transformation with fimY altered in the first three rare arginine codons. Reverse transcriptase-polymerase chain reaction confirmed that the fimY transcript was present at similar levels in the fimU mutant and parental strain. These results indicated that the observed inhibition of protein expression was not occurring at the transcriptional level. Analysis of expression of the malEfimY fusion in the S. typhimurium fimU mutant and parental strain confirmed the data observed in E. coli. In contrast, a FimW fusion was found to be produced at similar levels in both the fimU mutant and the parental strain. Together, these data indicate that the absence of a functional fimU results in the inhibition of efficient FimY translation, and thus type 1 fimbrial production in S. typhimurium.

Bacterial Proteins↗
PubMed2001

Type 3 fimbrial shaft (MrkA) of Klebsiella pneumoniae, but not the fimbrial adhesin (MrkD), facilitates biofilm formation.

Isolates of Klebsiella pneumoniae are responsible for opportunistic infections, particularly of the urinary tract and respiratory tract, in humans. These bacteria express type 3 fimbriae that have been implicated in binding to eucaryotic cells and matrix proteins. The type 3 fimbriae mediate binding to target tissue using the MrkD adhesin that is associated with the fimbrial shaft comprised of the MrkA protein. The formation of biofilms in vitro by strains of K. pneumoniae was shown to be affected by the production of fimbriae on the bacterial surface. However, a functional MrkD adhesin was not necessary for efficient biofilm formation. Nonfimbriate strains were impaired in their ability to form biofilms. Using isogenic fimbriate and nonfimbriate strains of K. pneumoniae expressing green fluorescent protein it was possible to demonstrate that the presence of type 3 fimbriae facilitated the formation of dense biofilms in a continuous-flowthrough chamber. Transformation of nonfimbriate mutants with a plasmid possessing an intact mrk gene cluster restored the fimbrial phenotype and the rapid ability to form biofilms.

Adhesins, Bacterial↗
PubMed2001

FimW is a negative regulator affecting type 1 fimbrial expression in Salmonella enterica serovar typhimurium.

Type 1 fimbriae are proteinaceous surface appendages that carry adhesins specific for mannosylated glycoproteins. These fimbriae are found on most members of the family Enterobacteriaceae and are known to facilitate binding to a variety of eukaryotic cells, including those found on the mucosal surfaces of the alimentary tract. We have shown that the regulation of type 1 fimbrial expression in Salmonella enterica serovar Typhimurium is controlled, in part, by the products of four genes found within the fim gene cluster: fimZ, fimY, fimW, and fimU. To better understand the specific role of FimW in fimbrial expression, a mutation was constructed in this gene by the insertion of a kanamycin resistance DNA cassette into the chromosome. The resulting fimW mutation was characterized by mannose-sensitive hemagglutination and agglutination with fimbria-specific antiserum. Assays suggested that this mutant was more strongly fimbriate than the parental strain, exhibiting a four- to eightfold increase in fimbrial production. The fimW mutation was introduced into a second strain of Salmonella enterica serovar Typhimurium, and this mutant was also found to be strongly fimbriate compared to the parental strain. Consistent with the role of this protein as a negative regulator, fimA-lacZ expression in serovar Typhimurium, as well as in Escherichia coli, was increased twofold in the absence of functional FimW. Primer extension analysis determined that fimW transcription is initiated from its own promoter 31 bp upstream of the translation start site. Analysis using a fimW-lacZ reporter indicated that fimW expression in serovar Typhimurium was increased under conditions that select for poorly fimbriate bacteria and low fimA expression. FimW also appears to act as an autoregulator, since expression from the fimW-lacZ reporter was increased in a fimW mutant. FimW was partially purified by fusion with the E. coli maltose-binding protein. Use of this FimW protein extract, as well as others, in DNA-binding assays was unable to identify a specific binding site for FimW in the fimA, fimZ, fimY, or fimW promoter regions. To analyze protein-protein interactions, FimW was expressed in a LexA-based two-hybrid system in E. coli. A significant interaction between FimW and the DNA-binding activator protein, FimZ, was detected using this system. These results indicate that FimW is a negative regulator of serovar Typhimurium type 1 fimbrial expression and may function by interfering with FimZ-mediated activation of fimA expression.

Bacterial Proteins↗
PubMed2000

Characterization of FimY as a coactivator of type 1 fimbrial expression in Salmonella enterica serovar Typhimurium.

Type 1 fimbriae of Salmonella enterica serovar Typhimurium are surface appendages that carry adhesins specific for mannosylated host glycoconjugates. Regulation of the major fimbrial subunit is thought to be controlled by a number of ancillary fim genes, including fimZ, fimY, fimW, and fimU. Previous studies using a FimZ mutant have indicated that this protein is necessary for fimA expression, and in vitro DNA binding assays determined that FimZ is a transcriptional activator that binds directly to the fimA promoter. To determine the role of FimY as a potential regulator of fimbrial expression, a fimY mutant of serovar Typhimurium was generated by allelic exchange. This mutant was found to be phenotypically nonfimbriate. No transcription from the fimA promoter was detected in a fimY mutant containing a fimA-lacZ reporter construct located on the chromosome. In addition, transcription from the cloned fimY promoter was not detected in Escherichia coli unless both FimZ and FimY were present, indicating that these proteins also act as coactivators of fimY expression. Consistent with these results, there is no transcription from a fimY-lacZ reporter construct within a serovar Typhimurium fimY or fimZ mutant. Studies using the fimY-lacZ construct reveal that expression of this gene varies with environmental conditions in a manner similar to fimA expression. Extensive in vitro DNA binding assays using extracts from E. coli that overexpress FimY, as well as partially purified FimY, were unable to identify a specific interaction between FimY and the fimA or fimY promoter. The results indicate that FimY is a positive regulator of fimbrial expression and that this protein acts in cooperation with FimZ to regulate the expression of Salmonella type 1 fimbrial appendages.

Adhesins, Bacterial↗
PubMed2000

Salmonella enterica serovars gallinarum and pullorum expressing Salmonella enterica serovar typhimurium type 1 fimbriae exhibit increased invasiveness for mammalian cells.

Salmonella enterica serovars Gallinarum and Pullorum are S. enterica biotypes that exhibit host specificity for poultry and aquatic birds and are not normally capable of causing disease in mammalian hosts. During their evolution toward host restriction serovars Gallinarum and Pullorum lost their ability to mediate mannose-sensitive hemagglutination (MSHA), a phenotype correlated with adherence to certain cell types. Because adherence is an essential requirement for invasion of cells by bacterial pathogens, we examined whether MHSA type 1 fimbriae would increase the ability of serovars Pullorum and Gallinarum to invade normally restrictive cells. Serovars Gallinarum and Pullorum expressing S. enterica serovar Typhimurium strain LT2 type 1 fimbriae exhibited a 10- to 20-fold increased ability to adhere to and a 20- to 60-fold increased invasion efficiency of the human epithelial HEp-2 cell line. Invasion was accompanied by extensive ruffling of the membranes of the HEp-2 cells. In a murine ligated ileal loop model, a 32% increase in the number of M-cell ruffles was seen when serovar Gallinarum expressed serovar Typhimurium type 1 fimbriae.

Animals↗
PubMed1999

Construction and characterization of mutations within the Klebsiella mrkD1P gene that affect binding to collagen type V.

The fimbria-associated MrkD1P protein mediates adherence of type 3 fimbriate strains of Klebsiella pneumoniae to collagen type V. Currently, three different MrkD adhesins have been described in Klebsiella species, and each possesses a distinctive binding pattern. Therefore, the binding abilities of mutants possessing defined mutations within the mrkD1P gene were examined in order to determine whether specific regions of the adhesin molecule were responsible for collagen binding. Both site-directed and chemically induced mutations were constructed within mrkD1P, and the ability of the gene products to be incorporated into fimbrial appendages or bind to collagen was determined. Binding to type V collagen was not associated solely with one particular region of the MrkD1P protein, and two classes of nonadhesive mutants were isolated. In one class of mutants, the MrkD adhesin was not assembled into the fimbrial shaft, whereas in the second class of mutants, the adhesin was associated with fimbriae but did not bind to collagen. Both hemagglutinating and collagen-binding activities were associated with the MrkD1P molecule, since P pili and type 3 fimbriae carrying adhesive MrkD proteins exhibited identical binding properties.

Adhesins, Bacterial↗
PubMed1998

Characterization of the type 3 fimbrial adhesins of Klebsiella strains.

The Klebsiella pneumoniae fimbrial adhesin, MrkD, mediates adherence to the basolateral surfaces of renal and pulmonary epithelia and to the basement membranes of tissues. Although all isolates possessing the MrkD adhesin mediate the agglutination, in vitro, of erythrocytes treated with tannic acid, the mrkD gene is not conserved within species. The ability of a plasmid-borne mrkD gene product to mediate binding to type V collagen is associated frequently with strains of K. oxytoca and rarely with strains of K. pneumoniae. In K. pneumoniae, the MrkD adhesin is located within a chromosomally borne gene cluster and mediates binding to collagen types IV and V. The plasmid-borne determinant, mrkD1P, and the chromosomally borne gene, mrkD1C, are not genetically related. Some strains of enterobacteria possess a mrkD1C allele that is associated with hemagglutinating activity but does not bind to either type IV or type V collagen.

Adhesins, Bacterial↗
PubMed1997

Construction and characterization of type 1 non-fimbriate and non-adhesive mutants of Salmonella typhimurium.

Mutations in the fimH gene of Salmonella typhimurium result in a non-fimbriate, non-adhesive phenotype. This phenotype was shown to be due to the lack of both fimH and fimF expression since disruption of the fimH gene by insertion of a DNA cassette into this determinant results in mutants that are complemented by plasmids carrying both fimH and fimF. Deletion mutations within the S. typhimurium fimH gene carried on a recombinant plasmid can be used to complement the mutant, and these transformants are non-adhesive but fully fimbriate, consistent with the role of FimH as being necessary for fimbrial adhesin expression. Adherence to erythrocytes, HeLa, and Hep-2 cells is associated with expression of the FimH polypeptide, and fimbriate strains that cannot synthesize FimH are non-adhesive. Discrete differences in the amino acid sequences of the adhesive type 1 and the non-hemagglutinating type 2 FimH polypeptides were detected, and are most likely responsible for the differences in hemagglutinating activity.

Adhesins, Bacterial↗
PubMed1997

Binding of the type 3 fimbriae of Klebsiella pneumoniae to human endothelial and urinary bladder cells.

Binding of the two identified type 3 fimbrial variants of Klebsiella pneumoniae to human endothelial EA-hy926 and bladder T24 cells was assessed. The recombinant Escherichia coli strain LE392(pFK12), expressing plasmid-encoded type 3 fimbriae of K. pneumoniae, adhered to both cell lines, and the fimbriae purified from the strain bound to both cell lines in a dose-dependent manner. Adhesiveness to both cell lines of chromosomally encoded type 3 fimbriae from K. pneumoniae IApc35 was lower. No binding was detected with type 1 fimbriae of K. pneumoniae. Both type 3 fimbrial variants exhibited a significantly lower affinity for the cell lines than did S fimbriae of meningitis-associated E. coli.

Cell Line↗
PubMed1997

Hyperthermic treatment of malignant diseases: current status and a view toward the future.

New studies in hyperthermia at the basic science, engineering, and clinical level have stimulated renewed enthusiasm for re-investigating its potential as an anticancer therapy. This article reviews the salient features of these recent results and points out areas for additional investigation. Highlighting these new results is the publication of several positive phase III trials for thermoradiotherapy compared to radiotherapy alone. Important highlights are the encouraging results using magnetic resonance imaging for noninvasive thermometry. If this technology is successfully implemented with real time power control it will revolutionize the clinical application of hyperthermia.

Animals↗