Biomedical subjects

Dominic P Kwiatkowski

Publications and source records attributed to Dominic P Kwiatkowski.

At least 19 recordsLinked to original sources

PubMed2006

Localization of a long-range cis-regulatory element of IL13 by allelic transcript ratio mapping.

It appears that, for many genes, the two alleles possessed by an individual may produce different amounts of transcript. When such allelic differences in transcription are observed for some individuals but not others, a plausible explanation is genetic variation in the cis-acting elements that regulate the gene in question. Here we describe a novel analytical approach that uses such observations, combined with genotyping data from the HapMap project, to define the genomic location of cis-acting regulatory elements. When applied to the human 5q31 chromosomal region, where complex regulatory mechanisms are known to exist, we demonstrate the sensitivity of this approach by locating a highly significant cis-regulatory element operating on IL13 at long range from a position 250 kb upstream from the gene (P = 2 x 10(-6)). As this method is unaffected by other sources of variation, such as environmental and trans-acting genetic factors, it provides a tractable approach for dissecting the complexities of genetic variation in gene regulation.

Alleles↗

PubMed2006

Data sharing and intellectual property in a genomic epidemiology network: policies for large-scale research collaboration.

Genomic epidemiology is a field of research that seeks to improve the prevention and management of common diseases through an understanding of their molecular origins. It involves studying thousands of individuals, often from different populations, with exacting techniques. The scale and complexity of such research has required the formation of research consortia. Members of these consortia need to agree on policies for managing shared resources and handling genetic data. Here we consider data-sharing and intellectual property policies for an international research consortium working on the genomic epidemiology of malaria. We outline specific guidelines governing how samples and data are transferred among its members; how results are released into the public domain; when to seek protection for intellectual property; and how intellectual property should be managed. We outline some pragmatic solutions founded on the basic principles of promoting innovation and access.

Access to Information↗

PubMed2006

Seasonal childhood anaemia in West Africa is associated with the haptoglobin 2-2 genotype.

BACKGROUND: Anaemia is a major cause of morbidity and mortality for children in Africa. The plasma protein haptoglobin (Hp) binds avidly to free haemoglobin released following malaria-induced haemolysis. Haptoglobin polymorphisms result in proteins with altered haemoglobin-binding capacity and different antioxidant, iron-recycling, and immune functions. Previous studies examined the importance of haptoglobin polymorphism in malaria and iron homeostasis, but it is unknown whether haptoglobin genotype might be a risk factor for anaemia in children in a malaria-endemic area. METHODS AND FINDINGS: A cohort of 780 rural Gambian children aged 2-6 y was surveyed at the start and end of the malaria season. Samples were taken to assess haemoglobin (Hb) concentration, iron status (ferritin, zinc protoporphyrin, transferrin saturation, and soluble transferrin receptor concentration), haptoglobin concentration, alpha-1-antichymotrypsin (a measure of inflammation), and malaria parasites on blood film. We extracted DNA and genotyped for haptoglobin, sickle cell, and glucose-6-phosphate (G6PD) deficiency. Mean Hb levels fell over the malaria season. Children with the haptoglobin 2-2 genotype (17%) had a greater mean drop in Hb level over the malaria season (an 8.9 g/l drop; confidence interval [CI] 5.7, 12.1) compared to other children (a 5.1 g/l drop; CI 3.8, 6.4). In multivariate regression analysis, controlling for baseline Hb level, age group, village, malaria parasites on blood film, iron status, haptoglobin concentration, and G6PD deficiency, haptoglobin genotype predicted Hb level at the end of the malaria season (p = 0.0009, coefficient = -4.2). Iron status was not influenced by haptoglobin genotype. CONCLUSIONS: The finding that haptoglobin 2-2 genotype is a risk factor for anaemia in children in a malaria-endemic area may reflect the reduced ability of the Hp2-2 polymer to scavenge free haemoglobin-iron following malaria-induced haemolysis. The magnitude of the effect of haptoglobin genotype (4 g/l Hb difference, p = 0.0009) was comparable to that of G6PD deficiency or HbAS (3 g/l difference, p = 0.03; and 2 g/l difference, p = 0.68, respectively).

Anemia↗

PubMed2006

A coding polymorphism in matrix metalloproteinase 9 reduces risk of scarring sequelae of ocular Chlamydia trachomatis infection.

BACKGROUND: Trachoma, an infectious disease of the conjunctiva caused by Chlamydia trachomatis, is an important global cause of blindness. A dysregulated extracellular matrix (ECM) proteolysis during the processes of tissue repair following infection and inflammation are thought to play a key role in the development of fibrotic sequelae of infection, which ultimately leads to blindness. Expression and activity of matrix metalloproteinase 9 (MMP-9), a major effector of ECM turnover, is up-regulated in the inflamed conjunctiva of trachoma subjects. Genetic variation within the MMP9 gene affects in vitro MMP9 expression levels, enzymatic activity and susceptibility to various inflammatory and fibrotic conditions. METHODS: We genotyped 651 case-control pairs from trachoma endemic villages in The Gambia for coding single nucleotide polymorphisms (SNPs) in the MMP9 gene using the high-throughput Sequenom system. Single marker and haplotype conditional logistic regression (CLR) analysis for disease association was performed. RESULTS: The Q279R mutation located in exon 6 of MMP9 was found to be associated with lower risk for severe disease sequelae of ocular Chlamydia trachomatis infection. This mutation, which leads to a nonsynonymous amino-acid change within the active site of the enzyme may reduce MMP-9-induced degradation of the structural components of the ECM during inflammatory episodes in trachoma and its associated fibrosis. CONCLUSION: This work supports the hypothesis that MMP-9 has a role in the pathogenesis of blinding trachoma.

Case-Control Studies↗

PubMed2006

Context-specific functional effects of IFNGR1 promoter polymorphism.

We report evidence of a polymorphism in the promoter region of IFNGR1 (encoding interferon-gamma receptor 1) that has opposite functional effects in different cellular contexts. It is a deletion/insertion polymorphism that is found in Africans but not Europeans or Asians, and has been associated with resistance to severe malaria. We find that the IFNGR1-470del allele acts to suppress binding of nuclear proteins to the IFNGR1 promoter region in a manner that is specific for cell type. In B-lymphocytes, the IFNGR1-470del allele suppresses the binding of a approximately 35 kDa nuclear protein and acts to increase reporter gene expression. In epithelial cells, the same allele acts to decrease gene expression and suppresses the binding of approximately 90 kDa STAT-1 and STAT-2 proteins. In T-lymphocytes, this allele causes only subtle differences in nuclear protein binding and has no significant effect on gene expression. These findings suggest a mechanism by which a single genetic variant may cause a broad range of phenotypic consequences.

Cell Line, Tumor↗

PubMed2006

Haplotype homozygosity and derived alleles in the human genome.

Haplotype-based techniques are being used to estimate the relative age of alleles--particularly in screening loci for signals of recent positive selection--but does this approach capture even coarse age differences? Using simulations and empirical data from the International HapMap Project, we show that a simple pairwise metric of haplotype homozygosity gives significantly higher mean values for human single-nucleotide-polymorphism alleles that appear to be derived than for those that appear to be ancestral, as determined by comparison with the chimpanzee genome. Our results support the use of haplotype-based techniques, such as extended haplotypic homozygosity, to assess the age of alleles.

Age Factors↗

PubMed2006

Searching for the regulators of human gene expression.

Many common human traits are believed to be a composite reflection of multiple genetic and non-genetic factors and the genetic contribution is consequently often difficult to characterise. Recent advances suggest that subtle variation in the regulation of gene expression may contribute to complex human traits. In two reports, Cheung and colleagues scale up human genetics analysis to an impressive level in a genome-wide search for the regulators of gene expression. They perform linkage analysis on expression profiles for over 3,500 genes and then employ the HapMap resource to take positive findings through to association studies at the genome-wide level. This work gives new insights into the complexities of gene regulation and the plausibility of genome-wide study design.

Cell Line↗

PubMed2005

Screening for recently selected alleles by analysis of human haplotype similarity.

There is growing interest in the use of haplotype-based methods for detecting recent selection. Here, we describe a method that uses a sliding window to estimate similarity among the haplotypes associated with any given single-nucleotide polymorphism (SNP) allele. We used simulations of natural selection to provide estimates of the empirical power of the method to detect recently selected alleles and found it to be comparable in power to the popular long-range haplotype test and more powerful than methods based on nucleotide diversity. We then applied the method to a recently selected allele--the sickle mutation at the HBB locus--and found it to have a signal of selection that was significantly stronger than that of simulated models both with and without strong selection. Using this method, we also evaluated >4,000 SNPs on chromosome 20, indicating the applicability of the method to regional data sets.

Alleles↗

PubMed2005

Investigation of familial segregation of hyperreactive malarial splenomegaly in Kumasi, Ghana.

Hyperreactive malarial splenomegaly (HMS), a common cause of massive splenomegaly in malaria-endemic regions, is defined as persistent splenomegaly without demonstrable underlying disease. Previous studies have found HMS more frequently in certain tribes in Papua New Guinea, Uganda and Nigeria, with strong familial associations in Uganda and Papua New Guinea. This case-control study aimed to determine the extent of familial association of splenomegaly and the pattern of segregation of the condition in families in Ghana. It involved 22 HMS cases with 99 relatives, and 15 population controls of similar socio-economic background with 51 relatives. The pedigree of each family was recorded. Clinical and laboratory data were collected on all participants, including the presence and degree of splenomegaly. Relatives with splenomegaly were identified for 27% of HMS cases and for 6.7% of population controls (P=0.04). There were significant differences in the IgM levels, which were higher (P=0.005), and the haemoglobin levels, which were lower (P=0.009), in cases compared with controls. In Ghana, relatives of HMS cases are more likely to have splenomegaly than population controls, but with no obvious pattern of Mendelian segregation. HMS aetiology in Ghana is likely to be complex, involving multiple genetic and environmental factors.

Adult↗

PubMed2005

How malaria has affected the human genome and what human genetics can teach us about malaria.

Malaria is a major killer of children worldwide and the strongest known force for evolutionary selection in the recent history of the human genome. The past decade has seen growing evidence of ethnic differences in susceptibility to malaria and of the diverse genetic adaptations to malaria that have arisen in different populations: epidemiological confirmation of the hypotheses that G6PD deficiency, alpha+ thalassemia, and hemoglobin C protect against malaria mortality; the application of novel haplotype-based techniques demonstrating that malaria-protective genes have been subject to recent positive selection; the first genetic linkage maps of resistance to malaria in experimental murine models; and a growing number of reported associations with resistance and susceptibility to human malaria, particularly in genes involved in immunity, inflammation, and cell adhesion. The challenge for the next decade is to build the global epidemiological infrastructure required for statistically robust genomewide association analysis, as a way of discovering novel mechanisms of protective immunity that can be used in the development of an effective malaria vaccine.

Animals↗

PubMed2005

Genomewide analysis of the host response to malaria in Kenyan children.

Malaria is a global problem, and there is a critical need for further understanding of the disease process. When malarial parasites invade and develop within the bloodstream, they stimulate a profound host response whose main clinical sign is fever. To explore this response, we measured host gene expression in whole blood from Kenyan children hospitalized with either acute malaria or other febrile illnesses. Genomewide analysis of expression identified 2 principal gene-expression profiles related to neutrophil and erythroid activity. In addition to these general acute responses, a third gene-expression profile was associated with host parasitemia; mediators of erythrophagocytosis and cellular stress were notable components of this response. The delineation of subjects on the basis of patterns of gene expression provides a molecular perspective of the host response to malaria and further functional insight into the underlying processes of pathogenesis.

Child↗

PubMed2005

The complexity of genetic variation in a simple immune system.

Fruitflies derived from a wild population vary in their resistance to infection with the bacterial pathogen Serratia marcescens. A survey of nucleotide diversity in 21 genes involved in innate immunity concluded that 16 genes had polymorphisms associated with resistance to this specific pathogen. However, the effects of individual polymorphisms on the resistance phenotype were modest, and epistatic interactions appeared to be common. What might these findings tell us about genetic resistance to infection in humans?

Animals↗

PubMed2004

Allele-specific repression of lymphotoxin-alpha by activated B cell factor-1.

Genetic variation at the human LTA locus, encoding lymphotoxin-alpha, is associated with susceptibility to myocardial infarction, asthma and other diseases. By detailed haplotypic analysis of the locus, we identified a single-nucleotide polymorphism (SNP) at LTA+80 as a main predictor of LTA protein production by human B cells. We found that activated B-cell factor-1 (ABF-1) binds to this site in vitro and suppresses reporter gene expression, but only in the presence of the LTA+80A allele. Using haplotype-specific chromatin immunoprecipitation, we confirmed that ABF-1 is preferentially recruited to the low-producer allele in vivo. These findings provide a molecular model of how LTA expression may be genetically regulated by allele-specific recruitment of the transcriptional repressor ABF-1.

Alleles↗

PubMed2004

Quantitative high-throughput analysis of transcription factor binding specificities.

We present a general high-throughput approach to accurately quantify DNA-protein interactions, which can facilitate the identification of functional genetic polymorphisms. The method tested here on two structurally distinct transcription factors (TFs), NF-kappaB and OCT-1, comprises three steps: (i) optimized selection of DNA variants to be tested experimentally, which we show is superior to selecting variants at random; (ii) a quantitative protein-DNA binding assay using microarray and surface plasmon resonance technologies; (iii) prediction of binding affinity for all DNA variants in the consensus space using a statistical model based on principal coordinates analysis. For the protein-DNA binding assay, we identified a polyacrylamide/ester glass activation chemistry which formed exclusive covalent bonds with 5'-amino-modified DNA duplexes and hindered non-specific electrostatic attachment of DNA. Full accessibility of the DNA duplexes attached to polyacrylamide-modified slides was confirmed by the high degree of data correlation with the electromobility shift assay (correlation coefficient 93%). This approach offers the potential for high-throughput determination of TF binding profiles and predicting the effects of single nucleotide polymorphisms on TF binding affinity. New DNA binding data for OCT-1 are presented.

Acrylic Resins↗

PubMed2004

Improved allelic differentiation using sequence-specific oligonucleotide hybridization incorporating an additional base-analogue mismatch.

Sequence-specific oligonucleotide hybridization (SSOH, 'dot-blotting') is a widely employed method of typing single nucleotide polymorphisms (SNPs), but it is often compromised by lack of allelic differentiation. We describe a novel improvement to SSOH that incorporates an additional mismatch into the oligonucleotide probe using the universal base analogue 3-nitropyrrole. This method greatly increases allelic differentiation compared to standard SSOH where oligonucleotides contain only SNP-defining base changes. Moreover, stringency of the hybridisation is predictably maintained over a wide range of temperatures, which can be calculated empirically, thus facilitating the genotyping of multiple SNPs using similar conditions. This improved method increases the usefulness of hybridisation-based methods of rapid genotyping of SNPs and may have implications for array methodologies.

Alleles↗

PubMed2004

Response of the splenic dendritic cell population to malaria infection.

Dendritic cells, particularly those residing in the spleen, are thought to orchestrate acquired immunity to malaria, but it is not known how the splenic dendritic cell population responds to malaria infection and how this response compares with the responses of other antigen-presenting cells. We investigated this question for Plasmodium chabaudi AS infection in C57BL/6 mice. We found that dendritic cells, defined here by the CD11c marker, migrated from the marginal zone of the spleen into the CD4(+) T-cell area within 5 days after parasites entered the bloodstream. This contrasted with the results observed for the macrophage and B-cell populations, which expanded greatly but did not show any comparable migration. Over the same time period dendritic cells showed upregulation of CD40, CD54, and CD86 costimulatory molecules that are required for successful T-cell activation. In dendritic cells, the peak intracellular gamma interferon expression (as shown by fluorescence-activated cell sorting) was on day 5, 2 days earlier than the peak expression in B-cells or macrophages. These findings show that splenic dendritic cells are actively engaged in the earliest phase of malarial infection in vivo and are likely to be critical in shaping the subsequent immune response.

Animals↗

PubMed2003

Haplotype mapping of the bronchiolitis susceptibility locus near IL8.

Susceptibility to viral bronchiolitis, the commonest cause of infant admissions to hospital in the industrialised world, is associated with polymorphism at the IL8 locus. Here we map the genomic boundaries of the disease association by case-control analysis and TDT in 580 affected UK infants. Markers for association mapping were chosen after determining patterns of linkage disequilibrium across the surrounding region of chromosome 4q, a 550-kb segment containing nine genes, extending from AFP to PPBP. The region has three major clusters of high linkage disequilibrium and is notable for its low haplotypic diversity. We exclude adjacent chemokine genes as the cause of the association, and identify a disease-associated haplotype that spans a 250-kb region from AFM to IL8. In between these two genes there is only one structural feature of interest, a novel gene RASSF6, which is predicted to encode a Ras effector protein.

Base Sequence↗

PubMed2003

Haplotypic analysis of the TNF locus by association efficiency and entropy.

BACKGROUND: To understand the causal basis of TNF associations with disease, it is necessary to understand the haplotypic structure of this locus. We genotyped 12 single-nucleotide polymorphisms (SNPs) distributed over 4.3 kilobases in 296 healthy, unrelated Gambian and Malawian adults. We generated 592 high-quality haplotypes by integrating family- and population-based reconstruction methods. RESULTS: We found 32 different haplotypes, of which 13 were shared between the two populations. Both populations were haplotypically diverse (gene diversity = 0.80, Gambia; 0.85, Malawi) and significantly differentiated (p < 10-5 by exact test). More than a quarter of marker pairs showed evidence of intragenic recombination (29% Gambia; 27% Malawi). We applied two new methods of analyzing haplotypic data: association efficiency analysis (AEA), which describes the ability of each SNP to detect every other SNP in a case-control scenario; and the entropy maximization method (EMM), which selects the subset of SNPs that most effectively dissects the underlying haplotypic structure. AEA revealed that many SNPs in TNF are poor markers of each other. The EMM showed that 8 of 12 SNPs (Gambia) and 7 of 12 SNPs (Malawi) are required to describe 95% of the haplotypic diversity. CONCLUSIONS: The TNF locus in the Gambian and Malawi sample is haplotypically diverse and has a rich history of intragenic recombination. As a consequence, a large proportion of TNF SNPs must be typed to detect a disease-modifying SNP at this locus. The most informative subset of SNPs to genotype differs between the two populations.

Adult↗