Biomedical subjects

Mathew T Pletcher

Publications and source records attributed to Mathew T Pletcher.

15 recordsLinked to original sources

PubMed2006

Comparative analysis of haplotype association mapping algorithms.

BACKGROUND: Finding the genetic causes of quantitative traits is a complex and difficult task. Classical methods for mapping quantitative trail loci (QTL) in miceuse an F2 cross between two strains with substantially different phenotype and an interval mapping method to compute confidence intervals at each position in the genome. This process requires significant resources for breeding and genotyping, and the data generated are usually only applicable to one phenotype of interest. Recently, we reported the application of a haplotype association mapping method which utilizes dense genotyping data across a diverse panel of inbred mouse strains and a marker association algorithm that is independent of any specific phenotype. As the availability of genotyping data grows in size and density, analysis of these haplotype association mapping methods should be of increasing value to the statistical genetics community. RESULTS: We describe a detailed comparative analysis of variations on our marker association method. In particular, we describe the use of inferred haplotypes from adjacent SNPs, parametric and nonparametric statistics, and control of multiple testing error. These results show that nonparametric methods are slightly better in the test cases we study, although the choice of test statistic may often be dependent on the specific phenotype and haplotype structure being studied. The use of multi-SNP windows to infer local haplotype structure is critical to the use of a diverse panel of inbred strains for QTL mapping. Finally, because the marginal effect of any single gene in a complex disease is often relatively small, these methods require the use of sensitive methods for controlling family-wise error. We also report our initial application of this method to phenotypes cataloged in the Mouse Phenome Database. CONCLUSION: The use of inbred strains of mice for QTL mapping has many advantages over traditional methods. However, there are also limitations in comparison to the traditional linkage analysis from F2 and RI lines. Application of these methods requires careful consideration of algorithmic choices based on both theoretical and practical factors. Our findings suggest general guidelines, though a complete evaluation of these methods can only be performed as more genetic data in complex diseases becomes available.

Algorithms↗

PubMed2006

Marked interindividual variability in the response to selective inhibitors of cyclooxygenase-2.

BACKGROUND & AIMS: Variability in response to drugs may influence both efficacy and safety. Cyclooxygenase (COX)-2 inhibitors pose a cardiovascular risk by potentially increasing the likelihood of thrombosis, hypertension, and atherogenesis. Differences between individuals in the response to COX-2 inhibitors would be expected to influence their susceptibility to cardiovascular complications. We examined the variability in degree and selectivity of COX-2 inhibition in humans in response to celecoxib and rofecoxib. METHODS: Fifty healthy volunteers received placebo, rofecoxib (25 mg), and celecoxib (200 mg), randomized by order. COX-1 and COX-2 inhibition was determined using ex vivo and in vivo indices of enzymatic activity. A subset of 5 individuals underwent 5 replicate studies to estimate variability in drug response both within and between subjects. RESULTS: Despite the higher COX-2 selectivity of rofecoxib in vitro, the average selectivity attained by 25 mg rofecoxib and 200 mg celecoxib in vivo were not different. However, there was considerable variability at an individual level in the degree of COX-2 inhibition and selectivity attained by both drugs. Approximately one third of the variability was attributable to differences between individuals, suggesting the contribution of genetic sources of variance, such as candidate polymorphisms detected in COX-1 and CYP2C9. CONCLUSIONS: The actual degree of selectivity for inhibition of COX-2 achieved by the coxibs relates both to chemical properties of the drug and to factors within an individual that modulate drug response. These sources of variability might be exploited to identify patients uniquely susceptible to benefit or at developing risk of cardiovascular complications.

Adult↗

PubMed2005

Quantitative trait loci that determine BMD in C57BL/6J and 129S1/SvImJ inbred mice.

UNLABELLED: BMD is highly heritable; however, little is known about the genes. To identify loci controlling BMD, we conducted a QTL analysis in a (B6 x 129) F2 population of mice. We report on additional QTLs and also narrow one QTL by combining the data from multiple crosses and through haplotype analysis. INTRODUCTION: Previous studies have identified quantitative trait loci (QTL) that determine BMD in mice; however, identification of genes underlying QTLs is impeded by the large size of QTL regions. MATERIALS AND METHODS: To identify loci controlling BMD, we performed a QTL analysis of 291 (B6 x 129) F2 females. Total body and vertebral areal BMD (aBMD) were determined by peripheral DXA when mice were 20 weeks old and had consumed a high-fat diet for 14 weeks. RESULTS AND CONCLUSIONS: Two QTLs were common for both total body and vertebral aBMD: Bmd20 on chromosome (Chr) 6 (total aBMD; peak cM 26, logarithm of odds [LOD] 3.8, and vertebral aBMD; cM 32, LOD 3.6) and Bmd22 on Chr 1 (total aBMD; cM 104, LOD 2.5, and vertebral aBMD; cM 98, LOD 2.6). A QTL on Chr 10 (Bmd21, cM 68, LOD 3.0) affected total body aBMD and a QTL on Chr 7 (Bmd9, cM 44, LOD 2.7) affected vertebral aBMD. A pairwise genome-wide search did not reveal significant gene-gene interactions. Collectively, the QTLs accounted for 21.6% of total aBMD and 17.3% of vertebral aBMD of the F(2) population variances. Bmd9 was previously identified in a cross between C57BL/6J and C3H/HeJ mice, and we narrowed this QTL from 34 to 22 cM by combining the data from these crosses. By examining the Bmd9 region for conservation of ancestral alleles among the low allele strains (129S1/SvImJ and C3H/HeJ) that differed from the high allele strain (C57BL/6J), we further narrowed the region to approximately 9.9 cM, where the low allele strains share a common haplotype. Identifying the genes for these QTLs will enhance our understanding of skeletal biology.

Animals↗

PubMed2005

Tumor suppressor in lung cancer 1 (TSLC1) alters tumorigenic growth properties and gene expression.

BACKGROUND: Introduction of cDNA or genomic clones of the tumor suppressor in lung cancer 1 (TSLC1) gene into the non-small cell lung cancer line, A549, reverses tumorigenic growth properties of these cells. These results and the observation that TSLC1 is down-regulated in a number of tumors suggest that TSLC1 functions as a critical switch mediating repression of tumorigenesis. RESULTS: To investigate this mechanism, we compared growth properties of A549 with the TSLC1-containing derivative. We found a G1/S phase transition delay in 12.2. Subtractive hybridization, quantitative PCR, and TranSignal Protein/DNA arrays were used to identify genes whose expression changed when TSLC1 was up-regulated. Members of common G1/S phase regulatory pathways such as TP53, MYC, RB1 and HRAS were not differentially expressed, indicating that TSLC1 may function through an alternative pathway(s). A number of genes involved in cell proliferation and tumorigenesis were differentially expressed, notably genes in the Ras-induced senescence pathway. We examined expression of several of these key genes in human tumors and normal lung tissue, and found similar changes in expression, validating the physiological relevance of the A549 and 12.2 cell lines. CONCLUSION: Gene expression and cell cycle differences provide insights into potential downstream pathways of TSLC1 that mediate the suppression of tumor properties in A549 cells.

Carcinoma, Non-Small-Cell Lung↗

PubMed2005

Uncovering regulatory pathways that affect hematopoietic stem cell function using 'genetical genomics'.

We combined large-scale mRNA expression analysis and gene mapping to identify genes and loci that control hematopoietic stem cell (HSC) function. We measured mRNA expression levels in purified HSCs isolated from a panel of densely genotyped recombinant inbred mouse strains. We mapped quantitative trait loci (QTLs) associated with variation in expression of thousands of transcripts. By comparing the physical transcript position with the location of the controlling QTL, we identified polymorphic cis-acting stem cell genes. We also identified multiple trans-acting control loci that modify expression of large numbers of genes. These groups of coregulated transcripts identify pathways that specify variation in stem cells. We illustrate this concept with the identification of candidate genes involved with HSC turnover. We compared expression QTLs in HSCs and brain from the same mice and identified both shared and tissue-specific QTLs. Our data are accessible through WebQTL, a web-based interface that allows custom genetic linkage analysis and identification of coregulated transcripts.

Carrier Proteins↗

PubMed2005

c-Myb and p300 regulate hematopoietic stem cell proliferation and differentiation.

Precise control of hematopoietic stem cell (HSC) proliferation and differentiation is needed to maintain a lifetime supply of blood cells. Using genome-wide ENU mutagenesis and phenotypic screening, we have identified a mouse line that harbors a point mutation in the transactivation (TA) domain of the transcription factor c-Myb (M303V), which reduces c-Myb-dependent TA by disrupting its interaction with the transcriptional coactivator p300. The biological consequences of the c-Myb(M303V/M303V) mutation include thrombocytosis, megakaryocytosis, anemia, lymphopenia, and the absence of eosinophils. Detailed analysis of hematopoiesis in c-Myb(M303V/M303V) mice reveals distinct blocks in T cell, B cell, and red blood cell development, as well as a remarkable 10-fold increase in the number of HSCs. Cell cycle analyses show that twice as many HSCs from c-Myb(M303V/M303V) animals are actively cycling. Thus c-Myb, through interaction with p300, controls the proliferation and differentiation of hematopoietic stem and progenitor cells.

Animals↗

PubMed2004

Use of a dense single nucleotide polymorphism map for in silico mapping in the mouse.

Rapid expansion of available data, both phenotypic and genotypic, for multiple strains of mice has enabled the development of new methods to interrogate the mouse genome for functional genetic perturbations. In silico mapping provides an expedient way to associate the natural diversity of phenotypic traits with ancestrally inherited polymorphisms for the purpose of dissecting genetic traits. In mouse, the current single nucleotide polymorphism (SNP) data have lacked the density across the genome and coverage of enough strains to properly achieve this goal. To remedy this, 470,407 allele calls were produced for 10,990 evenly spaced SNP loci across 48 inbred mouse strains. Use of the SNP set with statistical models that considered unique patterns within blocks of three SNPs as an inferred haplotype could successfully map known single gene traits and a cloned quantitative trait gene. Application of this method to high-density lipoprotein and gallstone phenotypes reproduced previously characterized quantitative trait loci (QTL). The inferred haplotype data also facilitates the refinement of QTL regions such that candidate genes can be more easily identified and characterized as shown for adenylate cyclase 7.

Adenylyl Cyclases↗

PubMed2004

Inositol (1,4,5) trisphosphate 3 kinase B controls positive selection of T cells and modulates Erk activity.

The mechanisms governing positive selection of T cells in the thymus are still incompletely understood. Here, we describe a N-ethyl-N-nitrosourea induced recessive mouse mutant, Ms. T-less, which lacks T cells in the peripheral blood because of a complete block of thymocyte development at the CD4(+)CD8(+) stage. Single nucleotide polymorphism mapping and candidate gene sequencing revealed a nonsense mutation in the inositol (1,4,5) trisphosphate 3 kinase B (Itpkb) gene in Ms. T-less mice. Accordingly, Ms. T-less thymocytes do not show detectable expression of Itpkb protein and have drastically reduced basal inositol (1,4,5) trisphosphate kinase activity. Itpkb converts inositol (1,4,5) trisphosphate to inositol (1,3,4,5) tetrakisphosphate, soluble second messengers that have been implicated in Ca(2+) signaling. Surprisingly, Ca(2+) responses show no significant differences between wild type (WT) and mutant thymocytes. However, extracellular signal-regulated kinase (Erk) activation in response to suboptimal antigen receptor stimulation is attenuated in Ms. T-less thymocytes, suggesting a role for Itpkb in linking T cell receptor signaling to efficient and sustained Erk activation.

Animals↗

PubMed2004

Transcript level alterations reflect gene dosage effects across multiple tissues in a mouse model of down syndrome.

Human trisomy 21, which results in Down syndrome (DS), is one of the most complicated congenital genetic anomalies compatible with life, yet little is known about the molecular basis of DS. It is generally accepted that chromosome 21 (Chr21) transcripts are overexpressed by about 50% in cells with an extra copy of this chromosome. However, this assumption is difficult to test in humans due to limited access to tissues, and direct support for this idea is available for only a few Chr21 genes or in a limited number of tissues. The Ts65Dn mouse is widely used as a model for studies of DS because it is at dosage imbalance for the orthologs of about half of the 284 Chr21 genes. Ts65Dn mice have several features that directly parallel developmental anomalies of DS. Here we compared the expression of 136 mouse orthologs of Chr21 genes in nine tissues of the trisomic and euploid mice. Nearly all of the 77 genes which are at dosage imbalance in Ts65Dn showed increased transcript levels in the tested tissues, providing direct support for a simple model of increased transcription proportional to the gene copy number. However, several genes escaped this rule, suggesting that they may be controlled by additional tissue-specific regulatory mechanisms revealed in the trisomic situation.

Animals↗

PubMed2003

Global disruption of the cerebellar transcriptome in a Down syndrome mouse model.

Trisomy 21 (Down syndrome) results in cerebellar dysmorphology with direct parallels in the Ts65Dn mouse. Despite pronounced changes in morphology, cerebellar function is not markedly different. As a first test of whether those cerebellar cells that have survived to adulthood in trisomic mice are equivalent to euploid cells, we used microarrays to assess the trisomic and euploid cerebella. Trisomic and euploid transcriptomes were robustly distinguished. Changes in expression of individual genes were very subtle, but the differences in respective transcriptome phenotypes extended deeply into the set of nearly 7000 probes (genes) located throughout the genome. In contrast to deterministic models of gene action in trisomy, examination of the discriminating genes in two independent experiments suggests that the global perturbation includes a significant stochastic component. Thus, dosage imbalance of 124 genes in Ts65Dn mice alters the expression of thousands of genes to create a variable trisomic transcriptome. This global destabilization has important implications for approaches to ameliorative therapies in Down syndrome.

Animals↗

PubMed2003

Melanopsin is required for non-image-forming photic responses in blind mice.

Although mice lacking rod and cone photoreceptors are blind, they retain many eye-mediated responses to light, possibly through photosensitive retinal ganglion cells. These cells express melanopsin, a photopigment that confers this photosensitivity. Mice lacking melanopsin still retain nonvisual photoreception, suggesting that rods and cones could operate in this capacity. We observed that mice with both outer-retinal degeneration and a deficiency in melanopsin exhibited complete loss of photoentrainment of the circadian oscillator, pupillary light responses, photic suppression of arylalkylamine-N-acetyltransferase transcript, and acute suppression of locomotor activity by light. This indicates the importance of both nonvisual and classical visual photoreceptor systems for nonvisual photic responses in mammals.

Animals↗

PubMed2003

Genome-wide single-nucleotide polymorphism analysis defines haplotype patterns in mouse.

The nature and organization of polymorphisms, or differences, between genomes of individuals are of great interest, because these variations can be associated with or even underlie phenotypic traits, including disease susceptibility. To gain insight into the genetic and evolutionary factors influencing such biological variation, we have examined the arrangement (haplotype) of single-nucleotide polymorphisms across the genomes of eight inbred strains of mice. These analyses define blocks of high or low diversity, often extending across tens of megabases that are delineated by abrupt transitions. These observations provide a striking contrast to the haplotype structure of the human genome.

Animals↗

PubMed2002

Genetic and comparative mapping of genes dysregulated in mouse hearts lacking the Hand2 transcription factor gene.

The helix-loop-helix transcription factor HAND2 plays a vital role in the development of the heart, limb, facies, and other neural crest-derived structures. We used differential display analysis to identify 33 putative HAND2-regulated ESTs that are differentially expressed in Hand2(-/-) vs wild-type mice. We determined the positions on mouse and human genetic maps of 29 of these by using the T31 mouse Radiation Hybrid panel, comparison to human genomic sequence, and comparative mapping. We examined the conserved chromosomal locations for phenotypes that involve development of heart, face, and limb structures that are affected by HAND2. One EST mapped to a region of conserved synteny between mouse chromosome 2 and human chromosome 10p. RACE analysis extended the sequence and identified this cDNA as the mouse ortholog of human nebulette, an actin-binding protein expressed in fetal heart. Nebulette was shown to be deleted in DiGeorge Syndrome 2 patients with the proximal deletion of human 10p13-p14 that is associated with cardiac and craniofacial abnormalities.

Animals↗

PubMed2002

Comparative genomic sequence analysis of the human chromosome 21 Down syndrome critical region.

Comprehensive knowledge of the gene content of human chromosome 21 (HSA21) is essential for understanding the etiology of Down syndrome (DS). Here we report the largest comparison of finished mouse and human sequence to date for a 1.35-Mb region of mouse chromosome 16 (MMU16) that corresponds to human chromosome 21q22.2. This includes a portion of the commonly described "DS critical region," thought to contain a gene or genes whose dosage imbalance contributes to a number of phenotypes associated with DS. We used comparative sequence analysis to construct a DNA feature map of this region that includes all known genes, plus 144 conserved sequences > or =100 bp long that show > or =80% identity between mouse and human but do not match known exons. Twenty of these have matches to expressed sequence tag and cDNA databases, indicating that they may be transcribed sequences from chromosome 21. Eight putative CpG islands are found at conserved positions. Models for two human genes, DSCR4 and DSCR8, are not supported by conserved sequence, and close examination indicates that low-level transcripts from these loci are unlikely to encode proteins. Gene prediction programs give different results when used to analyze the well-conserved regions between mouse and human sequences. Our findings have implications for evolution and for modeling the genetic basis of DS in mice.

Animals↗

PubMedDate not supplied

A mutant mouse with a highly specific contextual fear-conditioning deficit found in an N-ethyl-N-nitrosourea (ENU) mutagenesis screen.

Targeted mutagenesis in mice has shown that genes from a wide variety of gene families are involved in memory formation. The efficient identification of genes involved in learning and memory could be achieved by random mutagenesis combined with high-throughput phenotyping. Here, we provide the first report of a mutagenesis screen that has generated memory mutants in the mouse. We tested a group of N-ethyl-N-nitrosourea (ENU) mutagenized mice in the conditioned fear paradigm. We screened for both dominant and recessive mutations that caused impairments in contextual or tone fear conditioning. Heritability testing confirmed three fear conditioning mutants, i.e., Forgetful, Slowlearner, and Scatterbrain. All three have a learning or short-term memory deficit in contextual fear conditioning. Forgetful was further characterized and showed a highly specific phenotype. The contextual fear-conditioning deficit was apparent when Forgetful was trained with tone-shock pairings, but not when trained with shock alone. The deficit was not due to changes in shock sensitivity or anxiety. Forgetful was not impaired in two other memory tests (hidden platform version of Morris water maze and object recognition). Our data show that a mutagenesis screen can generate mutant mice with highly specific memory phenotypes that can supplement existing mice with targeted mutations. Mapping of Slowlearner found linkage to a region of chromosome 12 (LOD score of 6.5 close to D12Mit171), which suggests that ENU mutants should enable the positional cloning of genes involved in memory formation.

Animals↗