Biomedical subjects

Jonathan Pevsner

Publications and source records attributed to Jonathan Pevsner.

At least 19 recordsLinked to original sources

PubMed2007

SNPchip: R classes and methods for SNP array data.

UNLABELLED: High-density single nucleotide polymorphism microarrays (SNP chips) provide information on a subject's genome, such as copy number and genotype (heterozygosity/homozygosity) at a SNP. While fluorescence in situ hybridization and karyotyping reveal many abnormalities, SNP chips provide a higher resolution map of the human genome that can be used to detect, e.g., aneuploidies, microdeletions, microduplications and loss of heterozygosity (LOH). As a variety of diseases are linked to such chromosomal abnormalities, SNP chips promise new insights for these diseases by aiding in the discovery of such regions, and may suggest targets for intervention. The R package SNPchip contains classes and methods useful for storing, visualizing and analyzing high density SNP data. Originally developed from the SNPscan web-tool, SNPchip utilizes S4 classes and extends other open source R tools available at Bioconductor. This has numerous advantages, including the ability to build statistical models for SNP-level data that operate on instances of the class, and to communicate with other R packages that add additional functionality. AVAILABILITY: The package is available from the Bioconductor web page at www.bioconductor.org. SUPPLEMENTARY INFORMATION: The supplementary material as described in this article (case studies, installation guidelines and R code) is available from http://biostat.jhsph.edu/~iruczins/publications/sm/

Models, Statistical↗

PubMed2006

Analysis and visualization of chromosomal abnormalities in SNP data with SNPscan.

BACKGROUND: A variety of diseases are caused by chromosomal abnormalities such as aneuploidies (having an abnormal number of chromosomes), microdeletions, microduplications, and uniparental disomy. High density single nucleotide polymorphism (SNP) microarrays provide information on chromosomal copy number changes, as well as genotype (heterozygosity and homozygosity). SNP array studies generate multiple types of data for each SNP site, some with more than 100,000 SNPs represented on each array. The identification of different classes of anomalies within SNP data has been challenging. RESULTS: We have developed SNPscan, a web-accessible tool to analyze and visualize high density SNP data. It enables researchers (1) to visually and quantitatively assess the quality of user-generated SNP data relative to a benchmark data set derived from a control population, (2) to display SNP intensity and allelic call data in order to detect chromosomal copy number anomalies (duplications and deletions), (3) to display uniparental isodisomy based on loss of heterozygosity (LOH) across genomic regions, (4) to compare paired samples (e.g. tumor and normal), and (5) to generate a file type for viewing SNP data in the University of California, Santa Cruz (UCSC) Human Genome Browser. SNPscan accepts data exported from Affymetrix Copy Number Analysis Tool as its input. We validated SNPscan using data generated from patients with known deletions, duplications, and uniparental disomy. We also inspected previously generated SNP data from 90 apparently normal individuals from the Centre d'Etude du Polymorphisme Humain (CEPH) collection, and identified three cases of uniparental isodisomy, four females having an apparently mosaic X chromosome, two mislabelled SNP data sets, and one microdeletion on chromosome 2 with mosaicism from an apparently normal female. These previously unrecognized abnormalities were all detected using SNPscan. The microdeletion was independently confirmed by fluorescence in situ hybridization, and a region of homozygosity in a UPD case was confirmed by sequencing of genomic DNA. CONCLUSION: SNPscan is useful to identify chromosomal abnormalities based on SNP intensity (such as chromosomal copy number changes) and heterozygosity data (including regions of LOH and some cases of UPD). The program and source code are available at the SNPscan website http://pevsnerlab.kennedykrieger.org/snpscan.htm.

Base Sequence↗

PubMed2005

Primary and secondary transcriptional effects in the developing human Down syndrome brain and heart.

BACKGROUND: Down syndrome, caused by trisomic chromosome 21, is the leading genetic cause of mental retardation. Recent studies demonstrated that dosage-dependent increases in chromosome 21 gene expression occur in trisomy 21. However, it is unclear whether the entire transcriptome is disrupted, or whether there is a more restricted increase in the expression of those genes assigned to chromosome 21. Also, the statistical significance of differentially expressed genes in human Down syndrome tissues has not been reported. RESULTS: We measured levels of transcripts in human fetal cerebellum and heart tissues using DNA microarrays and demonstrated a dosage-dependent increase in transcription across different tissue/cell types as a result of trisomy 21. Moreover, by having a larger sample size, combining the data from four different tissue and cell types, and using an ANOVA approach, we identified individual genes with significantly altered expression in trisomy 21, some of which showed this dysregulation in a tissue-specific manner. We validated our microarray data by over 5,600 quantitative real-time PCRs on 28 genes assigned to chromosome 21 and other chromosomes. Gene expression values from chromosome 21, but not from other chromosomes, accurately classified trisomy 21 from euploid samples. Our data also indicated functional groups that might be perturbed in trisomy 21. CONCLUSIONS: In Down syndrome, there is a primary transcriptional effect of disruption of chromosome 21 gene expression, without a pervasive secondary effect on the remaining transcriptome. The identification of dysregulated genes and pathways suggests molecular changes that may underlie the Down syndrome phenotypes.

Astrocytes↗

PubMed2005

The use of genomic microarrays to study chromosomal abnormalities in mental retardation.

Mental retardation affects 2 to 3% of the US population. It is defined by broad criteria, including significantly subaverage intelligence, onset by age 18, and impaired function in a group of adaptive skills. A myriad of genetic and environmental causes have been described, but for approximately half of individuals diagnosed with mental retardation the molecular basis remains unknown. Genomic microarrays, also called array comparative genomic hybridization (array CGH), represent one of several novel technologies that allow the detection of chromosomal abnormalities, such as microdeletions and microduplications, in a rapid, high throughput fashion from genomic DNA samples. In one early application of this technology, genomic microarrays have been used to characterize the extent of chromosomal changes in a group of patients diagnosed with one particular type of disorder that causes mental retardation, such as deletion 1p36 syndrome. In another application, DNA samples from individuals with idiopathic mental retardation have been assayed to scan the entire genome in attempts to identify chromosomal changes. Genomic microarrays offer both a genome-wide perspective of chromosomal aberrations as well as higher resolution (to the level of approximately one megabase) compared to alternative available technologies.

Chromosome Aberrations↗

PubMed2004

mVps24p functions in EGF receptor sorting/trafficking from the early endosome.

Yeast Vps24p (vacuolar protein sorting) is part of a protein complex suggested to function in sorting/trafficking during endocytosis. We have characterized a mammalian homolog of the yeast protein, mVps24p, and examined its role in epidermal growth factor receptor trafficking. Endogenous mVps24p was distributed in both cytosol and in puncta and partially colocalized with markers for the trans-Golgi network. Adventitious expression of hrs or a mVps4p mutant deficient in ATPase activity caused a redistribution of both mVps24p and the M6PR to the resultant clustered/enlarged early endosomes. Expression of an mVps24p N-terminal fragment, that interacts with phosphatidylinositol 3,5-bisphosphate but not with mVps4p, produces enlarged early endosomes. More importantly, the mVps24p N-terminal fragment resulted in not only enhanced recycling, but also decreased degradation of the EGF receptor. These findings are consistent with a model in which mVps24p has a role in trafficking from the early endosome.

Amino Acid Sequence↗

PubMed2004

Mitochondrial fission proteins regulate programmed cell death in yeast.

The possibility that single-cell organisms undergo programmed cell death has been questioned in part because they lack several key components of the mammalian cell death machinery. However, yeast encode a homolog of human Drp1, a mitochondrial fission protein that was shown previously to promote mammalian cell death and the excessive mitochondrial fragmentation characteristic of apoptotic mammalian cells. In support of a primordial origin of programmed cell death involving mitochondria, we found that the Saccharomyces cerevisiae homolog of human Drp1, Dnm1, promotes mitochondrial fragmentation/degradation and cell death following treatment with several death stimuli. Two Dnm1-interacting factors also regulate yeast cell death. The WD40 repeat protein Mdv1/Net2 promotes cell death, consistent with its role in mitochondrial fission. In contrast to its fission function in healthy cells, Fis1 unexpectedly inhibits Dnm1-mediated mitochondrial fission and cysteine protease-dependent cell death in yeast. Furthermore, the ability of yeast Fis1 to inhibit mitochondrial fission and cell death can be functionally replaced by human Bcl-2 and Bcl-xL. Together, these findings indicate that yeast and mammalian cells have a conserved programmed death pathway regulated by a common molecular component, Drp1/Dnm1, that is inhibited by a Bcl-2-like function.

Adaptor Proteins, Signal Transducing↗

PubMed2004

Gene expression alterations over large chromosomal regions in cancers include multiple genes unrelated to malignant progression.

In solid tumors, the relationship between DNA copy number and global expression over large chromosomal regions has not been systematically explored. We used a 12,626-gene expression array analysis of head and neck squamous cell carcinoma and normal oral mucosa and annotated gene expression levels to specific chromosomal loci. Expression alterations correlated with reported data using comparative genomic hybridization. When genes with significant differences in expression between normal and malignant lesions, as defined by significance analysis of microarrays (SAM), were compared to nonsignificant genes, similar chromosomal patterns of alteration in expression were noted. Individual tumors underwent microsatellite analysis and chi(2) analysis of expression at 3p and 22q. Significant 3p underexpression and 22q overexpression were found in all primary tumors with 3p and 22q allelic imbalance, respectively, whereas no tumor without allelic imbalance on these chromosomal arms demonstrated expression differences. Loss and gain of chromosomal material in solid cancers can alter gene expression over large chromosomal regions, including multiple genes unrelated to malignant progression.

Allelic Imbalance↗

PubMed2004

Viral Bcl-2 homologs and their role in virus replication and associated diseases.

Cellular Bcl-2 family proteins regulate a critical step in the mammalian programmed cell death pathway by modulating mitochondrial permeability and function. Bcl-2 family proteins are also encoded by several large DNA viruses, including all known gamma herpesviruses, adenoviruses, and several other unrelated viruses. Viral Bcl-2 proteins can prevent cell death but often escape cellular regulatory mechanisms that govern their cellular counterparts. By evading the "altruistic" suicide of infected cells, viruses can ensure replication and propagation in the infected host, but sometimes in surprising ways. Many human cancers and other disorders are associated with viruses that encode Bcl-2 homologs. Here we consider the available mechanistic data for viral compared to cellular Bcl-2 protein function along with relevance to the virus life cycle and human disease states.

Apoptosis↗

PubMed2004

Discovering novel phenotype-selective neurotrophic factors to treat neurodegenerative diseases.

Astrocytes and neurons in the central nervous system (CNS) interact functionally to mediate processes as diverse as neuroprotection, neurogenesis and synaptogenesis. Moreover, the interaction can be homotypic, implying that astrocyte-derived secreted molecules affect their adjacent neurons optimally vs remote neurons. Astrocytes produce neurotrophic and extracellular matrix molecules that affect neuronal growth, development and survival, synaptic development, stabilization and functioning, and neurogenesis. This new knowledge offers the opportunity of developing astrocyte-derived, secreted proteins as a new class of therapeutics specifically to treat diseases of the CNS. However, primary astrocytes proliferate slowly in vitro, and when induced to immortalize by genetic manipulation, tend to lose their phenotype. These problems have limited the development of astrocytes as sources of potential drug candidates. We have successfully developed a method to induce spontaneous immortalization of astrocytes. Gene expression analysis, karyotyping and activity profiling data show that these spontaneously immortalized type-1 astrocyte cell lines retain the properties of their primary parents. The method is generic, such that cell lines can be prepared from any region of the CNS. To date, a library of 70 cell lines from four regions of the CNS: ventral mesencephalon, striatum, cerebral cortex and hippocampus, has been created. A phenotype-selective neurotrophic factor for dopaminergic neurons has been discovered from one of the cell lines (VMCL1). This mesencephalic astrocyte-derived neurotrophic factor (MANF) is a 20 kD, glycosylated, human secreted protein. Homologs of this protein have been identified in 16 other species including C. elegans. These new developments offer the opportunity of creating a library of astrocyte-derived molecules, and developing the ones with the best therapeutic indices for clinical use.

Animals↗

PubMed2004

Progress in the use of microarray technology to study the neurobiology of disease.

The diverse functions of the brain are mediated by neurons and glia whose phenotype is defined by a dynamically maintained set of gene transcripts, or 'transcriptome'. Large-scale analysis of gene expression in postmortem brain using microarray technology has the potential to elucidate molecular changes that occur in disease states. There are unique challenges associated with studies of postmortem brain, including limited sample sizes and variable clinical phenotypes that are typical of complex disorders. Nevertheless, recent microarray-based studies have implicated both individual dysregulated genes and abnormal patterns of gene expression in brain disorders.

Animals↗

PubMed2003

Nitric oxide regulates exocytosis by S-nitrosylation of N-ethylmaleimide-sensitive factor.

Nitric oxide (NO) inhibits vascular inflammation, but the molecular basis for its anti-inflammatory properties is unknown. We show that NO inhibits exocytosis of Weibel-Palade bodies, endothelial granules that mediate vascular inflammation and thrombosis, by regulating the activity of N-ethylmaleimide-sensitive factor (NSF). NO inhibits NSF disassembly of soluble NSF attachment protein receptor (SNARE) complexes by nitrosylating critical cysteine residues of NSF. NO may regulate exocytosis in a variety of physiological processes, including vascular inflammation, neurotransmission, thrombosis, and cytotoxic T lymphocyte cell killing.

Animals↗

PubMed2003

A transcriptional progression model for head and neck cancer.

PURPOSE: A genetic progression model for head and neck squamous cell carcinoma (HNSC) has been established and implies the presence of transcriptional dysregulation as a consequence of accumulation of genetic alterations. Although expression array data have been provided for HNSC, the timing of transcriptional dysregulation in the progression from normal mucosa to dyplastic epithelium to invasive HNSC has not been described. Here, we describe a transcriptional progression model of HNSC. EXPERIMENTAL DESIGN: Expression arrays representing >12,000 genes and expressed sequence tags were used to examine malignant lesions (M), premalignant lesions (PM), distant, histopathologically normal mucosa from patients with premalignant or malignant lesions (MN), and normal mucosa from the upper aerodigestive tract of patients with noncancer diagnoses (N). Significance analysis of microarrays, hierarchical clustering, and principal components analysis was used to identify genes with differential expression patterns. RESULTS: Using a false discovery rate of <5% for significance analysis of microarray, the M group revealed 965 up-regulated and 1106 down-regulated genes relative to the N group. The PM group demonstrated 108 up-regulated and 226 down-regulated genes relative to the N group, whereas the M group demonstrated only 5 up-regulated and 13 down-regulated genes relative to the PM group. Both hierarchical cluster analysis and principal components analysis revealed a consistent separation between the N, PM, and M groups, with a closer association between the PM and M groups. To provide independent validation of the microarray data, quantitative reverse transcription-PCR was performed for a significantly up-regulated gene, integrin alpha 6, correlating well with microarray data (linear regression analysis, P < 0.0001). CONCLUSIONS: Similarly to the genetic progression model of HNSC, this transcriptional model shows that the majority of alterations occurs before the development of malignancy and identifies key targets of transcriptional dysregulation during progression from a normal to a premalignant state and from a premalignant to a malignant state.

Carcinoma↗

PubMed2003

Increased fibronectin expression in sturge-weber syndrome fibroblasts and brain tissue.

Sturge-Weber syndrome (SWS) is a neurocutaneous disorder that presents with a facial port-wine stain and a leptomeningeal angioma. Fibronectin expression regulates angiogenesis and vasculogenesis and participates in brain tissue responses to ischemia and seizures. We therefore hypothesized that abnormal gene expression of fibronectin and other extracellular matrix genes would be found in SWS brain tissue and SWS port-wine skin fibroblasts. Fibronectin gene and protein expression from port-wine-derived fibroblasts were compared with that from normal skin-derived fibroblasts of four individuals with SWS using microarrays, reverse transcriptase-PCR, Western analysis, and immunocytochemistry. Fibronectin gene and/or protein expression from eight SWS surgical brain samples was compared with that in two surgical epilepsy brain samples and six postmortem brain samples using microarrays, reverse transcriptase-PCR, and Western analysis. The gene expression of fibronectin was significantly increased (p < 0.05) in the SWS port-wine-derived fibroblasts compared with that of fibroblasts from SWS normal skin. A trend for increased protein levels of fibronectin in port-wine fibroblasts was found by Western analysis. No difference in the pattern of fibronectin staining was detected. The gene expression of fibronectin was significantly increased (p < 0.05), and a trend for increased fibronectin protein expression was found in the SWS surgical brain samples compared with the postmortem controls. These results suggest a potential role for fibronectin in the pathogenesis of SWS and in the brain's response to chronic ischemic injury in SWS. The reproducible differences in fibronectin gene expression between the SWS port-wine-derived fibroblasts and the SWS normal skin-derived fibroblasts are consistent with the presence of a hypothesized somatic mutation underlying SWS.

Adolescent↗

PubMed2003

Global up-regulation of chromosome 21 gene expression in the developing Down syndrome brain.

Down syndrome (DS) results from complete or partial triplication of human chromosome 21. It is assumed that the neurological and other symptoms are caused by the overexpression of genes on chromosome 21, but this hypothesis has not yet been assessed on a chromosome-wide basis. Here we show that expression of genes localized to chromosome 21 is globally up-regulated in human fetal trisomy 21 cases, both in cerebral cortex extracts and in astrocytic cell lines cultured from cerebral cortex. This abnormal regulation of gene expression is specific to chromosome 21. Our data describe transcriptional changes that are specific to many genes assigned to chromosome 21 and do not directly measure the clinical phenotype of DS. However, it is possible that these gene expression changes ultimately relate to the phenotypic variability of DS.

Animals↗

PubMed2003

Expression of MeCP2 in olfactory receptor neurons is developmentally regulated and occurs before synaptogenesis.

Rett syndrome, a neurodevelopmental disorder hypothesized to be due to defective neuronal maturation, is a result of mutations in the mecp2 gene encoding the transcriptional repressor methyl-CpG binding protein (MeCP2). We utilized the olfactory system, which displays postnatal neurogenesis, as a model to investigate MeCP2 expression during development and after injury. MeCP2 expression increased postnatally, localizing to mature olfactory receptor neurons (ORNs) and sustentacular supporting cells. The timing of MeCP2 expression was defined by using detergent ablation (to remove the ORNs) and unilateral olfactory bulbectomy (to remove the ORN target), both of which increase neurogenesis. MeCP2 expression in the ORNs reached prelesioning levels as cells matured after ablation, whereas expression was not completely restored after bulbectomy, in which functional synaptogenesis cannot occur. Thus, MeCP2 expression correlates with the maturational state of ORNs, and precedes synaptogenesis. Identifying the time window of MeCP2 expression should help further clarify the biological defects in Rett syndrome.

Animals↗

PubMed2003

Development of human protein reference database as an initial platform for approaching systems biology in humans.

Human Protein Reference Database (HPRD) is an object database that integrates a wealth of information relevant to the function of human proteins in health and disease. Data pertaining to thousands of protein-protein interactions, posttranslational modifications, enzyme/substrate relationships, disease associations, tissue expression, and subcellular localization were extracted from the literature for a nonredundant set of 2750 human proteins. Almost all the information was obtained manually by biologists who read and interpreted >300,000 published articles during the annotation process. This database, which has an intuitive query interface allowing easy access to all the features of proteins, was built by using open source technologies and will be freely available at http://www.hprd.org to the academic community. This unified bioinformatics platform will be useful in cataloging and mining the large number of proteomic interactions and alterations that will be discovered in the postgenomic era.

BRCA1 Protein↗

PubMed2003

Clinical variability in Rett syndrome.

The clinical variability of Rett syndrome, associated with mutations in the MECP2 gene, varies from classically symptomatic female patients to asymptomatic female patients, and male patients who have none of the diagnostic features considered pathognomonic of this disease. Multiple factors contribute to this variability. In our studies, mutations closer to the amino-terminus, prior to amino acid 255, led to severe clinical manifestations, such as inability to walk, severe dysphagia, and urinary organic acid abnormalities, compared with mutations toward the carboxyl-terminus. However, we found no correlation between severity and mutation type (missense versus nonsense). Despite the importance of mutation location to clinical severity, the widely varying severity within the same mutation suggests that in females, X-chromosome inactivation or other epigenetic phenomena also have roles in determining severity. We propose that stages 1 and 2 of the disease are a consequence of failed, time-linked, postnatal expression of MeCP2 in cerebellar neurons. This, in association with glutamate N-methyl-D-aspartate receptor-mediated neuroexcitotoxic injury to the differentiating neurons, results in the transient age-specific autistic-like behavior, motor, and cognitive dysfunction associated with these stages.

Animals↗

PubMed2003

MANF: a new mesencephalic, astrocyte-derived neurotrophic factor with selectivity for dopaminergic neurons.

We describe the discovery of a novel, 20 kDa, secreted human protein named mesencephalic astrocyte-derived neurotrophic factor, or MANF. The homologous, native molecule was initially derived from a rat mesencephalic type-1 astrocyte cell line and recombinant MANF subcloned from a cDNA encoding human arginine-rich protein. MANF selectively protects nigral dopaminergic neurons, versus GABAergic or serotonergic neurons. The discovery of MANF marks a more systematic approach in the search for astrocyte-derived, secreted proteins that selectively protect specific neuronal phenotypes. Compared to glial cell line-derived neurotrophic factor (GDNF) and brain-derived neurotrophic factor (BDNF), MANF was more selective in the protection of dopaminergic neurons at lower (0.05-0.25 ng/mL) and middle (0.5-2.5 ng/mL) concentrations: MANF>GDNF>BDNF. GDNF was more selective at higher concentrations (25-50 ng/ml): GDNF>MANF>BDNF. Two domains in MANF of 39-AA and 109-AA respectively, and eight cysteines are conserved from C. elegans to man. MANF is encoded by a 4.3 Kb gene with 4 exons, and is located on the short arm of human chromosome 3. The secondary structure is dominated by alpha-helices (47%) and random coils (37%). Studies to determine the localization of MANF in the brains of rat, monkey, and man, as well as the receptor, signaling pathways, and biologically active peptide mimetics are in progress. The selective, neuroprotective effect of MANF for dopaminergic neurons suggests that it may be indicated for the treatment of Parkinson's disease.

Amino Acid Sequence↗