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At least 1,351 records · Page 75Linked to original sources
PubMed1986

Induction of thymidine kinase activity by viruses with group B DNA genomes: bovine cytomegalovirus (bovine herpesvirus 4).

The bovine herpesvirus type 4 (BHV-4) group has a slow replication cycle, a narrow host range, and cytopathogenic effects characteristic of cytomegaloviruses (CMV), but a Group B genome structure similar to that of lymphotropic Herpesvirus saimiri (HVS). Reference BHV-4 strain DN599 and BHV-4 strains N124 and FHV-2 induced in the cytosol fraction of thymidine kinase-negative (TK-) rabbit skin (RAB-BU) cell mutants a novel TK activity. The BHV-4-induced thymidine kinase (TK) differed from the principal cytosol TK of mock-infected cells in PAGE mobility (Rm) under non-denaturing conditions and in the capacity to efficiently substitute CTP for ATP as a phosphate donor. The BHV-4 thymidine phosphorylating activity could also be distinguished from many common herpesvirus-induced TKs because it lacked iododeoxycytidine phosphorylating activity. Iododeoxyuridine, trifluorothymidine and bromovinyldeoxyuridine inhibited [3H]thymidine (0.01 mM) phosphorylation by the BHV-4 enzyme in a dose-dependent manner, but arabinosylthymine and 2'-fluoro-5-methyl-arabinosyluracil (FMAU) were poor inhibitors of [3H]thymidine phosphorylation, and acyclovir and (dihydroxy-2-propoxymethyl)guanine (DHPG) did not inhibit at all at 60 and 40 times the concentrations of [3H]thymidine, respectively.

Animals↗
PubMed2003

Activation of AP-1 signal transduction pathway by SARS coronavirus nucleocapsid protein.

In March 2003, a novel coronavirus was isolated from patients exhibiting atypical pneumonia and subsequently proven to be the causative agent of the disease now referred to as severe acute respiratory syndrome (SARS). The complete genome of the SARS coronavirus (SARS-CoV) has since been sequenced. The SARS-CoV nucleocapsid (SARS-CoV N) shares little homology with other members of the coronavirus family. To determine if the N protein is involved in the regulation of cellular signal transduction, an ELISA-based assay on transcription factors was used. We found that the amount of transcription factors binding to promoter sequences of c-Fos, ATF2, CREB-1, and FosB was increased by the expression of SARS-CoV N. Since these factors are related to AP-1 signal transduction pathway, we investigated whether the AP-1 pathway was activated by SARS-CoV N protein using the PathDetect system. The results demonstrated that the expression of N protein, not the membrane protein (M), activated AP-1 pathway. We also found that SARS-CoV N protein does not activate NF-kappaB pathway, demonstrating that activation of important cellular pathways by SAS-CoV N protein is selective. Thus our data for the first time indicate that SARS-CoV has encoded a strategy to regulate cellular signaling process.

Animals↗
PubMed2004

Rapid chromosomal assignment of medaka mutants by bulked segregant analysis.

Genetic screens in medaka are leading to the identification of an increasing number of unique mutant phenotypes. However, so far only a few genes responsible for these phenotypes have been characterized. Furthermore, no protocols using a systematic positional cloning strategy have been developed to determine the implicated genes. The PCR-based bulked segregant analysis is a fast and reliable tool to accomplish the initial steps of the positional cloning of a mutation. Here we describe the selection of a panel of genetic markers that, evenly distributed over the 24 chromosomes of medaka, provide a full coverage of the compact medaka genome (800 Mb) when used in bulked segregant analysis. The reference panel, which consists of 48 EST-derived markers, is anchored to a collection of more than 2000 polymorphic markers, thus facilitating a rapid transition from chromosomal assignment to fine mapping of the mutants. More importantly, since most of the genetic screens have been performed in the inbred Cab strain (derived from the Southern population), the selection of markers included in this panel was intended to optimize the recognition of polymorphisms between Cab and the polymorphic inbred mapping strain Kaga. Here we present a reliable mapping panel, confirmed both by the assignment of the locus responsible for the medaka mutation eyeless/Rx3 to chromosome 12, and by the analysis of its resolution power using representative markers.

Animals↗
PubMed2025

Monogenic disorders associated with motor speech phenotypes in children and adolescents undergoing clinical exome sequencing.

PURPOSE: Prior studies investigating the genetic architecture of pediatric motor speech disorders (MSDs) have been limited by small sample sizes and an exclusive focus on apraxia. We aimed to identify pathogenic genomic variants associated with MSDs in a large pediatric population referred for exome sequencing (ES). METHODS: We identified pediatric patients with MSDs who had clinical ES between 2012 and 2022. The rate of pathogenic/likely pathogenic (P/LP) findings considered causative of the MSD phenotype was determined and delineated by sex and neurodevelopmental comorbidity. Gene-based burden testing compared the rate of P/LP variants in each gene in MSD cases with a comparison clinical ES cohort. RESULTS: Positive diagnostic results were detected in 527 of 2004 (26.3%) patients with MSDs, with higher diagnostic rates in females and individuals with neurodevelopmental comorbidities. P/LP sequence variants were detected in 262 genes. Gene-based case-referent burden analysis revealed that 30 genes were nominally associated with MSDs, 2 of which (SETBP1 and ADCY5) survived exome-wide correction. CONCLUSION: Over 25% of patients with MSDs were found to harbor P/LP variants in 262 genes, many of which have not previously been associated with MSDs. Potential clinical implications include early implementation of intensive speech therapy for children diagnosed with monogenic causes of MSDs.

Humans↗
PubMed2006

Postmortem long QT syndrome genetic testing for sudden unexplained death in the young.

OBJECTIVES: This study sought to determine the spectrum and prevalence of long QT syndrome (LQTS)-associated mutations in a large cohort of autopsy-negative sudden unexplained death (SUD). BACKGROUND: Potentially heritable arrhythmia syndromes may explain a significant proportion of SUD in the young. Here, comprehensive postmortem LQTS genetic testing was performed in a cohort of SUD cases. METHODS: From September 1998 to March 2004, 49 cases of SUD (30 male patients, average age at death 14.2 +/- 10.9 years) were referred by medical examiners/coroners to Mayo Clinic's Sudden Death Genomics Laboratory. Using polymerase chain reaction, denaturing high-performance liquid chromatography, and direct DNA sequencing, open reading frame/splice site mutational analysis was conducted for all 8 genes implicated in the pathogenesis of either LQTS (LQT1 to LQT6) or multisystem disorders involving either QT or QU prolongation. RESULTS: Ten LQTS-associated mutations (4 novel) were discovered in 10 SUD cases (20%, 8 female patients, average age at death 18.0 +/- 11.8 years). The LQTS susceptibility mutations LQT1 (5), LQT2 (3), and LQT3 (2) were far more common among women (8 of 18, 44%) than men (2 of 30, 6.7%, p < 0.008). The activities at the time of SUD included sleep (5), exertion (2), auditory arousal (1), and undetermined (2). Sudden death was the sentinel event in two-thirds of the cases. CONCLUSIONS: In this cardiac channel-focused molecular autopsy investigation of SUD, over one-third of decedents harbored a putative cardiac channel mutation: 7 previously reported to host mutations in the RyR2-encoded calcium release channel and now 10 with LQTS susceptibility mutations. Accordingly, postmortem cardiac channel genetic testing should be pursued in the evaluation of autopsy-negative SUD.

Adolescent↗
PubMed2004

Rapid actions of progesterone on granulosa cells.

Ovarian granulosa cells are responsive to progesterone but do not express the nuclear progesterone receptor. In an attempt to identify a receptor for progesterone (P4) in granulosa cells (GCs), an antibody built against the ligand binding site of the P4 receptor (i.e. C-262) was used. This antibody detected a 60 kDa protein in GCs as well as spontaneously immortalized granulosa cells (SIGCs). This C-262 detectable protein localizes to the plasma membrane and binds P4. Importantly, this C-262 detectable protein appears to be involved in mediating P4's biological actions. This is based on the findings that C-262 1) blocks P4's ability to inhibit mitogen-induced mitosis and apoptosis and 2) FITC-BSA-conjugated P4 binding to granulosa cells. A C-262 detectable protein was isolated using a C-262 affinity column and sequenced. This analysis identified an unnamed protein referred to as RDA288 that was found in the rat genome (Accession number: XM216160). A nearly identical unnamed protein was found in a cDNA library of mouse lung (Accession number: AK004678). Whether RDA288 functions as a membrane receptor for P4 remains to be determined.

Animals↗
PubMed2004

Construction of representative transcript and protein sets of human, mouse, and rat as a platform for their transcriptome and proteome analysis.

The number of mammalian transcripts identified by full-length cDNA projects and genome sequencing projects is increasing remarkably. Clustering them into a strictly nonredundant and comprehensive set provides a platform for functional analysis of the transcriptome and proteome, but the quality of the clustering and predictive usefulness have previously required manual curation to identify truncated transcripts and inappropriate clustering of closely related sequences. A Representative Transcript and Protein Sets (RTPS) pipeline was previously designed to identify the nonredundant and comprehensive set of mouse transcripts based on clustering of a large mouse full-length cDNA set (FANTOM2). Here we propose an alternative method that is more robust, requires less manual curation, and is applicable to other organisms in addition to mouse. RTPSs of human, mouse, and rat have been produced by this method and used for validation. Their comprehensiveness and quality are discussed by comparison with other clustering approaches. The RTPSs are available at .

Animals↗
PubMed1998

Occurrence of MEN 2a in familial Hirschsprung's disease: a new indication for genetic testing of the RET proto-oncogene.

PURPOSE: The association of the rare hereditary cancer syndrome, multiple endocrine neoplasia type 2a (MEN 2a) with Hirschsprung's disease, both linked to germline mutations in the RET proto-oncogene, has been reported recently. With the widespread availability of genetic screening for MEN 2a, it is necessary to define the indications for genetic testing of MEN 2a and population subgroups at high risk for inheriting the disease. The purpose of this study was to assess the prevalence of Hirschsprung's disease in MEN 2a and investigate the value of genetic analysis for MEN 2a in children with familial Hirschsprung's disease. METHODS: The ethnically diverse study group consisted of unselected consecutive patients (n=426) at risk for hereditary medullary thyroid cancer (MTC) referred to a single laboratory for genetic testing. Analysis used genomic DNA and a polymerase chain reaction-based heteroduplex mutation detection strategy for exons 10, 11, 13, and 14 of the RET proto-oncogene followed by direct DNA sequencing. Significance of RET genotype-phenotype correlation was determined by Fisher's two-tailed Exact test and a 2 x 2 contingency table. RESULTS: Thirty-six distinctly new MEN 2a kindreds were identified. Hirschsprung's disease cosegregated among siblings with MEN 2a in 15 patients from 6 of the 36 (17%) families. The extent of aganglionosis in the 15 patients ranged from midrectum to duodenum. Of the 15 patients with Hirschsprung's disease, 10 (six boys, four girls) underwent thyroidectomy for MTC (n=5) or C-cell hyperplasia (n = 5) at ages 2 to 47 years (mean, 15.6 years), and the remaining five patients died in childhood of complications related to the aganglionosis. In retrospect, Hirschsprung's disease was the presenting feature of MEN 2a in five of the six families rather than MTC or pheochromocytoma. In all six MEN 2a families expressing Hirschsprung's disease, the RET mutation predisposing to the combined phenotype occurred in exon 10 at codons 609 (n=2), 618 (n=3), or 620 (n = 1). By contrast, the MEN 2a with Hirschsprung's phenotype was not found in any of the 22 families with a RETexon 11, 13, or 14 mutation (P=.0007). CONCLUSIONS: The authors conclude that Hirschsprung's disease is a phenotypic marker for MEN 2a and possibly more common than originally appreciated. The expression of Hirschsprung's disease with MEN 2a may be uniquely linked to RETexon 10 mutations. The authors recommend that (1) patients affected with MEN 2a may be counseled regarding the potential risk of Hirschsprung's disease in offspring and (2) a family history of MTC be explored in children with familial Hirschsprung's disease and genetic screening for MEN 2a be considered.

Adolescent↗
PubMedDate not supplied

HTS in the new millennium: the role of pharmacology and flexibility.

Over the past decade, high throughput screening (HTS) has become the focal point for discovery programs within the pharmaceutical industry. The role of this discipline has been and remains the rapid and efficient identification of lead chemical matter within chemical libraries for therapeutics development. Recent advances in molecular and computational biology, i.e., genomic sequencing and bioinformatics, have resulted in the announcement of publication of the first draft of the human genome. While much work remains before a complete and accurate genomic map will be available, there can be no doubt that the number of potential therapeutic intervention points will increase dramatically, thereby increasing the workload of early discovery groups. One current drug discovery paradigm integrates genomics, protein biosciences and HTS in establishing what the authors refer to as the "gene-to-screen" process. Adoption of the "gene-to-screen" paradigm results in a dramatic increase in the efficiency of the process of converting a novel gene coding for a putative enzymatic or receptor function into a robust and pharmacologically relevant high throughput screen. This article details aspects of the identification of lead chemical matter from HTS. Topics discussed include portfolio composition (molecular targets amenable to small molecule drug discovery), screening file content, assay formats and plating densities, and the impact of instrumentation on the ability of HTS to identify lead chemical matter.

Animals↗
PubMed2004

The SARS coronavirus nucleocapsid protein induces actin reorganization and apoptosis in COS-1 cells in the absence of growth factors.

In March 2003, a novel coronavirus was isolated from patients exhibiting atypical pneumonia, and was subsequently proven to be the causative agent of the disease now referred to as SARS (severe acute respiratory syndrome). The complete genome of the SARS-CoV (SARS coronavirus) has since been sequenced. The SARS-CoV nucleocapsid (SARS-CoV N) protein shares little homology with other members of the coronavirus family. In the present paper, we show that SARS-CoV N is capable of inducing apoptosis of COS-1 monkey kidney cells in the absence of growth factors by down-regulating ERK (extracellular-signal-regulated kinase), up-regulating JNK (c-Jun N-terminal kinase) and p38 MAPK (mitogen-activated protein kinase) pathways, and affecting their downstream effectors. SARS-CoV N expression also down-regulated phospho-Akt and Bcl-2 levels, and activated caspases 3 and 7. However, apoptosis was independent of the p53 and Fas signalling pathways. Furthermore, activation of the p38 MAPK pathway was found to induce actin reorganization in cells devoid of growth factors. At the cytoskeletal level, SARS-CoV N down-regulated FAK (focal adhesion kinase) activity and also down-regulated fibronectin expression. This is the first report showing the ability of the N protein of SARS-CoV to induce apoptosis and actin reorganization in mammalian cells under stressed conditions.

Actins↗
PubMed2003

Combined pituitary hormone deficiency in Australian children: clinical and genetic correlates.

OBJECTIVE: Mutations in the gene for the POU domain transcription factor POU1F1 (human Pit-1) have been reported in patients with GH, TSH and PRL deficiencies. PROP1 (Prophet of Pit-1) gene mutations also cause gonadotrophin deficiencies and in some cases partial ACTH deficiency. This study analyses the POU1F1 and PROP1 genes in a cohort of Australian children with combined pituitary hormone deficiency (CPHD) and correlates results with patient phenotype. PATIENTS AND DESIGN: Genomic analysis was carried out on 33 patients with CPHD referred from centres around Australia. Clinical data were collected from medical records and referring physicans. RESULTS: POU1F1 mutations were identified in two of four patients with a suggestive phenotype. In a female patient, novel compound heterozygous POU1F1 mutations were identified: Arg143Leu in exon 3 and Leu194Gln in exon 4. This patient presented with failure to thrive at 6 weeks of age and has deficiencies of TSH and GH. A previously described heterozygous Arg271Trp mutation in exon 6 of the POU1F1 gene was identified in a female infant who presented with growth failure and was diagnosed with TSH then GH deficiencies. No PROP1 mutations were identified; however, we describe a number of previously unreported PROP1 polymorphisms. No patients presenting with deficiencies of all anterior pituitary hormones early in life had POU1F1 or PROP1 gene mutations. CONCLUSIONS: In 33 Australian children with CPHD we have identified POU1F1 mutations in two patients and no PROP1 mutations. We speculate that in the majority of children other genes must be responsible for the CPHD phenotype.

Adolescent↗
PubMed2002

An anatomy of normal and malignant gene expression.

A gene's expression pattern provides clues to its role in normal physiology and disease. To provide quantitative expression levels on a genome-wide scale, the Cancer Genome Anatomy Project (CGAP) uses serial analysis of gene expression (SAGE). Over 5 million transcript tags from more than 100 human cell types have been assembled. To enhance the utility of this data, the CGAP SAGE project created SAGE Genie, a web site for the analysis and presentation of SAGE data (http://cgap.nci.nih.gov/SAGE). SAGE Genie provides an automatic link between gene names and SAGE transcript levels, accounting for alternative transcription and many potential errors. These informatics advances provide a rapid and intuitive view of transcript expression in the human body or brain, displayed on the SAGE Anatomic Viewer. We report here an easily accessible view of nearly any gene's expression in a wide variety of malignant and normal tissues.

DNA, Mitochondrial↗
PubMed2001

CCAAT/enhancer-binding protein-beta is a mediator of the nutrient-sensing response pathway that activates the human asparagine synthetase gene.

Transcription from the human asparagine synthetase (AS) gene is increased in response to either amino acid (amino acid response) or glucose (unfolded protein response) deprivation. These two independent pathways converge on the same set of genomic cis-elements within the AS promoter, which are referred to as nutrient-sensing response element (NSRE)-1 and -2, both of which are absolutely necessary for gene activation. The NSRE-1 sequence was used to identify the corresponding transcription factor by yeast one-hybrid screening. Based on those results, electrophoretic mobility shift assays for individual CCAAT/enhancer-binding protein-beta (C/EBP) family members were performed to test for supershifting of complexes by specific antibodies. The results indicated that of all the family members, C/EBPbeta bound to the NSRE-1 sequence to the greatest extent and that the absolute amount of this complex was increased when extracts from amino acid- or glucose-deprived cells were tested. Using electrophoretic mobility shift assays, mutation of the NSRE-1 sequence completely prevented formation of the C/EBPbeta-containing complexes. In contrast, mutation of the NSRE-2 sequence did not block C/EBPbeta binding. Overexpression in HepG2 hepatoma cells of the activating isoform of C/EBPbeta increased AS promoter-driven transcription, whereas the inhibitory dominant-negative isoform of C/EBPbeta blocked enhanced transcription following amino acid or glucose deprivation. Collectively, the results provide both in vitro and in vivo evidence for a role of C/EBPbeta in the transcriptional activation of the AS gene in response to nutrient deprivation.

Aspartate-Ammonia Ligase↗
PubMed2003

Effects of caffeine and chlorogenic acid on propidium iodide accessibility to DNA: consequences on genome size evaluation in coffee tree.

Estimates of genome size using flow cytometry can be biased by the presence of cytosolic compounds, leading to pseudo-intraspecific variation in genome size. Two important compounds present in coffee trees-caffeine and chlorogenic acid-modify accessibility of the dye propidium iodide to Petunia DNA, a species used as internal standard in our genome size evaluation. These compounds could be responsible for intraspecific variation in genome size since their contents vary between trees. They could also be implicated in environmental variations in genome size, such as those revealed when comparing the results of evaluations carried out on different dates on several genotypes.

Caffeine↗
PubMed2005

DNA-BAR: distinguisher selection for DNA barcoding.

DNA-BAR is a software package for selecting DNA probes (henceforth referred to as distinguishers) that can be used in genomic-based identification of microorganisms. Given the genomic sequences of the microorganisms, DNA-BAR finds a near-minimum number of distinguishers yielding a distinct hybridization pattern for each microorganism. Selected distinguishers satisfy user specified bounds on length, melting temperature and GC content, as well as redundancy and cross-hybridization constraints.

Algorithms↗
PubMed1999

MLH1 promoter methylation and gene silencing is the primary cause of microsatellite instability in sporadic endometrial cancers.

Defective DNA mismatch repair in human tumors leads to genome-wide instability of microsatellite repeats and a molecular phenotype referred to as microsatellite instability (MSI). MSI has been reported in a variety of cancers and is a consistent feature of tumors from patients with hereditary non-polyposis colorectal cancer. Approximately 20% of cancers of the uterine endometrium, the fifth most common cancer of women world-wide, exhibit MSI. Although the frequency of MSI is higher in endometrial cancers than in any other common malignancy, the genetic basis of MSI in these tumors has remained elusive. We investigated the role that methylation of the MLH1 DNA mismatch repair gene plays in the genesis of MSI in a large series of sporadic endometrial cancers. The MLH1 promoter was methylated in 41 of 53 (77%) MSI-positive cancers investigated. In MSI-negative tumors on the other hand, there was evidence for limited methylation in only one of 11 tumors studied. Immunohistochemical investigation of a subset of the tumors revealed that methylation of the MLH1 promoter in MSI-positive tumors was associated with loss of MLH1 expression. Immunohistochemistry proved that two MSI-positive tumors lacking MLH1 methylation failed to express the MSH2 mismatch repair gene. Both of these cancers came from women who had family and medical histories suggestive of inherited cancer susceptibility. These observations suggest that epigenetic changes in the MLH1 locus account for MSI in most cases of sporadic endometrial cancers and provide additional evidence that the MSH2 gene may contribute substantially to inherited forms of endometrial cancer.

Adaptor Proteins, Signal Transducing↗
PubMed2006

Ensembl 2007.

The Ensembl (http://www.ensembl.org/) project provides a comprehensive and integrated source of annotation of chordate genome sequences. Over the past year the number of genomes available from Ensembl has increased from 15 to 33, with the addition of sites for the mammalian genomes of elephant, rabbit, armadillo, tenrec, platypus, pig, cat, bush baby, common shrew, microbat and european hedgehog; the fish genomes of stickleback and medaka and the second example of the genomes of the sea squirt (Ciona savignyi) and the mosquito (Aedes aegypti). Some of the major features added during the year include the first complete gene sets for genomes with low-sequence coverage, the introduction of new strain variation data and the introduction of new orthology/paralog annotations based on gene trees.

Animals↗