Biomedical subjects

Boris Lenhard

Publications and source records attributed to Boris Lenhard.

At least 19 recordsLinked to original sources

PubMed2006

RNAdb 2.0--an expanded database of mammalian non-coding RNAs.

RNAdb is a comprehensive database of mammalian non-protein-coding RNAs (ncRNAs). There is increasing recognition that ncRNAs play important regulatory roles in multicellular organisms, and there is an expanding rate of discovery of novel ncRNAs as well as an increasing allocation of function. In this update to RNAdb, we provide nucleotide sequences and annotations for tens of thousands of non-housekeeping ncRNAs, including a wide range of mammalian microRNAs, small nucleolar RNAs and larger mRNA-like ncRNAs. Some of these have documented functions and/or expression patterns, but the majority remain of unclear significance, and include PIWI-interacting RNAs, ncRNAs identified from the latest rounds of large-scale cDNA sequencing projects, putative antisense transcripts, as well as ncRNAs predicted on the basis of structural features and alignments. Improvements to the database comprise not only new and updated ncRNA datasets, but also provision of microarray-based expression data and closer interface with more specialized ncRNA resources such as miRBase and snoRNA-LBME-db. To access RNAdb, visit http://research.imb.uq.edu.au/RNAdb.

Animals↗

PubMed2006

In vivo transcript profiling and phylogenetic analysis identifies suppressor of cytokine signaling 2 as a direct signal transducer and activator of transcription 5b target in liver.

The GH-activated signal transducer and activator of transcription 5b (STAT5b) is an essential regulator of somatic growth. The transcriptional response to STAT5b in liver is poorly understood. We have combined microarray-based expression profiling and phylogenetic analysis of gene regulatory regions to study the interplay between STAT5b and GH in the regulation of hepatic gene expression. The acute transcriptional response to GH in vivo after a single pulse of GH was studied in the liver of hypophysectomized rats in the presence of either constitutively active or a dominant-negative STAT5b delivered by adenoviral gene transfer. Genes showing differential expression in these two situations were analyzed for the presence of STAT5b binding sites in promoter and intronic regions that are phylogenetically conserved between rats and humans. Using this approach, we showed that most rapid transcriptional effects of GH in the liver are not results of direct actions of STAT5b. In addition, we identified novel STAT5b cis regulatory elements in genes such as Frizzled-4, epithelial membrane protein-1, and the suppressor of cytokine signaling 2 (SOCS2). Detailed analysis of SOCS2 promoter demonstrated its direct transcriptional regulation by STAT5b upon GH stimulation. A novel response element was identified within the first intron of the human SOCS2 gene composed of an E-box followed by tandem STAT5b binding sites, both of which are required for full GH responsiveness. In summary, we demonstrate the power of combining transcript profiling with phylogenetic sequence analysis to define novel regulatory paradigms.

Animals↗

PubMed2006

Transcriptional and structural impact of TATA-initiation site spacing in mammalian core promoters.

BACKGROUND: The TATA box, one of the most well studied core promoter elements, is associated with induced, context-specific expression. The lack of precise transcription start site (TSS) locations linked with expression information has impeded genome-wide characterization of the interaction between TATA and the pre-initiation complex. RESULTS: Using a comprehensive set of 5.66 x 10(6) sequenced 5' cDNA ends from diverse tissues mapped to the mouse genome, we found that the TATA-TSS distance is correlated with the tissue specificity of the downstream transcript. To achieve tissue-specific regulation, the TATA box position relative to the TSS is constrained to a narrow window (-32 to -29), where positions -31 and -30 are the optimal positions for achieving high tissue specificity. Slightly larger spacings can be accommodated only when there is no optimally spaced initiation signal; in contrast, the TATA box like motifs found downstream of position -28 are generally nonfunctional. The strength of the TATA binding protein-DNA interaction plays a subordinate role to spacing in terms of tissue specificity. Furthermore, promoters with different TATA-TSS spacings have distinct features in terms of consensus sequence around the initiation site and distribution of alternative TSSs. Unexpectedly, promoters that have two dominant, consecutive TSSs are TATA depleted and have a novel GGG initiation site consensus. CONCLUSION: In this report we present the most comprehensive characterization of TATA-TSS spacing and functionality to date. The coupling of spacing to tissue specificity at the transcriptome level provides important clues as to the function of core promoters and the choice of TSS by the pre-initiation complex.

Animals↗

PubMed2006

A global genomic transcriptional code associated with CNS-expressed genes.

Highly conserved non-coding DNA regions (HCNR) occur frequently in vertebrate genomes, but their functional roles remain unclear. Here, we provide evidence that a large portion of HCNRs are enriched for binding sites for Sox, POU and Homeodomain transcription factors, and such HCNRs can act as cis-regulatory regions active in neural stem cells. Strikingly, these HCNRs are linked to several hundreds of genes expressed in the developing CNS and they may exert locus-wide regulatory effects on multiple genes flanking their genomic location. Moreover, these data imply a unifying transcriptional logic for a large set of CNS-expressed genes in which Sox and POU proteins act as generic promoters of transcription while Homeodomain proteins control the spatial expression of genes through active repression.

Animals↗

PubMed2006

Genome-wide analysis of mammalian promoter architecture and evolution.

Mammalian promoters can be separated into two classes, conserved TATA box-enriched promoters, which initiate at a well-defined site, and more plastic, broad and evolvable CpG-rich promoters. We have sequenced tags corresponding to several hundred thousand transcription start sites (TSSs) in the mouse and human genomes, allowing precise analysis of the sequence architecture and evolution of distinct promoter classes. Different tissues and families of genes differentially use distinct types of promoters. Our tagging methods allow quantitative analysis of promoter usage in different tissues and show that differentially regulated alternative TSSs are a common feature in protein-coding genes and commonly generate alternative N termini. Among the TSSs, we identified new start sites associated with the majority of exons and with 3' UTRs. These data permit genome-scale identification of tissue-specific promoters and analysis of the cis-acting elements associated with them.

3' Untranslated Regions↗

PubMed2006

Complex Loci in human and mouse genomes.

Mammalian genomes harbor a larger than expected number of complex loci, in which multiple genes are coupled by shared transcribed regions in antisense orientation and/or by bidirectional core promoters. To determine the incidence, functional significance, and evolutionary context of mammalian complex loci, we identified and characterized 5,248 cis-antisense pairs, 1,638 bidirectional promoters, and 1,153 chains of multiple cis-antisense and/or bidirectionally promoted pairs from 36,606 mouse transcriptional units (TUs), along with 6,141 cis-antisense pairs, 2,113 bidirectional promoters, and 1,480 chains from 42,887 human TUs. In both human and mouse, 25% of TUs resided in cis-antisense pairs, only 17% of which were conserved between the two organisms, indicating frequent species specificity of antisense gene arrangements. A sampling approach indicated that over 40% of all TUs might actually be in cis-antisense pairs, and that only a minority of these arrangements are likely to be conserved between human and mouse. Bidirectional promoters were characterized by variable transcriptional start sites and an identifiable midpoint at which overall sequence composition changed strand and the direction of transcriptional initiation switched. In microarray data covering a wide range of mouse tissues, genes in cis-antisense and bidirectionally promoted arrangement showed a higher probability of being coordinately expressed than random pairs of genes. In a case study on homeotic loci, we observed extensive transcription of nonconserved sequences on the noncoding strand, implying that the presence rather than the sequence of these transcripts is of functional importance. Complex loci are ubiquitous, host numerous nonconserved gene structures and lineage-specific exonification events, and may have a cis-regulatory impact on the member genes.

Animals↗

PubMed2006

Alternative promoter usage of the membrane glycoprotein CD36.

BACKGROUND: CD36 is a membrane glycoprotein involved in a variety of cellular processes such as lipid transport, immune regulation, hemostasis, adhesion, angiogenesis and atherosclerosis. It is expressed in many tissues and cell types, with a tissue specific expression pattern that is a result of a complex regulation for which the molecular mechanisms are not yet fully understood. There are several alternative mRNA isoforms described for the gene. We have investigated the expression patterns of five alternative first exons of the CD36 gene in several human tissues and cell types, to better understand the molecular details behind its regulation. RESULTS: We have identified one novel alternative first exon of the CD36 gene, and confirmed the expression of four previously known alternative first exons of the gene. The alternative transcripts are all expressed in more than one human tissue and their expression patterns vary highly in skeletal muscle, heart, liver, adipose tissue, placenta, spinal cord, cerebrum and monocytes. All alternative first exons are upregulated in THP-1 macrophages in response to oxidized low density lipoproteins. The alternative promoters lack TATA-boxes and CpG islands. The upstream region of exon 1b contains several features common for house keeping gene and monocyte specific gene promoters. CONCLUSION: Tissue-specific expression patterns of the alternative first exons of CD36 suggest that the alternative first exons of the gene are regulated individually and tissue specifically. At the same time, the fact that all first exons are upregulated in THP-1 macrophages in response to oxidized low density lipoproteins may suggest that the alternative first exons are coregulated in this cell type and environmental condition. The molecular mechanisms regulating CD36 thus appear to be unusually complex, which might reflect the multifunctional role of the gene in different tissues and cellular conditions.

Adipose Tissue↗

PubMed2006

A new generation of JASPAR, the open-access repository for transcription factor binding site profiles.

JASPAR is the most complete open-access collection of transcription factor binding site (TFBS) matrices. In this new release, JASPAR grows into a meta-database of collections of TFBS models derived by diverse approaches. We present JASPAR CORE--an expanded version of the original, non-redundant collection of annotated, high-quality matrix-based transcription factor binding profiles, JASPAR FAM--a collection of familial TFBS models and JASPAR phyloFACTS--a set of matrices computationally derived from statistically overrepresented, evolutionarily conserved regulatory region motifs from mammalian genomes. JASPAR phyloFACTS serves as a non-redundant extension to JASPAR CORE, enhancing the overall breadth of JASPAR for promoter sequence analysis. The new release of JASPAR is available at http://jaspar.genereg.net.

Animals↗

PubMed2006

New technologies, new findings, and new concepts in the study of vertebrate cis-regulatory sequences.

All vertebrates share a similar early embryonic body plan and use the same regulatory genes for their development. The availability of numerous sequenced vertebrate genomes and significant advances in bioinformatics have resulted in the finding that the genomic regions of many of these developmental regulatory genes also contain highly conserved noncoding sequence. In silico discovery of conserved noncoding regions and of transcription factor binding sites as well as the development of methods for high throughput transgenesis in Xenopus and zebrafish are dramatically increasing the speed with which regulatory elements can be discovered, characterized, and tested in the context of whole live embryos. We review here some of the recent technological developments that will likely lead to a surge in research on how vertebrate genomes encode regulation of transcriptional activity, how regulatory sequences constrain genomic architecture, and ultimately how vertebrate form has evolved.

Animals↗

PubMed2006

Transcript annotation in FANTOM3: mouse gene catalog based on physical cDNAs.

The international FANTOM consortium aims to produce a comprehensive picture of the mammalian transcriptome, based upon an extensive cDNA collection and functional annotation of full-length enriched cDNAs. The previous dataset, FANTOM2, comprised 60,770 full-length enriched cDNAs. Functional annotation revealed that this cDNA dataset contained only about half of the estimated number of mouse protein-coding genes, indicating that a number of cDNAs still remained to be collected and identified. To pursue the complete gene catalog that covers all predicted mouse genes, cloning and sequencing of full-length enriched cDNAs has been continued since FANTOM2. In FANTOM3, 42,031 newly isolated cDNAs were subjected to functional annotation, and the annotation of 4,347 FANTOM2 cDNAs was updated. To accomplish accurate functional annotation, we improved our automated annotation pipeline by introducing new coding sequence prediction programs and developed a Web-based annotation interface for simplifying the annotation procedures to reduce manual annotation errors. Automated coding sequence and function prediction was followed with manual curation and review by expert curators. A total of 102,801 full-length enriched mouse cDNAs were annotated. Out of 102,801 transcripts, 56,722 were functionally annotated as protein coding (including partial or truncated transcripts), providing to our knowledge the greatest current coverage of the mouse proteome by full-length cDNAs. The total number of distinct non-protein-coding transcripts increased to 34,030. The FANTOM3 annotation system, consisting of automated computational prediction, manual curation, and final expert curation, facilitated the comprehensive characterization of the mouse transcriptome, and could be applied to the transcriptomes of other species.

Animals↗

PubMed2005

Exploring hepatic hormone actions using a compilation of gene expression profiles.

BACKGROUND: Microarray analysis is attractive within the field of endocrine research because regulation of gene expression is a key mechanism whereby hormones exert their actions. Knowledge discovery and testing of hypothesis based on information-rich expression profiles promise to accelerate discovery of physiologically relevant hormonal mechanisms of action. However, most studies so-far concentrate on the analysis of actions of single hormones and few examples exist that attempt to use compilation of different hormone-regulated expression profiles to gain insight into how hormone act to regulate tissue physiology. This report illustrates how a meta-analysis of multiple transcript profiles obtained from a single tissue, the liver, can be used to evaluate relevant hypothesis and discover novel mechanisms of hormonal action. We have evaluated the differential effects of Growth Hormone (GH) and estrogen in the regulation of hepatic gender differentiated gene expression as well as the involvement of sterol regulatory element-binding proteins (SREBPs) in the hepatic actions of GH and thyroid hormone. RESULTS: Little similarity exists between liver transcript profiles regulated by 17-alpha-ethinylestradiol and those induced by the continuos infusion of bGH. On the other hand, strong correlations were found between both profiles and the female enriched transcript profile. Therefore, estrogens have feminizing effects in male rat liver which are different from those induced by GH. The similarity between bGH and T3 were limited to a small group of genes, most of which are involved in lipogenesis. An in silico promoter analysis of genes rapidly regulated by thyroid hormone predicted the activation of SREBPs by short-term treatment in vivo. It was further demonstrated that proteolytic processing of SREBP1 in the endoplasmic reticulum might contribute to the rapid actions of T3 on these genes. CONCLUSION: This report illustrates how a meta-analysis of multiple transcript profiles can be used to link knowledge concerning endocrine physiology to hormonally induced changes in gene expression. We conclude that both GH and estrogen are important determinants of gender-related differences in hepatic gene expression. Rapid hepatic thyroid hormone effects affect genes involved in lipogenesis possibly through the induction of SREBP1 proteolytic processing.

Animals↗

PubMed2005

Identification of functional SNPs in the 5-prime flanking sequences of human genes.

BACKGROUND: Over 4 million single nucleotide polymorphisms (SNPs) are currently reported to exist within the human genome. Only a small fraction of these SNPs alter gene function or expression, and therefore might be associated with a cell phenotype. These functional SNPs are consequently important in understanding human health. Information related to functional SNPs in candidate disease genes is critical for cost effective genetic association studies, which attempt to understand the genetics of complex diseases like diabetes, Alzheimer's, etc. Robust methods for the identification of functional SNPs are therefore crucial. We report one such experimental approach. RESULTS: Sequence conserved between mouse and human genomes, within 5 kilobases of the 5-prime end of 176 GPCR genes, were screened for SNPs. Sequences flanking these SNPs were scored for transcription factor binding sites. Allelic pairs resulting in a significant score difference were predicted to influence the binding of transcription factors (TFs). Ten such SNPs were selected for mobility shift assays (EMSA), resulting in 7 of them exhibiting a reproducible shift. The full-length promoter regions with 4 of the 7 SNPs were cloned in a Luciferase based plasmid reporter system. Two out of the 4 SNPs exhibited differential promoter activity in several human cell lines. CONCLUSIONS: We propose a method for effective selection of functional, regulatory SNPs that are located in evolutionary conserved 5-prime flanking regions (5'-FR) regions of human genes and influence the activity of the transcriptional regulatory region. Some SNPs behave differently in different cell types.

Algorithms↗

PubMed2005

RNAdb--a comprehensive mammalian noncoding RNA database.

In recent years, there have been increasing numbers of transcripts identified that do not encode proteins, many of which are developmentally regulated and appear to have regulatory functions. Here, we describe the construction of a comprehensive mammalian noncoding RNA database (RNAdb) which contains over 800 unique experimentally studied non-coding RNAs (ncRNAs), including many associated with diseases and/or developmental processes. The database is available at http://research.imb.uq.edu.au/RNAdb and is searchable by many criteria. It includes microRNAs and snoRNAs, but not infrastructural RNAs, such as rRNAs and tRNAs, which are catalogued elsewhere. The database also includes over 1100 putative antisense ncRNAs and almost 20,000 putative ncRNAs identified in high-quality murine and human cDNA libraries, with more to be added in the near future. Many of these RNAs are large, and many are spliced, some alternatively. The database will be useful as a foundation for the emerging field of RNomics and the characterization of the roles of ncRNAs in mammalian gene expression and regulation.

Animals↗

PubMed2004

Arrays of ultraconserved non-coding regions span the loci of key developmental genes in vertebrate genomes.

BACKGROUND: Evolutionarily conserved sequences within or adjoining orthologous genes often serve as critical cis-regulatory regions. Recent studies have identified long, non-coding genomic regions that are perfectly conserved between human and mouse, termed ultra-conserved regions (UCRs). Here, we focus on UCRs that cluster around genes involved in early vertebrate development; genes conserved over 450 million years of vertebrate evolution. RESULTS: Based on a high resolution detection procedure, our UCR set enables novel insights into vertebrate genome organization and regulation of developmentally important genes. We find that the genomic positions of deeply conserved UCRs are strongly associated with the locations of genes encoding key regulators of development, with particularly strong positional correlation to transcription factor-encoding genes. Of particular importance is the observation that most UCRs are clustered into arrays that span hundreds of kilobases around their presumptive target genes. Such a hallmark signature is present around several uncharacterized human genes predicted to encode developmentally important DNA-binding proteins. CONCLUSION: The genomic organization of UCRs, combined with previous findings, suggests that UCRs act as essential long-range modulators of gene expression. The exceptional sequence conservation and clustered structure suggests that UCR-mediated molecular events involve greater complexity than traditional DNA binding by transcription factors. The high-resolution UCR collection presented here provides a wealth of target sequences for future experimental studies to determine the nature of the biochemical mechanisms involved in the preservation of arrays of nearly identical non-coding sequences over the course of vertebrate evolution.

Animals↗

PubMed2004

A cladistic model of ACE sequence variation with implications for myocardial infarction, Alzheimer disease and obesity.

Sequence variation in ACE, which encodes angiotensin I converting enzyme, contributes to a large proportion of variability in plasma ACE levels, but the extent to which this impacts upon human disease is unresolved. Most efforts to associate ACE with other heritable traits have involved a single Alu insertion/deletion polymorphism, despite the probable existence of other functional sequence variants with effects that may not be consistently detectable by solely typing the Alu indel. Here, utilizing single nucleotide polymorphisms (SNPs) that differentiate major ACE clades in European populations, we demonstrate a number of significant phenotype associations across more than 4000 Swedish individuals. In a systematic analysis of metabolic phenotypes, effects were detected upon several traits, including fasting plasma glucose levels, insulin levels and measures of obesity (P-values ranging from 0.046 to 8.4 x 10(-6)). Extending cladistic models to the study of myocardial infarction and Alzheimer disease, significant associations were observed with greater effect sizes than those typically obtained in large-scale meta-analyses based on the Alu indel. Population frequencies of ACE genotypes were also found to change with age, congruent with previous data suggesting effects upon longevity. Clade models consistently outperformed those based upon single markers, reinforcing the importance of taking into consideration the possible confounding effects of allelic heterogeneity in this genomic region. Utilizing computational tools, potential functional variants are highlighted that may underlie phenotypic variability, which is discussed along with the broader implications these results may have for studies attempting to link variation in ACE to human disease.

Alzheimer Disease↗

PubMed2004

ConSite: web-based prediction of regulatory elements using cross-species comparison.

ConSite is a user-friendly, web-based tool for finding cis-regulatory elements in genomic sequences. Predictions are based on the integration of binding site prediction generated with high-quality transcription factor models and cross-species comparison filtering (phylogenetic footprinting). By incorporating evolutionary constraints, selectivity is increased by an order of magnitude as compared to single-sequence analysis. ConSite offers several unique features, including an interactive expert system for retrieving orthologous regulatory sequences. Programming modules and biological databases that form the foundation of the ConSite service are freely available to the research community. ConSite is available at http:/www.phylofoot.org/consite.

Animals↗

PubMed2004

Integrative annotation of 21,037 human genes validated by full-length cDNA clones.

The human genome sequence defines our inherent biological potential; the realization of the biology encoded therein requires knowledge of the function of each gene. Currently, our knowledge in this area is still limited. Several lines of investigation have been used to elucidate the structure and function of the genes in the human genome. Even so, gene prediction remains a difficult task, as the varieties of transcripts of a gene may vary to a great extent. We thus performed an exhaustive integrative characterization of 41,118 full-length cDNAs that capture the gene transcripts as complete functional cassettes, providing an unequivocal report of structural and functional diversity at the gene level. Our international collaboration has validated 21,037 human gene candidates by analysis of high-quality full-length cDNA clones through curation using unified criteria. This led to the identification of 5,155 new gene candidates. It also manifested the most reliable way to control the quality of the cDNA clones. We have developed a human gene database, called the H-Invitational Database (H-InvDB; http://www.h-invitational.jp/). It provides the following: integrative annotation of human genes, description of gene structures, details of novel alternative splicing isoforms, non-protein-coding RNAs, functional domains, subcellular localizations, metabolic pathways, predictions of protein three-dimensional structure, mapping of known single nucleotide polymorphisms (SNPs), identification of polymorphic microsatellite repeats within human genes, and comparative results with mouse full-length cDNAs. The H-InvDB analysis has shown that up to 4% of the human genome sequence (National Center for Biotechnology Information build 34 assembly) may contain misassembled or missing regions. We found that 6.5% of the human gene candidates (1,377 loci) did not have a good protein-coding open reading frame, of which 296 loci are strong candidates for non-protein-coding RNA genes. In addition, among 72,027 uniquely mapped SNPs and insertions/deletions localized within human genes, 13,215 nonsynonymous SNPs, 315 nonsense SNPs, and 452 indels occurred in coding regions. Together with 25 polymorphic microsatellite repeats present in coding regions, they may alter protein structure, causing phenotypic effects or resulting in disease. The H-InvDB platform represents a substantial contribution to resources needed for the exploration of human biology and pathology.

Alternative Splicing↗

PubMed2004

Variants of CYP46A1 may interact with age and APOE to influence CSF Abeta42 levels in Alzheimer's disease.

Recent studies have suggested that variants of CYP46A1, encoding cholesterol 24-hydroxylase (CYP46), confer risk for Alzheimer's disease (AD), a prospect substantiated by evidence of genetic association from several quantitative traits related to AD pathology, including cerebrospinal fluid (CSF) levels of the 42 amino-acid cleavage product of beta-amyloid (Abeta42) and the tau protein. In the present study, these claims have been explored by the genotyping of previously associated markers in CYP46A1 in three independent northern European case-control series encompassing 1323 individuals and including approximately 400 patients with measurements of CSF Abeta42 and phospho-tau protein levels. Tests of association in case-control models revealed limited evidence that CYP46A1 variants contributed to AD risk across these samples. However, models testing for potential effects upon CSF measures suggested a possible interaction of an intronic marker (rs754203) with age and APOE genotype. In stratified analyses, significant effects were evident that were restricted to elderly APOE epsilon4 carriers for both CSF Abeta42 ( P=0.0009) and phospho-tau ( P=0.046). Computational analyses indicate that the rs754203 marker probably does not impact the binding of regulatory factors, suggesting that other polymorphic sites underlie the observed associations. Our results provide an important independent replication of previous findings, supporting the existence of CYP46A1 sequence variants that contribute to variability in beta-amyloid metabolism.

Age Factors↗