The vasodilator-stimulated phosphoprotein (VASP) colocalizes with the ends of stress fibers in cell-matrix and cellcell contacts. We report here that bacterially expressed murine VASP directly interacts with skeletal muscle actin in several test systems including cosedimentation, viscometry and polymerization assays. It nucleates actin polymerization and tightly bundles actin filaments. The interaction with actin is salt-sensitive, indicating that the complex formation is primarily based on electrostatic interactions. Actin binding is confined to the Cterminal domain of VASP (EVH2). This domain, when expressed as a fusion protein with EGFP, associates with stress fibers in transiently transfected cells.z 1999 Federation of European Biochemical Societies.
Gene expression is controlled mainly by the binding of transcription factors to regulatory sequences. To generate a genomic map for regulatory sequences, the Arabidopsis thaliana genome was screened for putative transcription factor binding sites. Using publicly available data from the TRANSFAC database and from publications, alignment matrices for 23 transcription factors of 13 different factor families were used with the pattern search program Patser to determine the genomic positions of more than 2.4 x 10(6) putative binding sites. Due to the dense clustering of genes and the observation that regulatory sequences are not restricted to upstream regions, the prediction of binding sites was performed for the whole genome. The genomic positions and the underlying data were imported into the newly developed AthaMap database. This data can be accessed by positional information or the Arabidopsis Genome Initiative identification number. Putative binding sites are displayed in the defined region. Data on the matrices used and on the thresholds applied in these screens are given in the database. Considering the high density of sites it will be a valuable resource for generating models on gene expression regulation. The data are available at http://www.athamap.de.
The AthaMap database generates a map of cis-regulatory elements for the Arabidopsis thaliana genome. AthaMap contains more than 7.4 × 106 putative binding sites for 36 transcription factors (TFs) from 16 different TF families. A newly implemented functionality allows the display of subsets of higher conserved transcription factor binding sites (TFBSs). Furthermore, a web tool was developed that permits a user-defined search for co-localizing cis-regulatory elements. The user can specify individually the level of conservation for each TFBS and a spacer range between them. This web tool was employed for the identification of co-localizing sites of known interacting TFs and TFs containing two DNA-binding domains. More than 1.8 × 105 combinatorial elements were annotated in the AthaMap database. These elements can also be used to identify more complex co-localizing elements consisting of up to four TFBSs. The AthaMap database and the connected web tools are a valuable resource for the analysis and the prediction of gene expression regulation at .
Database‐assisted bioinformatic approaches facilitate the analysis of gene regulatory sequences and allow the identification of putative transcription factor binding sites or common regulatory motifs in similarly expressed genes. These approaches complement experiments that identify specific regulatory sites. They may be easily established if the respective transcription factors responsible for gene expression have already been cloned and annotated to the databases.
In this chapter we summarise recent results on transcriptional regulation in plants and describe the most abundant transcription factor families of
Arabidopsis thaliana
. We summarise experimental approaches to delineate gene regulatory sequences that should precede a database‐assisted analysis. The plant cis‐regulatory DNA element database PLACE and the transcription factor databases PlantCARE and TRANSFAC® are described, including connected software tools. PlanCARE is a plant specific database and TRANSFAC® covers transcription factors of all eukaryotes. The content and use of the TRANSFAC® database is presented in detail. Furthermore, we give examples on the identification of putative transcription factor binding sites in previously identified regulatory sequences and compile other databases that are relevant for the identification of regulatory sequences.
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