Computational protocol: Fascin-1 Promoter Activity Is Regulated by CREB and the Aryl Hydrocarbon Receptor in Human Carcinoma Cells

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Protocol publication

[…] Nucleotide sequences for 5.5 kb of DNA 5′ to the ATG codon of fascin-1 genes from six mammalian species were obtained from the Entrez Genomes division of NCBI (http://www.ncbi.nlm.nih.gov/sites/entrez/dbgenomeprj). The sequences were from: Homo sapiens , Build 36.2, chromosome 7; Pan troglodytes Build 2.1, chromosome 7; Macaca mulatta Build 1.1, chromosome 3; Canis lupus familiaris Build 2.1, chromosome 6; Mus musculus Build 37.1, chromosome 5 (5 86.0 cM), and Rattus norvegicus RGSC v3.4, chromosome 12.DNA sequence conservation between the species was examined by multiple sequence alignment of a 2 kb region 5′ to the ATG codon, using TCOFFEE Regular at EMBnet (http://ch.embnet.org/software/ClustalW-XXL.html). The sequences were also analysed by the algorithm FootPrinter 3.0 (http://genome.cs.mcgill.ca/cgi-bin/FootPrinter3.0/FootPrinterInput2.pl) that identifies short highly conserved regions, according to parsimony criteria in combination with the use of established phylogenetic relationships . The motif size was set to 10 and a maximum parsimony score of 2 was used. Candidate transcription factor binding sites were identified in each sequence using Match 1.0 Public (http://www.gene-regulation.com/) with the algorithm set to mimimise false negatives . Match 1.0 uses a library of mononucleotide position specific weight matrices from TRANSFAC® 6.0 to predict candidate binding sites for specific transcription factors . Pairwise analyses were also made through rVISTA 2.0 (http://rvista.dcode.org/) in which pairwise sequence alignment is combined with analysis against the TRANSFAC library of matrices to identify conserved candidate transcription factor binding sites . Because of the natural variety of TCF-binding nucleotide sequences, candidate TCF binding sites were also identified with the algorithm Target Explorer (http://luna.bioc.columbia.edu/Target_Explorer/) using a customised position specific weight matrix based on a library of motifs, compiled from a previous analysis of TCF binding motifs in combination with additional TCF motifs from the literature, with the minimum threshold set to 4. All results from the motif identification programmes were also examined against the TCOFFEE and FootPrinter sequence alignments. […]

Pipeline specifications

Software tools T-Coffee, Clustal W, FootPrinter, rVISTA, Target Explorer
Databases TRANSFAC
Application Phylogenetics
Organisms Homo sapiens
Diseases Carcinoma, Colonic Neoplasms