Gene expression and its regulation is a highly coordinated system, involved in many biological processes such as cell growth, division and differentiation. Transcriptional regions, involved in gene regulation, consist of a heterogeneous collection of smaller regulatory elements. In some cases, co-regulated genes contain a common set of transcription factor binding sites (TFBS).
Analysis of promoter regions is the major approach in understanding the transcriptional regulatory mechanisms. It is also useful for interpretation of mammalian gene expression studies, where co-expressed genes may share motifs representing putative TFBS. Motif identification also has the advantage that it can predict control regions in genes that have not been measured experimentally. However, a common problem is incomplete genomic sequence for the experimental species of interest. The approach here is to identify and use orthologous gene promoter sequences from a related and well-characterised species.
The primary aim of this study was to identify and predict regulatory TFBS in species where promoter sequence does not exist or is incomplete. The MEME programme was employed for the motif prediction step. The predicted elements were subsequently compared to known TFBS using TRANSFAC and JASPAR databases for identification. A methodology based on relative entropy was used. The validity of the method was confirmed as the predicted motifs in the training set were the expected sites involved in regulation of muscle development. The technique was applied to two data sets, generated from expressed sequence tag (EST) clustering analysis and microarray experiments. All data sets, software and results are available on the accompanying CD.
Bovine expression data was analysed for cardiac-specific expression using two separate approaches, combining bovine library EST frequency and human gene expression ratios. For each approach, the orthologous human and bovine promoter sequences were analysed for common motifs. Across all comparisons, 37% of motifs were identified as known TFBS using the TRANSFAC and JASPAR databases. As the human comparison had more promoter sequences available, this was the main limiting factor for the corresponding bovine analysis, rather than cross-species divergence or accuracy of gene expression measurement. Results from this study demonstrate that using promoter sequences from a related species is a viable approach when studying gene expression in species with limited amount of genomic sequence. As the bovine genome becomes more complete, it can in turn serve as the reference genome for other agriculturally important ruminants, such as sheep, goat and deer.
The second application concerned in silico analysis of gene regulation patterns in response to stimuli. Recently it has been shown that a mutation in the bone morphogenetic receptor IB leads to an increased ovulation rate in sheep. The objective of this study was to analyse gene expression patterns in cultured cells in response to four members of the BMP family, i.e. BMP2, BMP4, BMP6 and BMP7 and the control TGFβ. Microarray data was provided by J. Young. Twelve highly upregulated genes were stimulated by all BMPs, seven of which are known BMP target genes. Analysis of the predicted motifs identified four elements that may be involved in the regulation process. Cross-species comparison for one of the genes, ID1, showed high conservation of one of the motifs across 11 mammalian genomes. This particular motif had not been identified as a known binding site. In summary, the analysis of the expression data suggest an extension of the list of BMP targets.
The proposed method is relatively robust when sufficiently co-expressed (co-regulated) sequences can be identified, whether from the same or another species.
| Identifer | oai:union.ndltd.org:ADTP/217729 |
| Date | January 2007 |
| Creators | Zadissa, Amonida, n/a |
| Publisher | University of Otago. Department of Biochemistry |
| Source Sets | Australiasian Digital Theses Program |
| Language | English |
| Detected Language | English |
| Rights | http://policy01.otago.ac.nz/policies/FMPro?-db=policies.fm&-format=viewpolicy.html&-lay=viewpolicy&-sortfield=Title&Type=Academic&-recid=33025&-find), Copyright Amonida Zadissa |
Page generated in 0.0016 seconds