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Book Summary:
We used a combination of computational and experimental approaches to characterize the LSF (Late SV40 Factor)/GRH (Grainyhead) transcription factor family, members of which have roles in the cell cycle, growth and development. We investigated the phylogeny of the family, identified new members and predicted their biological roles. The family appears to have undergone an ancient gene duplication event and exhibits a structural and functional division into two distinct subfamilies. Bayesian prior-based protein sequence profiles were constructed to reveal distinctive as well as shared features between the subfamilies. The newly identified C. elegans member, Ce-GRH-1, groups with the GRH subfamily while the novel X. laevis member, Xl-LSF, groups with the LSF subfamily. In the process of identifying the C. elegans GRH homolog, we successfully resolved errors in annotation of the encompassing genomic sequence. Experimental characterization of the DNA binding site preferences of Ce-GRH-1 and D. melanogaster GRH demonstrated that they are identical. Further, RNA-mediated interference of ce-grh-1 resulted in embryonic lethality in C. elegans, accompanied by soft, defective cuticles. These phenotypes are strikingly similar to those observed in D. melanogaster grh mutants. Taken together, the results suggest evolutionarily conserved functional properties between these proteins. We designed a database of putative transcription start sites and promoters of all genes in C. elegans and D. melanogaster. In order to investigate GRH gene targets, we developed a method to detect statistically overabundant motifs in promoters of known or candidate GRH-regulated genes. These promoters contain potential binding sites for several transcription factors known or likely to be involved in ectodermal and developmental pathways. A set of novel GRH target genes was predicted based on the presence of clusters of these overabundant motifs in their promoters. This provides insight into the potential biological functions of GRH factors. In addition, mechanistic predictions are made regarding transcription factors that may coordinate with GRH in controlling developmental processes. |
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