Dissect Sequence

Next, we will focus on Ume6p to demonstrate the other functions of the Yeast Transfactome Database. From the analysis of their sporulation data, Friedlander et al. [2] provided a list of genes that are expressed in early sporulation. As an example, Msh5p is a meiosis-specific protein involved in recombination. Since Ume6p regulates early sporulation genes, we hypothesize that Ume6p regulates the expression of Msh5p. Imagine that we wish to experimentally delete the binding site for Ume6p from the promoter of msh5 so that we can eliminate regulation of Msh5p by Ume6p without otherwise disrupting sporulation, as would be the case if we deleted ume6. While we could replace the entire promoter of msh5, it would be more elegant and conclusive if we could only delete the specific Ume6p site. For this task, the "Dissect Sequence" tab is useful. As a convenience, the Yeast Transfactome website has remembered our previous selection of PSAMs using a browser cookie. However, we are only interested in Ume6p now, so we click on the "Reset Previous Selections" link. After the page refreshes, we enter "msh5" in the "gene name" text field. The database is preloaded with the 800bp upstream of each yeast open reading frame (ORF; non-overlapping with other ORFs) as approximate promoter sequences. If we wanted to score a different sequence, we could enter the nucleotide sequence ourselves in the "your own sequence" field. Next, we scroll down and select UME6_H2O2Hi from the list of PSAMs. Finally, we scroll back to the top of the page and click on the "Show Matches" button. In return, we are presented with a display of the upstream sequence of msh5.

Any sequence window that has an affinity that would cause the site to be bound at least 5% as much as the best binding site in the genome is displayed. The stronger the binding site, the darker the match marker box. In this display, we see that there is a strong match for the Ume6p binding site at position 178 (TCGGCGGTTAAT). Thus, we would want to delete or scramble this sequence in order to test Ume6p regulation of Msh5p expression.