Applicability of PhyloScan
The test cases described here reflect our past and present research interests in proteobacterial gene regulation, while simultaneously emphasizing PhyloScan's ability to handle multiple weak binding sites as well as mixed aligned and unaligned sequence data. However, the features of our data set are not unique; there are many examples where multiple binding sites are common (e.g., flies [33] and humans [34]) or where transcription factors and their cognate binding sites are conserved across diverse species for which multiple sequence alignments are not feasible (e.g., between eubacteria and archaea [13-15]). PhyloScan will have clear advantages in such contexts. However, it is important to note that in situations where orthologous regions are usually alignable and for which the multiple-weak-sites scenario is unlikely, PhyloScan will not perform better than existing approaches such as MONKEY. In another direction, in cases where sequences cannot be aligned, PhyloScan will not perform better than existing approaches that handle "independent species."
Here we have demonstrated significant improvement of scan results through the use of sequences from evolutionary distant species that have orthologous transcription factors. This is not unexpected, given results of a more theoretical nature that quantify the extent of such improvement [35].