Comparative analysis of ciliary gene regulation in nematodes

Cilia are highly-conserved organelles ubiquitously present in metazoans and some unicellular eukaryotes. In humans, ciliary defects result in a plethora of serious genetic diseases termed ciliopathies. Despite their diverse morphology and function, cilia are comprised of a core set of proteins, and many ciliary genes share similar but likely not identical regulation mechanisms. Our research aims to understand the variations in cis-regulatory elements in ciliary genes and the impact of such variations on transcriptional regulation. We hypothesize that cis-regulatory elements in different ciliary promoters are unique and that this uniqueness impacts the expression and function of ciliary genes. We focus on a particular cis-regulatory element, the X-box motif, which functions as the binding motif for RFX/DAF-19, a transcription factor that regulates ciliary gene expression. We identify and analyze X-box motifs for a set of 32 well-studied ciliary genes in C. elegans and their orthologs in 25 additional nematodes, including both free-living and parasitic species. My research consists of three modules. First, we curate ciliary gene orthologs using a combined approach, including homology-based gene finding and RNA-seq-based improvement. The primary goal of this step is to ensure that the 5' ends of the genes are accurately defined in order to properly locate ciliary promoters. Second, we search for putative X-box motifs in these promoters using computational tools to identify motifs that resemble the consensus. For the promoters from which consensus X-box motifs are not found, we will search for X-box motifs that may show more differences from the consensus using frequency matrix-based search and regular expressions, which we call "atypical" X-box motifs. Third, we analyze the putative atypical X-box motifs, focusing on their sequence similarities, positions in promoter sequences, and flanking sequences, and compare them against the consensus X-box motifs. In this study, I will highlight progress made and challenges encountered for defining X-box motifs in ciliary genes.