Bioinformatic, cell localization, and phylogenetic analyses reveal a novel family of putative lipases in Tetrahymena thermophila.

Lipid droplets are important and dynamic structures in all cells. Proteomic studies have shown that lipid droplets are decorated with up to hundreds of different proteins that modulate their formation, function, and interaction with other organelles. Work in our lab has shown that, like other organisms, the freshwater ciliate Tetrahymena thermophila increases lipid droplet formation during nutrient challenges such as starvation. Results of a T. thermophila proteomic screen identified lipid droplet-associated proteins, one of which (TTHERM_00784270, or 4270), is a hypothetical protein of no known function. BLAST searches of 4270 identified 15 other T. thermophila proteins and 19 proteins representing 6 other Tetrahymena species, but no orthologs outside of the genus. All of the T. thermophila genes are sequentially arranged along macronuclear chromosome 55, suggesting a possible gene duplication history. Phylogenetic analysis of all 35 protein sequences highlights the presence of both orthologs and paralogs in Tetrahymena species. Fluorescent tagging of 4 members of this family revealed that they colocalize with populations of lipid droplets. Molecular modeling as well as the presence of a pentapeptide GXSXG motif characteristic of lipases corroborate the bioinformatic evidence that we have identified a novel family of putative lipases that may play a role in how Tetrahymena cells store and use energy.