Electrical signals generated by the flow of ions across cellular membranes serve many physiological purposes. For example, neurons convey information in the form of action potentials, skeletal muscle contracts upon receiving electrical inputs, and hormones are released in response to depolarization. The currents that underlie these signals flow through ion channels. Appropriate abundance, localization, and activity of ion channels are crucial to ensure coordinated signaling and normal cellular function. Structure-function studies have revealed many sequence motifs in ion channels that are important for their biosynthesis, trafficking, and function. Less is known about the cellular machinery that interacts with ion channels during their biosynthesis and trafficking, and that regulates channel function at the plasma membrane. For my thesis project, I designed and implemented a screen to identify basic cellular machinery necessary for the functional expression of an inwardly rectifying K+ (Kir) channel.
The screen took advantage of previous knowledge gained from structure-function studies, and of the powerful genetic tools available in the yeast Saccharomyces cerevisiae. By assaying 374 yeast strains, each lacking one gene, for functional expression of the Kir channel, we identified seven yeast deletion strains (sur4?, csg2?, erv14?, emp24?, erv25?, bst1?, and yil039w/ted1?) in which functional expression of the Kir channel was disrupted. The seven strains lack proteins with conserved cellular functions in quality control in, and vesicle budding from the endoplasmic reticulum (ER), and in lipid biosynthesis.
The identification of these seven proteins as players in Kir channel biosynthesis, trafficking, and/or function, opens new avenues of research. Future studies will address the role of the mammalian homologs of the seven yeast genes in Kir channel ontogenesis. In addition to pointing toward specific genes, the results of our screen highlight the importance of protein-lipid interactions in trafficking and function of Kir channels. Last but not least, we propose that the previously uncharacterized protein Yil039wp, which we named Ted1p (Trafficking of Emp24p/Erv25p dependent cargo Disrupted), regulates the function of p24 proteins, possibly through dephosphorylation of a yet unknown target.