S.A. Herrera, B.H. Justesen, T. Dieudonné, C. Montigny, P. Nissen, G. Lenoir, and T. Günther Pomorski (2023).
Direct evidence of lipid transport by the Drs2-Cdc50 flippase upon truncation of its terminal regions.
Protein Science 2023: e4855
doi: 10.1002/pro.4855
P4-ATPases in complex with Cdc50 subunits are lipid flippases that couple ATP hydrolysis with lipid transport to the cytoplasmic leaflet of membranes to create lipid asymmetry. Such vectorial transport has been shown to contribute to vesicle formation in the late secretory pathway. Some flippases are regulated by autoinhibitory regions that can be destabilized by protein kinase-mediated phosphorylation and possibly by binding of cytosolic proteins. In addition, the binding of lipids to flippases may also induce conformational changes required for the activity of these transporters. Here, we address the role of phosphatidylinositol-4-phosphate (PI4P) and the terminal autoinhibitory tails on the lipid flipping activity of the yeast lipid flippase Drs2-Cdc50. By functionally reconstituting the full-length and truncated forms of Drs2 in a 1:1 complex with the Cdc50 subunit, we provide compelling evidence that lipid flippase activity is exclusively detected for the truncated Drs2 variant and is dependent on the presence of the phosphoinositide PI4P. These findings highlight the critical role of phosphoinositides as lipid co-factors in the regulation of lipid transport by the Drs2-Cdc50 flippase.