Recent advances in understanding the regulatory mechanisms of P4-ATPase lipid flippases.

P4-ATPases establish and maintain transbilayer lipid asymmetry in eukaryotic membranes by transporting phospholipids from the exoplasmic to the cytosolic membrane leaflet at the expense of ATP hydrolysis. In recent years, biochemical analysis together with an increasing number of cryo-electron microscopy structures has substantially increased our understanding of the lipid transport cycle and substrate recognition by P4-ATPases. However, the underlying molecular mechanism governing P4-ATPase activation and regulation remains less well understood. The present review aims to summarize current insights into the molecular mechanisms controlling P4-ATPase activation and regulation. We discuss autoinhibition by N- and C-terminal extensions, activation by regulatory lipids, particularly phosphoinositides, regulation by interacting proteins including small GTPases and guanine nucleotide exchange factors, and post-translational modifications, especially phosphorylation. Together, these studies reveal that P4-ATPase activity is controlled by a complex and multilayered regulatory network that enables precise control of lipid transport. Future studies combining structural, biochemical, and cellular approaches will be essential to elucidate how these mechanisms cooperate to maintain membrane lipid homeostasis and support diverse cellular functions.