ABSTRACT Every day ~50 billion cells undergo apoptosis, or programmed cell death. Efficient recognition and clearance of these cells is critical for tissue homeostasis in physiology and for its recovery following disease. Initiation of the apoptotic cascade triggers activation of phospholipid scramblases that externalize the lipid phosphatidylserine (PS) in the outer leaflet of the plasma membrane. Recognition of PS by dedicated receptors (PSRs) on immune cells is the first and critical step in the clearance of apoptotic cells. Immune cells with activated PSRs create an immunosuppressive environment that can be exploited by pathogens exposing PS in an immune-camouflage strategy called apoptotic mimicry. Recent studies implicated members of the XKR protein family in apoptotic scrambling. Dysfunction of XKR proteins results in an inflammatory environment and autoimmune disorders while their uncontrolled activation favors oncogenesis and facilitate viral entry. Thus, it is critical to elucidate the molecular mechanisms of XKR function and regulation to understand their physiological roles and their association with pathology. Currently, only structures of non-functional XKR proteins are available, hindering our understanding of how these protein mediate PS externalization. In preliminary experiments we show that two XKR homologues, CED-8 from C. elegans and human XKR4, scramble lipids. Using cryogenic electron microscopy, we determined the 3.55 Å resolution structure of hXKR4 in a novel, likely active conformation, providing insights into a potential mechanism for lipid transport. In our 1st aim, we propose to determine the structures of hXKR4 and CED-8 in different conformations and functional states to determine the molecular bases of XKR activation. We will use our newly developed biochemical assays to probe and elucidate the functional implications of these structures. The physiological implications of these mechanisms will be tested in cell-based measurements. Our