Throughout development, cells are programmed to die for homeostatic control. Corpse clearance is key for animals to avoid inflammation and auto-immune disease. To better understand the "eat me" signal of phosphatidylserine (PS) exposure for cell clearance, we are attempting to disrupt the externalization of PS on C. elegans polar bodies. The second polar body undergoes a non-apoptotic form of cell death, externalizes PS, and is cleared by embryonic cells during early development. PS exposure is known to be caused by lipid scramblases, enzymes present in the plasma membrane that translocate phospholipids between the leaflets of the lipid bilayer. However, the scramblases that regulate PS exposure in non-apoptotic dying cells are unknown. A combination of RNAi and the
ced-8(
n1891) loss-of-function allele suggested that the scramblase CED-8 plays a redundant role in externalizing PS on polar bodies with the mitochondrial factor WAH-1 as well as the scramblases SCRM-1, SCRM-2, and SCRM-3. As CED-8, WAH-1, and SCRM-1 are involved in PS externalization during apoptosis, we tested whether PS exposure depends upon the apoptotic caspase CED-3. However, PS was still externalized on polar bodies in
ced-3(
n717) mutants, confirming that scramblase activation is independent of the apoptotic pathway. To confirm the role of the redundant scramblases, we generated a
scrm-4 scrm-1 scrm-2;
scrm-3;
ced-8 quintuple scramblase mutant strain. These mutants still externalized PS on polar bodies, indicating that there are other ways to externalize PS on these cells. We are currently testing other scramblase proteins, including SCRM-6, SCRM-7, SCRM-8, and CED-7. Thus, we will define the molecular players in the regulation of PS exposure for non-apoptotic cell death, which will allow us to define the role of PS in phagocytic uptake as well as later steps of phagosome maturation.