Hyperactive Rac converts sublethal nibbling to lethal phagocytosis in vivo

Abstract

The small GTPase Rac is an essential regulator of cell shape, migration, macropinocytosis and phagocytosis. We recently reported that expression of constitutively active RacG12V is sufficient to cause a few migratory cells called border cells to cannibalize nurse cells in the Drosophila ovary. Building on that insight, we engineered mammalian Rac-enhanced chimeric-antigen-receptor macrophages (RaceCAR-Ms) to avidly engulf and kill cancer cells. Here we investigate the cellular and molecular mechanisms by which border cells efficiently kill the much larger nurse cells. Surprisingly, wild type border cells normally nibble on nurse cells as they migrate between them, and RacG12V causes border cells to take larger, lethal bites. These larger bites trigger rapid germline shrinkage, nuclear damage, and caspase activation, which spreads through the nurse cell syncytium. Then, many somatic follicle cells join in to engulf the dying germline. Rac and the engulfment receptor Draper are critical for both sublethal nibbling and lethal phagocytosis. Using clonal analysis, we show small groups of follicle cells expressing RacG12V induced caspase activation in neighboring follicle cells while larger RacG12V clones were required to cause germline killing. Increasing Draper expression or JNK activity in border cells also caused germline death, in a Rac-independent manner, suggesting that border cells can be activated to kill through multiple mechanisms. The series of events elucidated here reveals how hyperactivated Rac expressed in a few cells can trigger destruction of a much larger mass of cells. Significance StatementRac is a key protein in the cellular eating process called phagocytosis. Rac hyperactivity enhances the consumption of tumor cells by chimeric antigen receptor-macrophages (CAR-M), a promising type of cellular immunotherapy. Elucidating the mechanisms by which hyperactive Rac enhances cell killing may lead to improvements in CAR-M. Key insights into the in vivo effects of Rac have come from studying Drosophila oogenesis. Here we report molecular and cellular mechanisms by which hyperactivated Rac stimulates migratory cells to engulf and kill much larger cells in the fly ovary, ultimately resulting in destruction of the entire tissue. These insights have implications for how hyperactivating Rac might improve antibody and CAR-M therapies for cancer and other diseases.