RABEP1 amplifies front signaling in neutrophil migration

Abstract

Neutrophils are the first responders of our innate immune system, crucial for defense against various infections. The intricate regulation of neutrophil migration is essential for neutrophil function. However, a complete mechanistic understanding is missing. We performed a miRNA overexpression screen and identified miR-190 as a potent suppressor of neutrophil migration in zebrafish. Through a second round of small-scale screening using neutrophil-specific knockouts of putative miR-190 targets, we identified that rabep1 (Rabaptin-5, RAB GTPase binding effector protein 1) is essential for neutrophil motility and chemotaxis in zebrafish. Re-expressing full-length RABEP1 in the knockout, but not a truncated form lacking the Rab4/Rab5 binding domain, rescued cell motility. Knocking down RABEP1 in human dHL-60 cells consistently reduced cell motility. RABEP1-deficient dHL-60 cells accumulate excessive members inside the cells. The Rab5 GTP level is unaffected, but the RABEP1 knockdown cells displayed reduced PAK phosphorylation and overall reduced actin polymerization, but still appropriately polarized upon chemokine stimulation. Overexpression of dominant-negative Rab4 or Rab5 similarly inhibits neutrophil migration. Our data suggests that RABEP1 drives endosomal recycling, Rac activation, and leading-edge actin polymerization, providing significant insights into the role of the endocytic pathway in neutrophil motility.