Front Signal-Dependent Accumulation of RHOA Inhibitor FAM65B at Leading Edges Polarizes Neutrophils

A hallmark of neutrophil polarization is the back localization of active RHOA and phosphorylated myosin light chain (pMLC). However, the mechanism for the polarization is not entirely clear. Here we show that FAM65B, a newly identified RHOA inhibitor, is important for the polarization. When FAM65B is phosphorylated, it binds to 14-3-3 and becomes more stable. In neutrophils, chemoattractants stimulate FAM65B phosphorylation largely depending on the front signals that include those mediated by PLCβ and PI3Kγ, leading to FAM65B accumulation at the leading edge. Concordantly, FAM65B-deficiency in neutrophils resulted in an increase in RHOA activity and front localization of pMLC as well as defects in chemotaxis directionality and adhesion to endothelial cells under flow. These data together elucidate a mechanism for RHOA and pMLC polarization in stimulated neutrophils through direct inhibition of RHOA by FAM65B at the leading edge.