ABSTRACT ROLES OF TNFAIP8 IN CELL MIGRATION AND PHOSPHOINOSITIDE SIGNALING Mei Lin Youhai H. Chen, M.D., Ph.D. Well-coordinated directional cell migration is essential for diverse physiological processes such as embryogenesis and immune responses, while its dysfunction can lead to many pathological conditions including cancer and chronic inflammatory diseases. How shallow gradients of chemoattractants translate into the axes of cellular polarization and direct migration remain not well understood. In this study, we have employed loss- and gain-of-function approaches and found that TNFAIP8-deficient human myeloid HL-60 cells exhibit reduced growth and migratory capacity. TNFAIP8 can serve as a cytosolic inhibitor of Rac/Cdc42 and function through the PAK-LIMK-cofilin pathway, thus controlling trailing edge formation. Biochemical studies and cell-based approaches revealed that TNFAIP8 can interact with PtdIns(4,5)P2 and PtdIns(3,4,5)P3 through the conserved TIPE homology (TH) domain. TNFAIP8 can steer cells by enhancing PI3Ks activity and PtdIns(4,5)P2-dependent actin remodeling through cofilin, reinforcing a positive feedback loop that amplifies phosphoinositide signaling at the leading edge. Consequently, TNFAIP8 is required for chemoattractant-induced polarization by acting as a “dual-role regulator” in cytoskeletal dynamics in response to external stimuli. Additionally, using experimental autoimmune encephalomyelitis (EAE) as a mouse model for multiple sclerosis (MS), we found TNFAIP8 and TIPE2 play redundant roles in controlling lymphocyte migration and infiltration. Our results support that TNFAIP8 is important in inflammation and tumorigenesis, and represents a potential pharmaceutical drug target to block cell migration for treating human diseases such as cancer.