NMAAP1 Maintains Nil Phenotype in Macrophages Through Binding to IP3R and Activating Calcium-related Signaling Pathways

Background: NMAAP1 plays a role in regulating macrophage differentiation to the M1 type and exerting antitumoral functions. It is not clear what role and mechanism NMAAP1 does play in the reversal of macrophages from M1 to M2. Methods: We detected the typing of macrophages with high or low expression of NMAAP1 by QPCR and ELISA, and detected the colocalization of NMAAP1 and endogenous IP3R by laser confocal microscopy, and detected the protein expression in cells by Western-blotting. Results: Our study found that knockdown NMAAP1 in RAW264.7 cells induced macrophage polarization to the M2 type and up-regulation of NMAAP1 in RAW264.7 cells maintain M1 Phenotype even in the presence of IL-4, a stronger inducer of the M2 type. Additionally, Co-immunoprecipitation revealed a protein-protein interaction between NMAAP1 and IP3R and then activates key molecules in the PKC-dependent Raf/MEK/ERK and Ca2+/CaM/CaMKII signaling pathways. Activation of PKC (Thr638/641), ERK1/2 (Thr202/Tyr204) and CaMKII (Thr286) is involved in the regulation of cell differentiation. Conclusion: NMAAP1 interacts with IP3R, which in turn activates the PKC-dependent Raf/MEK/ERK and Ca2+/CaM/CaMKII signaling pathways. These results provide a new explanation of the mechanism underlying M1 differentiation.