PKC-? pseudosubstrate and catalytic activity are necessary for membrane delivery during IgG-mediated phagocytosis.

ABSTRACT: In RAW 264.7 cells, PKC-? regulates Fc?R-mediated phagocytosis. BMDM behave similarly; PKC-? concentrates at phagosomes and internalization are reduced in PKC-??/? cells. Two questions were asked: what is the role of PKC-?? and what domains are necessary for PKC-? concentration? Function was studied using BMDM and frustrated phagocytosis. On IgG surfaces, PKC-??/? macrophages spread less than WT. Patch-clamping revealed that the spreading defect is a result of the failure of PKC-??/? macrophages to add membrane. The defect is specific for Fc?R ligation and can be reversed by expression of full-length (but not the isolated RD) PKC-? in PKC-??/? BMDM. Thus, PKC-? function in phagocytosis requires translocation to phagosomes and the catalytic domain. The expression of chimeric PKC molecules in RAW cells identified the ?PS as necessary for PKC-? targeting. When placed into (nonlocalizing) PKC-?, ?PS was sufficient for concentration, albeit to a lesser degree than intact PKC-?. In contrast, translocation of ?(?PSC1B) resembled that of WT PKC-?. Thus, ?PS and ?C1B cooperate for optimal phagosome targeting. Finally, cells expressing ?K437W were significantly less phagocytic than their PKC-?-expressing counterparts, blocked at the pseudopod-extension phase. In summary, we have shown that ?PS and ?C1B are necessary and sufficient for targeting PKC-? to phagosomes, where its catalytic activity is required for membrane delivery and pseudopod extension.

SUBMITTER: Wood TR 

PROVIDER: S-EPMC3685018 | biostudies-literature | 2013 Jul

REPOSITORIES: biostudies-literature