Mutations in the cytoplasmic domain of the integrin β1 chain indicate a role for endocytosis factors in bacterial internalization

Abstract

Mutations that result in defective β1-integrin focal adhesion formation were analyzed for effects on bacterial internalization. Mutations in the cytoplasmic domain of the β1 chain that disrupt the sequence NPIY resulted in integrins deficient in bacterial uptake. Other mutations in the β1 chain that reduced cytoskeletal association showed enhanced bacterial uptake. Replacement of the NPIY sequence of the β1 subunit by the endocytosis internalization sequence PPGY resulted in integrin receptors highly proficient in bacterial internalization, yet severely defective in focal contact localization. Electron microscopy indicated that coated structures associated specifically with bacteria-binding β1-integrins, with an apparent recruitment of coated pits from ventral cell surfaces to apical surfaces corresponding to nascent bacterial phagosomes. Clathrin inhibition studies indicated a role for the adaptor molecule AP2 as well as clathrin in integrin-mediated bacterial internalization. These results indicate that association of β1-integrins with the cytoskeleton at focal contacts interferes with integrin-mediated bacterial internalization. Also, although actin polymerization is required for bacterial uptake, clathrin is probably involved in bacterial uptake promoted by β1-integrins.