Involvement of ezrin/moesin in de novo actin assembly on phagosomal membranes

The current study focuses on the molecular mechanisms responsible for actin assembly on a defined membrane surface: the phagosome. Mature phagosomes w ere surrounded by filamentous actin in vivo in two different cell types. Fl uorescence microscopy was used to study in vitro actin nucleation/polymeriz ation (assembly) on the surface of phagosomes isolated from J774 mouse macr ophages, In order to prevent non-specific actin polymerization during the a ssay, fluorescent G-actin was mixed with thymosin beta 4. The cytoplasmic s ide of phagosomes induced de novo assembly and barbed end growth of actin f ilaments. This activity varied cyclically with the maturation state of phag osomes, both in vivo and in vitro. Peripheral membrane proteins are crucial components of this actin assembly machinery, and we demonstrate a role for ezrin and/or moesin in this process. We propose that this actin assembly p rocess facilitates phagosome/endosome aggregation prior to membrane fusion.