Chlamydia trachomatis Species-Specific Induction of Ezrin Tyrosine Phosphorylation Functions in Pathogen Entry

Chlamydiae are obligate intracellular prokaryotes that, despite a considerable degree of shared developmental biology and genomic identity, exhibit distinct host infection tropisms and clinical expression of disease (32, 33, 39). This interesting spectrum in pathogen characteristics is frequently correlated with subtle differences in in vitro infection properties, particularly treatments that affect early chlamydial infection events of cultured mammalian cells (30). Chlamydiae have a biphasic developmental cycle whereby the infectious elementary body (EB) enters the host cell, evades lysosomal fusion, and differentiates into the replicating reticulate body (RB) within a host-derived vacuole (29). Infection of cells by EBs requires both attachment and entry steps. Initial attachment is temperature independent and heparin reversible. These mechanistic properties are shared by most chlamydial strains (48), suggesting a shared common chlamydial ligand and host receptor interaction. Work with chemically mutagenized Chinese hamster ovary (CHO) cells has shown that EB entry is phenotypically distinguishable from attachment since it is temperature dependent and irreversible, and the ligands and/or receptors appear to differ among chlamydial strains (7, 18). Numerous chlamydial ligands (40, 42, 47, 48) and several host receptors (10, 15) have been implicated in attachment, leaving the precise molecular mechanisms largely unresolved and controversial. Despite the apparent species specificity governing chlamydial entry mechanisms, all chlamydiae induce the reorganization and recruitment of actin that is necessary for the entry process (6, 8). The utilization of specific receptors by different chlamydial strains during EB entry is paralleled by observations that chlamydial entry also induces strain-specific tyrosine phosphorylation (Tyr-P) (44), suggesting a possible mechanistic relationship between these two independent observations. Previous studies have shown that entry is accompanied by the rapid Tyr-P of a number of proteins ranging in mass from 64 to 68, 97, and 140 kDa that were originally proposed to be of host cell origin (2, 16, 41). Recent reports, however, have identified an ∼150-kDa protein, termed translocated actin recruiting phosphoprotein (Tarp), as being a chlamydial type III effector that is translocated to the cytoplasmic face of the host cell plasma membrane (9). Although Tyr-P is not essential for Tarp function, Tarp plays an important role in actin recruitment at the entry site (8). We recently showed that the entry-mediated Tyr-P of a 70-kDa complex of proteins was chlamydial strain dependent where infection by human C. trachomatis strains, but not the guinea pig pathogen C. caviae, induced strong Tyr-P of this complex (44). The specific induction of these Tyr-P proteins by human strains suggested to us that they might be critical in the pathogenesis of human infection. The present study examined the 70-kDa protein complex and identified the host protein ezrin, a member of the ezrin-radixin-moesin (ERM) family that serves as a physical link between the host actin cytoskeleton and the plasma membrane. Here we show that ezrin is recruited to microvillus-like structures at sites of chlamydial entry. Importantly, ezrin is Tyr-P only upon C. trachomatis infection and plays an essential role in pathogen entry. This is the first confirmed identification of a host protein that is Tyr-P during chlamydial infection that is an integral component in the pathogenesis of C. trachomatis.