Chlamydial Entry Involves TARP Binding of Guanine Nucleotide Exchange Factors

Chlamydia trachomatis attachment to cells induces the secretion of the elementary body–associated protein TARP (Translocated Actin Recruiting Protein). TARP crosses the plasma membrane where it is immediately phosphorylated at tyrosine residues by unknown host kinases. The Rac GTPase is also activated, resulting in WAVE2 and Arp2/3-dependent recruitment of actin to the sites of chlamydia attachment. We show that TARP participates directly in chlamydial invasion activating the Rac-dependent signaling cascade to recruit actin. TARP functions by binding two distinct Rac guanine nucleotide exchange factors (GEFs), Sos1 and Vav2, in a phosphotyrosine-dependent manner. The tyrosine phosphorylation profile of the sequence YEPISTENIYESI within TARP, as well as the transient activation of the phosphatidylinositol 3-kinase (PI 3 -K), appears to determine which GEF is utilized to activate Rac. The first and second tyrosine residues, when phosphorylated, are utilized by the Sos1/Abi1/Eps8 and Vav2, respectively, with the latter requiring the lipid phosphatidylinositol 3,4,5-triphosphate. Depletion of these critical signaling molecules by siRNA resulted in inhibition of chlamydial invasion to varying degrees, owing to a possible functional redundancy of the two pathways. Collectively, these data implicate TARP in signaling to the actin cytoskeleton remodeling machinery, demonstrating a mechanism by which C. trachomatis invades non-phagocytic cells. The human pathogen Chlamydia trachomatis is the causative agent of the most prevalent bacterial sexually transmitted disease in industrialized nations, and of the preventable blinding condition trachoma in developing countries. Survival and replication of chlamydial species occur exclusively inside a host cell, and thus, gaining access to the protective intracellular niche is an absolute requirement. This report describes how the chlamydia protein TARP, which is secreted at the base of the bacteria and across the host membrane, acts as a scaffold to which host signaling proteins bind. This assembly of the complex of signaling proteins, which include Sos1, Abi1, Eps8, and Vav2 results in the remodeling of the host cytoskeleton to facilitate engulfment of the infecting chlamydia. We conclude that these proteins have a role in chlamydia based on a number of observations including their interaction with the TARP protein, their ability to switch on known signaling participants in chlamydia invasion, their localization at the site of chlamydia entry, and the inhibition of chlamydia invasion in their absence. Altogether, the data functionally link TARP with signaling pathways that function in chlamydial invasion, demonstrating the direct involvement of TARP in the invasion of host cells by C. trachomatis