Role of amphipathic helix of a herpesviral protein in membrane deformation and T cell receptor downregulation.

Lipid rafts are membrane microdomains that function as platforms for signal transduction and membrane trafficking. Tyrosine kinase interacting protein (Tip) of T lymphotropic Herpesvirus saimiri (HVS) is targeted to lipid rafts in T cells and downregulates TCR and CD4 surface expression. Here, we report that the membrane-proximal amphipathic helix preceding Tip's transmembrane (TM) domain mediates lipid raft localization and membrane deformation. In turn, this motif directs Tip's lysosomal trafficking and selective TCR downregulation. The amphipathic helix binds to the negatively charged lipids and induces liposome tubulation, the TM domain mediates oligomerization, and cooperation of the membrane-proximal helix with the TM domain is sufficient for localization to lipid rafts and lysosomal compartments, especially the mutivesicular bodies. These findings suggest that the membrane-proximal amphipathic helix and TM domain provide HVS Tip with the unique ability to deform the cellular membranes in lipid rafts and to downregulate TCRs potentially through MVB formation.

Fibronectin facilitates the invasion of Orientia tsutsugamushi into host cells through interaction with a 56-kDa type-specific antigen.

BACKGROUND: Orientia tsutsugamushi, the causative agent of scrub typhus, is an obligate intracellular bacterium. The pathogen's mechanism of cellular invasion is poorly characterized. METHODS: Through ligand immunoblots, glutathione S-transferase (GST) pull-down assays, and in vitro inhibition assays of intracellular invasion, a bacterial ligand was identified and was shown to interact with fibronectin (Fn) to enhance the intracellular invasion of O. tsutsugamushi. RESULTS: O. tsutsugamushi can bind to immobilized Fn in vitro, and exogenous Fn stimulates bacterial invasion of mammalian host cells. Bacterial invasion in the presence of Fn was abrogated by the addition of Arg-Gly-Asp peptides or by an anti-alpha5beta1 integrin antibody. Through a ligand immunoblot and GST pull-down assay, a 56-kDa type-specific antigen (TSA56) was identified as the bacterial ligand responsible for the interaction with Fn. Antigenic domain III and the adjacent C-terminal region (aa 243-349) of TSA56 interacted with Fn. Furthermore, we found that the enhanced invasion of the pathogen was abrogated by the addition of purified recombinant peptides derived from TSA56. CONCLUSION: Fn facilitates the invasion of O. tsutsugamushi through its interaction with TSA56.