Clathrin coated pit dependent pathway for Trypanosoma cruzi internalization into host cells.

A number of intracellular pathogens are internalized by host cells via multiple endocytic pathways, including Trypanosoma cruzi, the etiological agent of Chagas disease. Clathrin-mediated endocytosis is the most characterized endocytic pathway in mammalian cells. Its machinery was described as being required in mammalian cells for the internalization of large particles, including pathogenic bacteria, fungi, and large virus. To investigate whether T. cruzi entry into host cells can also take advantage of the clathrin-coated vesicle-dependent process, we utilized well-known inhibitors of clathrin-coated vesicle formation (sucrose hypertonic medium, chlorpromazine hydrochloride and pitstop 2) and small interference RNA (siRNA). All treatments drastically reduced the internalization of infective trypomastigotes and amastigotes of T. cruzi by phagocytic (macrophages) and epithelial cells. Clathrin labeling, as observed by fluorescence and electron microscopy, was also observed around the parasites from the initial stages of infection until the complete formation of the parasitophorous vacuole. These unexpected observations suggest the participation of the clathrin pathway in the T. cruzi entry process.

Macropinocytosis: a pathway to protozoan infection.

Among the various endocytic mechanisms in mammalian cells, macropinocytosis involves internalization of large amounts of plasma membrane together with extracellular medium, leading to macropinosome formation. These structures are formed when plasma membrane ruffles are assembled after actin filament rearrangement. In dendritic cells, macropinocytosis has been reported to play a role in antigen presentation. Several intracellular pathogens are internalized by host cells via multiple endocytic pathways and macropinocytosis has been described as an important entry site for various organisms. Some bacteria, such as Legionella pneumophila, as well as various viruses, use this pathway to penetrate and subvert host cells. Some protozoa, which are larger than bacteria and virus, can also use this pathway to invade host cells. As macropinocytosis is characterized by the formation of large uncoated vacuoles and is triggered by various signaling pathways, which is similar to what occurs during the formation of the majority of parasitophorous vacuoles, it is believed that this phenomenon may be more widely used by parasites than is currently appreciated. Here we review protozoa host cell invasion via macropinocytosis.

Trans-sialidase stimulates eat me response from epithelial cells.

Epithelial cell invasion by the protozoan parasite Trypanosoma cruzi is enhanced by the presence of an enzyme expressed on its cell surface during the trypomastigote life cycle stage. The enzyme, trans-sialidase (TS), is a member of one of the largest gene families expressed by the parasite and the role of its activity in mediating epithelial cell entry has not hitherto been understood. Here we show that the T. cruzi TS generates an eat me signal which is capable of enabling epithelial cell entry. We have utilized purified, recombinant, active (TcTS) and inactive (TcTS2V0) TS coated onto beads to challenge an epithelial cell line. We find that TS activity acts upon G protein coupled receptors present at the epithelial cell synapse with the coated bead, thereby enhancing cell entry. By so doing, we provide evidence that TS proteins bind glycans, mediate the formation of distinct synaptic domains and promote macropinocytotic uptake of microparticles into a perinuclear compartment in a manner which may emulate entosis.