Evolution of the multivesicular body ESCRT machinery; retention across the eukaryotic lineage

Evolution of the multivesicular body ESCRT machinery; retention across the eukaryotic lineage. functions in intimate connection with the trypanosome ESCRT system. INTRODUCTION is a highly divergent protozoan parasite and the causative agent of human and animal trypanosomiasis in sub-Saharan Africa. Despite a pressing need for new drugs and essentially no prospect for a vaccine, trypanosomiasis and related pathogens remain neglected by the pharmaceutical industry. has emerged as a model organism for multiple pathogenic trypanosomes, including and is also extremely valuable for comparative cell biology, due to the phylogenetic position within the Excavata supergroup coupled Cordycepin with an advanced experimental toolbox. is presently the most experimentally accessible member of the phylum Euglenozoa. evades the mammalian host acquired immune response through a combination of mechanisms, which include antigenic variation of the superabundant variant surface glycoprotein (VSG) and antibody clearance from the cell surface (discussed in reference Cordycepin 1). Both of these processes involve trafficking of the VSG protein through the flagellar pocket (FP), the sole site of endo- and exocytosis in these organisms. Endocytosis and recycling are extremely rapid processes and heavily reliant upon clathrin (CLH)-mediated mechanisms (2). Evolutionary modifications to the molecular machinery that subtend endocytosis include loss of the AP-2 cargo adaptor complex (3) and the presence of a cohort of lineage-specific clathrin-associated proteins (CAPs) (4). These adaptations, together with a very high endocytic flux, serve to maintain VSG surface density and rapidly remove surface-bound immune effectors, including anti-VSG antibodies (5). Further, endocytosis also contributes to mechanisms for evasion of the innate immune system that eliminate many subspecies of via uptake of trypanolytic factor (TLF). Here, human or higher-primate infectivity requires modifications to endocytic and receptor-mediated internalization mechanisms (6, 7). Most recently, it has emerged that endocytosis is a major mechanism for uptake of some first-line trypanocides and that this extreme rate of endocytosis offers a potential mechanism to deliver therapeutics to the parasite in a highly efficient manner (8, 9). Understanding the endocytic apparatus of trypanosomes is of direct therapeutic importance as well as providing insight into evolution of trafficking systems and has led to detailed characterization of several Rab proteins from this organism. Rab proteins constitute the largest family of the Ras GTPase superfamily and are important spatiotemporal regulators of membrane transport (10). Several endosomal compartments in are regulated by Rab proteins and have been described by us and others. Rab5A (TbRab5A)- and TbRab5B-containing endosomes receive distinct cargo in bloodstream-form (BSF) cells and, similarly to mammalian cells, are early compartments of the endocytic system (11, 12, 13). TbRab4 and TbRab11 are involved in recycling (11, 14, 15, 16, 17), and TbRab28 was recently shown to coordinate retromer-dependent trafficking and late endosomal pathways (18). TbRab7 mediates delivery from late endosomal compartments to the lysosome (19). A Rab21 orthologue is present in trypanosomes. Significantly, Cordycepin Rab5, Rab20, Rab21, Rab22, Rab24, and Rab50 likely form an ancient endocytic clade that is predicted to be present in the last eukaryotic common ancestor (LECA), and thus TbRab5 and TbRab21 are the only representatives of this clade conserved in trypanosomes (20, 21). In mammalian cells, Rab21 localizes to early endosomal vesicles and has extensive colocalization with early endosomal Rab4, Rab5, Rab17, and Rab22 but less with Rabs located at late and recycling endosomes. Cells expressing GTPase-inactive Rab21:T33N have defective transferrin and epidermal growth factor endocytosis and fail to deliver the latter to lysosomes (22). Rab21 localization Cordycepin also depends on cell polarity; Rab21 has an endoplasmic reticulum MAT1 (ER)-like localization in nonpolarized mammalian cells, whereas Rab21 is present.