Endosomal Localization of the Serum Resistance-Associated Protein in African Trypanosomes Confers Human Infectivity

Abstract

Trypanosoma brucei rhodesiense is the causative agent of human African sleeping sickness. While the closely related subspecies T. brucei brucei is highly susceptible to lysis by a subclass of human high-density lipoproteins (HDL) called trypanosome lytic factor (TLF), T. brucei rhodesiense is resistant and therefore able to establish acute and fatal infections in humans. This resistance is due to expression of the serum resistance-associated (SRA) gene, a member of the variant surface glycoprotein (VSG) gene family. Although much has been done to establish the role of SRA in human serum resistance, the specific molecular mechanism of SRA-mediated resistance remains a mystery. Thus, we report the trafficking and steady-state localization of SRA in order to provide more insight into the mechanism of SRA-mediated resistance. We show that SRA traffics to the flagellar pocket of bloodstream-form T. brucei organisms, where it localizes transiently before being endocytosed to its steady-state localization in endosomes, and we demonstrate that the critical point of colocalization between SRA and TLF occurs intracellularly.

Human trypanolytic factor APOL1 forms pH-gated cation-selective channels in planar lipid bilayers: Relevance to trypanosome lysis

“…We reasoned that if the APOL1-induced conductance was truly relevant to the mechanism of trypanosome lysis, then it should be prevented by preadding recombinant SRA to the cis compartment at pH 5.3, a condition that would mimic the endosomal lumen of T. b. rhodesiense (17). In fact, when APOL1 was added to the cis side in the presence of SRA at pH 5.3, we still observed a minor conductance increase, which was retained after perfusion of the cis side at acidic pH, but the usual amplification upon raising the cis pH was completely prevented (Fig.…”

“…In this model, membrane insertion of APOL1 is accomplished by three putative transmembrane sequences, one of which resides in the APOL1 C-terminal domain (19). The SRA protein of human serumresistant T. b. rhodesiense (oval), which is thought to be membraneanchored via a glycosylphosphatidylinositol group (not shown for clarity), is reported to cycle between the plasma membrane and the endocytic system, where it can bind to the C terminus of APOL1 at acidic pH (17). SRA may prevent insertion of the C-terminal transmembrane sequence of APOL1, allowing dissociation of APOL1 from the membrane upon recycling to neutral pH conditions of the cell surface, where APOL1 also dissociates from SRA.…”

“…Whereas multifactorial mechanisms were proposed to account for resistance of T. b. gambiense to human serum (13)(14)(15)(16), resistance in the case of T. b. rhodesiense is determined by the parasite's serum resistance-associated (SRA) protein, which localizes to parasite endosomes and binds to APOL1 under acidic conditions (10,17). Primate APOL1 orthologs, and two African human APOL1 variants, contain mutations in the C-terminal SRA-binding site that reduce binding affinity for SRA, allowing these variants to lyse SRA-producing trypanosomes (18)(19)(20).…”

Evolution of the primate trypanolytic factor APOL1

“…Upon TLF binding to HpHbR, TLF1/APOL1 traffics to the lysosome, where a pH-dependent conformational switch is believed to trigger APOL1 insertion into the endosomal/lysosomal membrane (12). Here, APOL1 forms a pore permeable to monovalent ions, leading to lysosomal membrane depolarization, depolarization of the plasma membrane, osmotic swelling, and rupture (22,23).…”

“…Human and gorilla sera protect against Trypanosoma brucei brucei, whereas baboon sera protect against T. b. brucei, Trypanosoma brucei rhodesiense, and potentially, Trypanosoma brucei gambiense (8,9). Human-infective T. b. rhodesiense, which causes acute African sleeping sickness, arose from animal-infective T. b. brucei through the evolution of a virulence factor called serum resistance-associated protein (SRA) (10) that binds to (11) and prevents human APOL1-mediated lysis (12), whereas T. b. gambiense evolved different mechanisms to neutralize human APOL1 (13)(14)(15).…”