Abstract
The human innate immunity factor apolipoprotein L-I (APOL1) protects against infection by several protozoan parasites, including Trypanosoma brucei brucei Endocytosis and acidification of high-density lipoprotein-associated APOL1 in trypanosome endosomes leads to eventual lysis of the parasite due to increased plasma membrane cation permeability, followed by colloid-osmotic swelling. It was previously shown that recombinant APOL1 inserts into planar lipid bilayers at acidic pH to form pH-gated nonselective cation channels that are opened upon pH neutralization. This corresponds to the pH changes encountered during endocytic recycling, suggesting APOL1 forms a cytotoxic cation channel in the parasite plasma membrane. Currently, the mechanism and domains required for channel formation have yet to be elucidated, although a predicted helix-loop-helix (H-L-H) was suggested to form pores by virtue of its similarity to bacterial pore-forming colicins. Here, we compare recombinant human and baboon APOL1 orthologs, along with interspecies chimeras and individual amino acid substitutions, to identify regions required for channel formation and pH gating in planar lipid bilayers. We found that whereas neutralization of glutamates within the H-L-H may be important for pH-dependent channel formation, there was no evidence of H-L-H involvement in either pH gating or ion selectivity. In contrast, we found two residues in the C-terminal domain, tyrosine 351 and glutamate 355, that influence pH gating properties, as well as a single residue, aspartate 348, that determines both cation selectivity and pH gating. These data point to the predicted transmembrane region closest to the APOL1 C terminus as the pore-lining segment of this novel channel-forming protein.
Highlights
Apolipoprotein L-I (APOL1) is a channel-forming innate immune factor that circulates on serum high-density lipoproteins [1]
Whereas C-terminal differences account for lysis of T. brucei rhodesiense, the mechanism by which T. brucei gambiense is lysed by baboon APOL1 has yet to be identified
It was previously shown that human APOL1 forms pH-gated channels in planar lipid bilayers and kills African trypanosomes via colloid-osmotic lysis [1, 12, 14]
Summary
Apolipoprotein L-I (APOL1) is a channel-forming innate immune factor that circulates on serum high-density lipoproteins [1]. Following endocytic uptake of high-density lipoprotein– associated APOL1 into acidic parasite endosomes, African trypanosomes undergo colloid-osmotic swelling and eventual lysis due to an increased plasma membrane cation permeability, which is proposed to result from APOL1 recycling to the plasma membrane, where it forms nonselective cation channels [12, 13]. In agreement with this model, cation channel formation by recombinant human APOL1 requires acidic pH (as in endosomes), whereas channel opening requires subsequent pH neutralization (as in recycling to the plasma membrane) [14, 15]. It remains to be seen whether these deficiencies result from misfolding of these protein fragments
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