Guillen Samander, A. | bioRxiv Science

bioRxiv2026

A new class of lipid transfer proteins is required for the recycling of lipids from the P. falciparum digestive vacuole

Malaria parasites endocytose large quantities of hemoglobin from the host erythrocyte, a process critical for parasite survival, leading to extensive membrane internalization. While hemoglobin degradation in the digestive vacuole (DV) is well studied, how the parasite deals with the membranes arriving within the DV is unknown. Here we identified PfTUPA, a previously uncharacterized lipid transfer protein in the DV membrane that is needed for this function. PfTUPA contains a soluble TULIP-like lipid transport domain exposed to the DV lumen and a transmembrane lipid transfer domain of bacterial origin (PqiA) in the DV membrane. Structural comparisons revealed proteins with various PqiA and TULIP-like domain combinations across distant eukaryotic clades, indicating this is a frequent functional partnership. Hence, PfTUPA belongs to a new class of eukaryotic lipid transfer proteins that in malaria parasites is needed for a key function of its biology.

microbiology↗

bioRxiv2026

Characterization of Host Cell Cytosol-filled Vesicles in the Human Malaria Parasite Plasmodium falciparum

Endocytosis of host cell cytosol is a key process in malaria blood stages. The endocytosed material consists mostly of haemoglobin (Hb) and is transported to the parasite's digestive vacuole (DV), where it is degraded. However, the endosomal transport pathway of the parasite is not well defined. A number of proteins are known that - when inactivated - lead to the appearance of Hb-filled vesicles (HbVs) in the parasite cytoplasm and prevent Hb from arriving in the DV, indicating they play a role in endosomal transport. However, despite the prominence of these HbVs, their morphology, molecular composition, and relationship to the endosomal transport pathway remain poorly understood. Here we carried out a morphological and surface proteome study of HbVs. CryoET showed HbVs are coatless, double membraned vesicles with a very narrow inter-membrane space devoid of larger protein densities. HbV surface proteomes generated by BioIDs most prominently detected DV proteases. Halo-based tracking of one of these DV protease confirmed it as a bona fide HbV cargo. The BioID also identified a number of vesicle trafficking proteins, including PfTBC8 and Rab11 proteins. Functional analysis of PfTBC8 showed its importance for the transport of Hb to the DV. This was due to a novel phenotype characterized by accumulation of diverse Hb-filled structures in the parasite cytosol and gradual decline of DV protease trafficking. PfTBC8 DiQ-BioID detected Rab11a and PfTBC8 inactivation altered Rab11a localization, suggesting it may be a Rab11a effector and that the new endosomal transport phenotype may result from a recycling defect. Together with the detection of VPS35 in the BioID and evidence from the CryoET of a disrupted inner HbV membrane in a proportion of vesicles, these findings support a model in which HbVs undergo maturation prior to fusion with the DV. Overall, our findings define morphological and molecular features of HbVs, identify protein cargo transported through this pathway, and reveal a previously unrecognized Rab-regulatory component required for endosomal trafficking in blood-stage P. falciparum parasites.

microbiology↗