PfATG18 is transported to the FV by haemoglobin trafficking pathway. Left and right: Localisation of PfATG18 upon treatment with haemoglobin trafficking inhibitors: 7 μM jasplakinolide (JAS), 25 mM 2,3-Butanedione monoxime (BDM) and 200 μM Dynasore. Immunofluorescence assay of PfATG18 with A. falcipain-2 (FP2), or B. phospholipid PI3P using Zenon antibody labelling system. Upon treatment with haemoglobin trafficking inhibitors, PfATG18 partially colocalises with FP2 and PI3P. Enlarged panel: magnified view of the dotted region shown in the Merge panel. Nucleus was stained with Hoechst. The Pearson’s coefficient (R) represents the degree of colocalisation between PfATG18 and FP2 or PI3P evaluated from the signal within the square region on the cytoplasmic vesicles shown in the enlarged panel, and their localisation is indicated by the arrows in Merge+Hoechst+DIC panel. N=10 cells, n=3 experiments, Scale bar=5 μm. To dissect the PfATG18 trafficking to the FV via HCv, this pathway was pharmacologically inhibited by jasplakinolide, BDM and Dynasore, and the localisation of PfATG18 was examined. In control parasites, PfATG18 was found on the periphery of the FV with FP2 signal inside it (let). Upon treatment with jasplakinolide or BDM, an increased number of PfATG18-labelled vesicles were observed at the parasite boundary (left), and colocalised with FP2 cytoplasmic vesicles (left). Additionally, in parasites treated with Dynasore, thin tube-like structures labelled with FP2 were observed and PfATG18 was found to localise on them. Since PfATG18 requires PI3P for it to localise to the membrane, we investigated if the trafficking of PfATG18 to FV is via PI3P-labelled vesicles. The parasites were treated with jasplakinolide, BDM and Dynasore to inhibit PfATG18 trafficking. In control parasites, PfATG18 colocalised with PI3P on FV (right), whereas upon treatments, the trafficking of PI3P- and PfATG18-decorated vesicles to FV was blocked and the colocalisation between them was observed at the cytoplasmic puncta (right), instead of on the FV, indicating that the targeting of PfATG18 to FV is PI3P-dependent.
Agrawal P, Manjithaya R, Surolia N. Autophagy-related protein PfATG18 participates in food vacuole dynamics and autophagy-like pathway in Plasmodium falciparum. Mol Microbiol. 2019 Dec 21
Other associated proteins
|PF3D7_1012900||autophagy-related protein 18|