The causative agent of malaria, Plasmodium, replicates inside a membrane-bound parasitophorous vacuole (PV), which shields this intracellular parasite from the cytosol of the host cell1. One common threat for intracellular pathogens is the homeostatic process of autophagy, through which cells capture unwanted intracellular material for lysosomal degradation2. During the liver stage of a malaria infection, Plasmodium parasites are targeted by the autophagy machinery of the host cell, and the PV membrane (PVM) becomes decorated with several autophagy markers, including LC3 (microtubule-associated protein 1 light chain 3)3,4. Here we show that Plasmodium berghei parasites infecting hepatic cells rely on the PVM transmembrane protein UIS3 to avoid elimination by host-cell-mediated autophagy. We found that UIS3 binds host LC3 through a non-canonical interaction with a specialized surface on LC3 where host proteins with essential functions during autophagy also bind. UIS3 acts as a bona fide autophagy inhibitor by competing with host LC3-interacting proteins for LC3 binding. Our work identifies UIS3, one of the most promising candidates for a genetically attenuated vaccine against malaria5, as a unique and potent mediator of autophagy evasion in Plasmodium. We propose that the protein–protein interaction between UIS3 and host LC3 represents a target for antimalarial drug development.
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Liehl, P., Zuzarte-Luis, V. & Mota, M. M. Unveiling the pathogen behind the vacuole. Nat. Rev. Microbiol. 13, 589–598 (2015).
Gomes, L. C. & Dikic, I. Autophagy in antimicrobial immunity. Mol. Cell 54, 224–233 (2014).
Prado, M. et al. Long-term live imaging reveals cytosolic immune responses of host hepatocytes against Plasmodium infection and parasite escape mechanisms. Autophagy 11, 1561–1579 (2015).
Thieleke-Matos, C. et al. Host cell autophagy contributes to Plasmodium liver development. Cell. Microbiol. 18, 437–450 (2016).
Mueller, A.-K., Labaied, M., Kappe, S. H. I. & Matuschewski, K. Genetically modified Plasmodium parasites as a protective experimental malaria vaccine. Nature 433, 164–167 (2005).
Shen, H.-M. & Mizushima, N. At the end of the autophagic road: an emerging understanding of lysosomal functions in autophagy. Trends Biochem. Sci. 39, 61–71 (2014).
Mueller, A.-K. et al. Plasmodium liver stage developmental arrest by depletion of a protein at the parasite–host interface. Proc. Natl Acad. Sci. USA 102, 3022–3027 (2005).
Hanson, K. K. et al. Torins are potent antimalarials that block replenishment of Plasmodium liver stage parasitophorous vacuole membrane proteins. Proc. Natl Acad. Sci. USA 110, E2838–E2847 (2013).
Spielmann, T., Montagna, G. N., Hecht, L. & Matuschewski, K. Molecular make-up of the Plasmodium parasitophorous vacuolar membrane. Int. J. Med. Microbiol. 302, 179–186 (2012).
Mizushima, N. et al. Dissection of autophagosome formation using Apg5-deficient mouse embryonic stem cells. J. Cell Biol. 152, 657–667 (2001).
Ganley, I. G., Wong, P.-M., Gammoh, N. & Jiang, X. Distinct autophagosomal–lysosomal fusion mechanism revealed by thapsigargin-induced autophagy arrest. Mol. Cell 42, 731–743 (2011).
Sturm, A. et al. Alteration of the parasite plasma membrane and the parasitophorous vacuole membrane during exo-erythrocytic development of malaria parasites. Protist 160, 51–63 (2009).
Sharma, A., Yogavel, M., Akhouri, R. R., Gill, J. & Sharma, A. Crystal structure of soluble domain of malaria sporozoite protein UIS3 in complex with lipid. J. Biol. Chem. 283, 24077–24088 (2008).
Mikolajczak, S. A., Jacobs-Lorena, V., MacKellar, D. C., Camargo, N. & Kappe, S. H. I. L-FABP is a critical host factor for successful malaria liver stage development. Int. J. Parasitol. 37, 483–489 (2007).
Favretto, F., Assfalg, M., Molinari, H. & D’Onofrio, M. Evidence from NMR interaction studies challenges the hypothesis of direct lipid transfer from L-FABP to malaria sporozoite protein UIS3. Prot. Sci. 22, 133–138 (2013).
Farré, J.-C. & Subramani, S. Mechanistic insights into selective autophagy pathways: lessons from yeast. Nat. Rev. Mol. Cell Biol. 17, 537–552 (2016).
Wacker, R. et al. LC3-association with the parasitophorous vacuole membrane of Plasmodium bergheiliver stages follows a noncanonical autophagy pathway. Cell. Microbiol. 19, e12754 (2017).
Martinez, J. et al. Molecular characterization of LC3-associated phagocytosis reveals distinct roles for Rubicon, NOX2 and autophagy proteins. Nat. Cell Biol. 17, 893–906 (2015).
Noda, N. N., Ohsumi, Y. & Inagaki, F. Atg8-family interacting motif crucial for selective autophagy. FEBS Lett. 584, 1379–1385 (2010).
Birgisdottir, Å. B., Lamark, T. & Johansen, T. The LIR motif—crucial for selective autophagy. J. Cell Sci. 126, 3237–3247 (2013).
Ichimura, Y. et al. Structural basis for sorting mechanism of p62 in selective autophagy. J. Biol. Chem. 283, 22847–22857 (2008).
Pankiv, S. et al. p62/SQSTM1 binds directly to Atg8/LC3 to facilitate degradation of ubiquitinated protein aggregates by autophagy. J. Biol. Chem. 282, 24131–24145 (2007).
Mizushima, N., Yoshimori, T. & Levine, B. Methods in mammalian autophagy research. Cell 140, 313–326 (2010).
Klionsky, D., Abdalla, F. & Abeliovich, H. Guidelines for the use and interpretation of assays for monitoring autophagy. Autophagy 8, 445–544 (2012).
Barrios-Rodiles, M. et al. High-throughput mapping of a dynamic signaling network in mammalian cells. Science 307, 1621–1625 (2005).
Zaffagnini, G. & Martens, S. Mechanisms of selective autophagy. J. Mol. Biol. 428, 1714–1724 (2016).
McEwan, D. G. et al. PLEKHM1 regulates autophagosome–lysosome fusion through HOPS complex and LC3/GABARAP proteins. Mol. Cell 57, 39–54 (2015).
Boonhok, R. et al. LAP-like process as an immune mechanism downstream of IFN-γ in control of the human malaria Plasmodium vivax liver stage. Proc. Natl Acad. Sci. USA 113, E3519–E3528 (2016).
Ruivo, M. T. G. et al. Host AMPK is a modulator of Plasmodiumliver infection. Cell Rep. 16, 2539–2545 (2016).
Kumar, H. et al. Protective efficacy and safety of liver stage attenuated malaria parasites. Sci. Rep. 6, 26824 (2016).
Kuma, A. et al. The role of autophagy during the early neonatal starvation period. Nature 432, 1032–1036 (2004).
Sou, Y. et al. The Atg8 conjugation system is indispensable for proper development of autophagic isolation membranes in mice. Mol. Biol. Cell 19, 4762–4775 (2008).
Lee, I. H. et al. Atg7 modulates p53 activity to regulate cell cycle and survival during metabolic stress. Science 336, 225–228 (2012).
Zhang, W. et al. PCB 126 and other dioxin-like PCBs specifically suppress hepatic PEPCK expression via the aryl hydrocarbon receptor. PLoS ONE 7, e37103 (2012).
Gonçalves, L. A., Vigário, A. M. & Penha-Gonçalves, C. Improved isolation of murine hepatocytes for in vitro malaria liver stage studies. Malar. J. 6, 169 (2007).
Itoe, M. A. A. et al. Host cell phosphatidylcholine is a key mediator of malaria parasite survival during liver stage infection. Cell Host Microbe 16, 778–786 (2014).
Janse, C. J., Ramesar, J. & Waters, A. P. High-efficiency transfection and drug selection of genetically transformed blood stages of the rodent malaria parasite Plasmodium berghei. Nat. Protoc. 1, 346–356 (2006).
Tsuji, M., Mattei, D., Nussenzweig, R. S., Eichinger, D. & Zavala, F. Demonstration of heat-shock protein 70 in the sporozoite stage of malaria parasites. Parasitol. Res. 80, 16–21 (1994).
Fentress, S. J. et al. Phosphorylation of immunity-related GTPases by a Toxoplasma gondii-secreted kinase promotes macrophage survival and virulence. Cell Host Microbe 8, 484–495 (2010).
Xu, D. & Zhang, Y. Improving the physical realism and structural accuracy of protein models by a two-step atomic-level energy minimization. Biophys. J. 101, 2525–2534 (2011).
Torchala, M., Moal, I. H., Chaleil, R. A. G., Fernandez-Recio, J. & Bates, P. A. SwarmDock: a server for flexible protein–protein docking. Bioinformatics 29, 807–809 (2013).
Pronk, S. et al. GROMACS 4.5: a high-throughput and highly parallel open source molecular simulation toolkit. Bioinformatics 29, 845–854 (2013).
Vangone, A., Oliva, R. & Cavallo, L. CONS-COCOMAPS: a novel tool to measure and visualize the conservation of inter-residue contacts in multiple docking solutions. BMC Bioinformatics 13(Suppl. 4), S19 (2012).
Laskowski, R. A. & Swindells, M. B. LigPlot+: multiple ligand–protein interaction diagrams for drug discovery. J. Chem. Inf. Model. 51, 2778–2786 (2011).
Kelly, S. M., Jess, T. J. & Price, N. C. How to study proteins by circular dichroism. Biochim. Biophys. Acta 1751, 119–139 (2005).
Van Wesenbeeck, L. et al. Involvement of PLEKHM1 in osteoclastic vesicular transport and osteopetrosis in incisors absent rats and humans. J. Clin. Invest. 117, 919–930 (2007).
The authors thank M. Komatsu (Tokyo Metropolitan Institute of Medical Science) for providing Atg3 MEFs, T. Finkel (NIH National Heart, Lung, and Blood Institute) for the Atg7 MEFs, F. Randow (MRC Laboratory of Molecular Biology) for the gift of p62-luciferase and GST-LC3 expression plasmids, Jacobus Pharmaceuticals for the WR99210 compound, A. Parreira for producing P. berghei-infected Anopheles mosquitoes, and S. Marques and K. Slavic for their help with parasite cloning. This work was supported by grants from the European Research Council (ERC‐2012‐StG_311502 to M.M.M.), Fundação para a Ciência e Tecnologia (EXCL/IMI-MIC/0056/2012 to M.M.M. and PTDC/IMI-MICC/1568/2012 to G.G.C.) and Institut Mérieux (MRG_20052016 to M.M.M). E.R. was the recipient of EMBO (ALTF 949-2008) and FCT (SFRH/BPD/68709/2010) fellowships. L.R. is the recipient of FCT fellowship SFRH/BPD/111323/2015. G.G.C. was sponsored by Marie Curie (PIEF-GA-2009-235864) and FCT (SFRH/BPD/74151/2010) fellowships. L.M.S. was supported by the European Community's Seventh Framework Programme (FP7/2007-2013) under grant agreement no. 242095 (EVIMalaR). V.Z.L. was sponsored by EMBO (ALTF 357-2009) and FCT (BPD-81953-2011) fellowships. I.M.V. and J.M.-V. were supported by NIH (1F32A11042-5 021) and FCT (SFRH/BD/52226/2013) fellowships, respectively.
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Scientific Reports (2019)
Nature Reviews Molecular Cell Biology (2018)