Leishmania major promastigote entry of the of the autophagic compartment and amastigote escape from the parasitophorous vacuole

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During the first three years we will (i) visualize the leishmania major containing parasitophorous vacuoles in human macrophages using state of the art high pressure freezing and electron tomography focusing on promastigote interactions with autophagy and amastigote escape from the parasitophorous vacuoles (ii) we will try to down regulate autophagy and lysosomal maturation in macrophage cell lines and primary macrophages (iii) we will establish a new assay for correlative electron microscopy. During the second 3-year funding period (i) we aim to dissect the molecular basis leading to promastigote entry of the autophagic pathway and amastigote escape. (ii) We will apply the correlative electron microcopy technique on new target proteins coming from our mass spectrometry screen. In addition we will test L. donovani known to block phagolysosomal maturation and L. amazonensis known to induce large parasitophorous vacuoles. (iii) Moreover we will focus on the parasitophorous vacuoles in human neutrophils that can only be infected with promastigotes and dendritic cells that can only be infected with amastigotes.