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Hubert Hilbi (2011-2014)

Functional phosphoinositide lipidomics of Legionella-containing vacuoles

Legionella spp. are environmental bacteria that grow within free-living amoebae and within macrophages in the lung, thereby causing a severe pneumonia termed “Legionnaires’ disease”. The bacteria replicate in host cells in distinct membrane-bound compartments, the “Legionella-containing vacuoles” (LCVs). Formation of LCVs is a complex and robust process, which involves a bacterial type IV secretion system and more than 100 different secreted “effector” proteins. These proteins are injected into host cells, where they modulate signal transduction and membrane dynamics. The aims of this project comprise a detailed molecular analysis using biochemical, genetic and cellular microbial methods of host phosphoinositide (PI) lipids and other lipids implicated in LCV formation by Legionella spp. To this end, we will purify intact LCVs and determine the PIs, the lipidome and the proteome by mass spectrometry. To functionally characterize the role of PIs in the formation of LCVs, we will (i) establish and compare PI and lipid catalogs of purified intact LCVs of Legionella pneumophila and Legionella longbeachae, (ii) analyze the dynamics of PIs and PImetabolizing enzymes on LCVs in real-time, (iii) characterize PI kinases and PI phosphatases implicated in LCV formation, and (iv) study the functional role of PIs for LCV formation.



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