Abstract
Legionella pneumophila infects human alveolar macrophages and is responsible for Legionnaire’s disease, a severe form of pneumonia. L. pneumophila encodes more than 300 putative effectors, which are translocated into the host cell via the Dot/Icm type IV secretion system. These effectors highjack the host’s cellular processes to allow bacterial intracellular growth and replication. Here we adopted a multidisciplinary approach to investigate WipB, a Dot/Icm effector of unknown function. The crystal structure of the N-terminal domain at 1.7 Å resolution comprising residues 25 to 344 revealed that WipB harbours a Ser/Thr phosphatase domain related to the eukaryotic phospho-protein phosphatase (PPP) family. The C-terminal domain (residues 365–524) is sufficient to pilot the effector to acidified LAMP1-positive lysosomal compartments, where WipB interacts with the v-ATPase and the associated LAMTOR1 phosphoprotein, key components of the lysosomal nutrient sensing (LYNUS) apparatus that controls the mammalian target of rapamycin (mTORC1) kinase complex at the lysosomal surface. We propose that WipB is a lysosome-targeted phosphatase that modulates cellular nutrient sensing and the control of energy metabolism during Legionella infection.
Highlights
Legionella pneumophila is a facultative intracellular pathogen responsible for a severe form of pneumonia termed Legionnaire’s Disease[1]
We investigated the ability of the WipB N-terminal domain to catalyse dephosphorylation reactions in vitro using a model phosphothreonine peptide with sequence RRA(pT)VA, a peptide derived from the rat liver pyruvate kinase and routinely used to monitor Ser/Thr phosphatase activities[15]
We determined the structure and characterized the catalytic domain of the Legionella pneumophila effector WipB. We showed that this domain belongs to the protein phosphatase (PPP) family of eukaryotic phosphatases and has a Ser/ Thr phosphatase activity in vitro[21]
Summary
Legionella pneumophila is a facultative intracellular pathogen responsible for a severe form of pneumonia termed Legionnaire’s Disease[1]. Legionella deletion mutants lacking wipB do not exhibit a replicative defect in macrophages, unless this is combined with a second mutation in lidA, a gene encoding another T4S effector capable of binding Rab GTPases. L. pneumophila wipB−lidA− double mutants exhibit attenuated intracellular replication in macrophages and increased host cell death, the significance of this remains unclear[2, 11]. Given this characteristic lack of attenuation in the single gene deletion mutant, a hallmark of functional redundancy, we adopted an alternative structural and biochemical approach to explore the properties and target/s of WipB. (PPP) family, and a C-terminal domain sufficient to pilot WipB to the acidified LAMP1-positive lysosomal compartments in eukaryotic cells, where WipB interacts with the v-ATPase and the associated LAMTOR1 phosphoprotein, key components of the lysosomal nutrient sensing (LYNUS) apparatus
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