Vol 5: Live-Cell Imaging of Phosphoinositide Dynamics and Membrane Architecture during Legionella Infection.Report as inadecuate



 Vol 5: Live-Cell Imaging of Phosphoinositide Dynamics and Membrane Architecture during Legionella Infection.


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This article is from mBio, volume 5.AbstractThe causative agent of Legionnaires’ disease, Legionella pneumophila, replicates in amoebae and macrophages in a distinct membrane-bound compartment, the Legionella-containing vacuole LCV. LCV formation is governed by the bacterial Icm-Dot type IV secretion system that translocates ~300 different -effector- proteins into host cells. Some of the translocated effectors anchor to the LCV membrane via phosphoinositide PI lipids. Here, we use the soil amoeba Dictyostelium discoideum, producing fluorescent PI probes, to analyze the LCV PI dynamics by live-cell imaging. Upon uptake of wild-type or Icm-Dot-deficient L. pneumophila, PtdIns3,4,5P3 transiently accumulated for an average of 40 s on early phagosomes, which acquired PtdIns3P within 1 min after uptake. Whereas phagosomes containing ΔicmT mutant bacteria remained decorated with PtdIns3P, more than 80% of wild-type LCVs gradually lost this PI within 2 h. The process was accompanied by a major rearrangement of PtdIns3P-positive membranes condensing to the cell center. PtdIns4P transiently localized to early phagosomes harboring wild-type or ΔicmT L. pneumophila and was cleared within minutes after uptake. During the following 2 h, PtdIns4P steadily accumulated only on wild-type LCVs, which maintained a discrete PtdIns4P identity spatially separated from calnexin-positive endoplasmic reticulum ER for at least 8 h. The separation of PtdIns4P-positive and ER membranes was even more pronounced for LCVs harboring ΔsidC-sdcA mutant bacteria defective for ER recruitment, without affecting initial bacterial replication in the pathogen vacuole. These findings elucidate the temporal and spatial dynamics of PI lipids implicated in LCV formation and provide insight into host cell membrane and effector protein interactions.



Author: Weber, Stephen; Wagner, Maria; Hilbi, Hubert

Source: https://archive.org/







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