Modulation of host phosphoinositide metabolism during Salmonella invasion by the type III secreted effector SopB.
Phosphoinositides (PI) play an important role in many different cellular processes. Their generation and functions, however, are very dynamic, and the detection of localized events usually requires very precise imaging techniques. Recent advances in lipid research raised the possibility of designing molecular probes to specifically detect lipids in different subcellular compartments and have provided new tools to directly image PI dynamics in living cells. Salmonella is a pathogenic bacterium that has the ability to invade host cells and grow intracellularly. To this end, they secrete specialized virulence proteins (effectors) directly into the cytosol of host cells. These effectors modulate signaling pathways to initiate bacterial uptake and promote intracellular survival. SopB, one of the many effector proteins that are translocated into host cells, has PI phosphatase activity and directly modulates PI metabolism. In this chapter, we describe a method to transfect PI-binding domains fused to fluorescent proteins as probes to monitor lipid dynamics during a Salmonella invasion in living cells using a spinning-disk confocal microscope.
Cell Biology Program, Hospital for Sick Children, Toronto, Ontario, Canada.
Bernhard
B
Roppenser
Sergio
S
Grinstein
John H
JH
Brumell
D000818:Animals; D001426:Bacterial Proteins / Q000378:metabolism / Q000502:physiology*; D015374:Biosensing Techniques; D002478:Cells, Cultured; D054884:Host-Pathogen Interactions; D006801:Humans; D008856:Microscopy, Fluorescence; D008954:Models, Biological; D010716:Phosphatidylinositols / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D012480:Salmonella Infections / Q000382:microbiology*; D012486:Salmonella typhimurium / Q000201:enzymology / Q000502:physiology*
D001426:Bacterial Proteins; D010716:Phosphatidylinositols; D011993:Recombinant Fusion Proteins; C489224:SopB protein, Salmonella