Monitoring phospholipid dynamics during phagocytosis: application of genetically-encoded fluorescent probes.
The internalization of foreign or unwanted particles by cells, a process that is important in many aspects of immunity and development, is known as phagocytosis. Rearrangement of cellular actin enables the phagocytic cell to gradually wrap itself around and ultimately engulf the target particle. Phagocytosis is initiated by receptor engagement that in turn triggers multiple signaling pathways, including extensive lipid remodeling. Lipid modification not only generates a variety of second messengers, but is important for the redistribution of key proteins involved in phagocytosis. Lipids can associate with proteins bearing stereospecific association domains. In addition the phosphoinositides and phosphatidylserine (PS), which are anionic, can also recruit proteins electrostatically by generating a considerable negative surface charge. We describe a method whereby lipids can be monitored dynamically in live cells performing phagocytosis. This method involves the expression by phagocytic cells of genetically encoded fluorescent lipid-binding probes, which can be monitored using confocal fluorescence microscopy.
Program in Cell Biology, The Hospital for Sick Children, Toronto, Canada.
Helen
H
Sarantis
Sergio
S
Grinstein
D000818:Animals; D001665:Binding Sites; D015374:Biosensing Techniques; D002462:Cell Membrane / Q000378:metabolism*; D002478:Cells, Cultured; D049452:Green Fluorescent Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D006801:Humans; D018613:Microscopy, Confocal; D010587:Phagocytosis; D010743:Phospholipids / Q000378:metabolism*; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D059010:Single-Cell Analysis
D010743:Phospholipids; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins