@article {65059,
	title = {Analysis of phosphatidylinositol transfer at ER-PM junctions in receptor-stimulated live cells.},
	journal = {Methods in Molecular Biology},
	volume = {1949},
	year = {2019},
	month = {2019 Feb 23},
	pages = {1-11},
	abstract = {<p>Phosphatidylinositol (PI) is an inositol-containing phospholipid synthesized in the endoplasmic reticulum (ER). PI is a precursor lipid for PI 4,5-bisphosphate (PI(4,5)P) in the plasma membrane (PM) important for Ca signaling in response to extracellular stimuli. Thus, ER-to-PM PI transfer becomes essential for cells to maintain PI(4,5)P homeostasis during receptor stimulation. In this chapter, we discuss two live-cell imaging protocols to analyze ER-to-PM PI transfer at ER-PM junctions, where the two membrane compartments make close appositions accommodating PI transfer. First, we describe how to monitor PI(4,5)P replenishment following receptor stimulation, as a readout of PI transfer, using a PI(4,5)P biosensor and total internal reflection fluorescence\&nbsp;microscopy. The second protocol directly visualizes PI transfer proteins that accumulate at ER-PM junctions and mediate PI(4,5)P replenishment with PI in the ER in stimulated cells. These methods provide spatial and temporal analysis of ER-to-PM PI transfer during receptor stimulation and can be adapted to other research questions related to this topic.</p>
},
	keywords = {Biological Transport, Biosensing Techniques, Cell Membrane, Endoplasmic Reticulum, Hela Cells, Humans, Image Processing, Computer-Assisted, Microscopy, Phosphatidylinositol 4,5-Diphosphate, Phosphatidylinositols},
	issn = {1940-6029},
	doi = {10.1007/978-1-4939-9136-5_1},
	author = {Chang, Chi-Lun and Liou, Jen}
}
