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Vol. 19, Issue 4, 1663-1669, April 2008
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Department of Physiology and Biophysics, Health Science Center, Stony Brook University, Stony Brook, NY 11794-8661
Submitted December 6, 2007;
Revised January 10, 2008;
Accepted January 24, 2008
Monitoring Editor: Tom U. Martin
Phosphatidylinositol 4,5-bisphosphate (PIP2) controls a surprisingly large number of processes in cells. Thus, many investigators have suggested that there might be different pools of PIP2 on the inner leaflet of the plasma membrane. If a significant fraction of PIP2 is bound electrostatically to unstructured clusters of basic residues on membrane proteins, the PIP2 diffusion constant, D, should be reduced. We microinjected micelles of Bodipy TMR-PIP2 into cells, and we measured D on the inner leaflet of fibroblasts and epithelial cells by using fluorescence correlation spectroscopy. The average ± SD value from all cell types was D = 0.8 ± 0.2 µm2/s (n = 218; 25°C). This is threefold lower than the D in blebs formed on Rat1 cells, D = 2.5 ± 0.8 µm2/s (n = 26). It is also significantly lower than the D in the outer leaflet or in giant unilamellar vesicles and the diffusion coefficient for other lipids on the inner leaflet of these cell membranes. The simplest interpretation is that approximately two thirds of the PIP2 on inner leaflet of these plasma membranes is bound reversibly.
Address correspondence to: Stuart McLaughlin (stuart.mclaughlin{at}stonybrook.edu).
Abbreviations used: FCS, fluorescence correlation spectroscopy; GUV, giant unilamellar vesicle; MARCKS, myristoylated alanine rich C kinase substrate.
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