Fig. 4: PI4P extraction assay.
From: Coordination of transporter, cargo, and membrane properties during non-vesicular lipid transport
A Scheme of the assay. The amount of accessible PI4P (nPI4P(acc.)) equals the amount of Osh6. The FCCS read-out, Gcc, monitoring the mutual motion of DiD and SidC-Atto488 disappears upon the Osh6 addition. B Scheme of the assay when other, PI4P-free LUVs (marked as B) are present during the extraction. The LUVs B either do not contain the competing cargo (1)—the extraction is not affected or contain the competing cargo (2)—the extraction is compromised. C Dependence of the Gcc read-out of SidC-Atto488 biosensor in our experimental system. The red line depicts the concentration region of PI4P where the extraction takes place. D Kinetics of the PI4P extraction from the PI4P containing LUVs when no other LUVs are present (black), and at presence of other, PI4P-free LUVs of various compositions: POPC (red), POPC/POPE (blue), POPC/POPG (green), POPC/POPA (orange), POPC/soy PI (dark yellow), POPC/POPS (violet), POPC/PIP2 (magenta). E PI4P extraction rates for each composition of LUVs B. Composition of the experiment in 3D, E: cPI4P = 500 nM, cOsh6 = 250 nM, cSidC-Atto488 = 100 nM, ctotal lipids LUV A = 50 µM, ctotal lipids LUV B = 200 µM. All error bars represent the standard error of the mean, n = 10 measurements. The p-values were obtained from the two-sample t-test.
