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. 2004 Feb;15(2):751–760. doi: 10.1091/mbc.E03-05-0307

Figure 2.

Figure 2.

PIM stimulates early endosomal fusion in ATP-, NSF-, and cytosol-dependent manner. (A) In vitro fusion was carried out, in the presence or absence of 80 μg/ml PIM for 30 min. Reactions contained: 1 mg/ml cytosol or no cytosol (control); ATP regenerating system or an ATP-depleting system (no ATP). Inset, quantitation of extravesicular probes (avidin and biotinylated HRP), in the presence or absence of 80 μg/ml PIM. (B) NSF dependency of PIM-stimulated fusion. Endosomes and cytosol were preincubated for 20 min with 0.25 mM NEM followed by initiating fusion reaction with 1 mg/ml cytosol and incubation for 30 min. (C) Cytosol concentration effects on PIM-mediated stimulation of endosomal fusion. Fusion reaction was allowed to occur for 30 min in the presence of different concentrations of cytosol in the presence or absence of 80 μg/ml PIM. (D) PIM effects on rate and extent of endosomal fusion. Fusion was conducted in the presence of 2 mg/ml cytosol. Inset, percentage of ATP-dependent fusion in the presence or absence of PIM at 10 and 20 min. All percent values are given relative to the control fusion reaction without PIM. Bars, SEM asterisk, p < 0.05 (ANOVA).