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. 2003 Nov 10;163(3):559–570. doi: 10.1083/jcb.200302157

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Copyright © 2003, The Rockefeller University Press

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Figure 9. — The effects of Rab27A-Q78L and MyRIP on secretion depend on the state of the actin cortex. (A) Rhodamine-phalloidin staining of the actin cortex of PC12 cells. Compared with control conditions (left), the thickness of actin is reduced upon latrunculin B treatment (middle) and increased by jasplakinolide (right). (B) The secretory activity of PC12 cells transfected with vectors encoding GFP (as control), Rab27A-Q78L, wild-type MyRIP (MyRIP 1–859), MyRIP-ΔCter (1–665), or granuphilin was measured as in Fig. 5 by means of the [³H]5-HT release assay. Cells were incubated without addition of drug (Std, black bars) or in the presence of 1 μM jasplakinolide (Jasp, open bars) or 5 μM latrunculin B (Lat B, gray bars) for 20 min before and during stimulation of secretion. The effect of the different proteins on secretion was expressed, after subtracting the basal release of [³H]5-HT in normal saline, as percentage of the stimulus-dependent secretory response of GFP-transfected cells measured under the same conditions (i.e., in the absence of drug, in the presence of jasp, or in the presence of lat B). Shown are the results (mean ± SEM) of several independent experiments (the number of experiments is shown above the bars). Latrunculin B significantly reduced (P < 0.001) the inhibitory effects of Rab27A-Q78L and MyRIP on secretion, whereas jasplakinolide increased that of MyRIP (P < 0.001). The net release of GFP-transfected cells was 19 ± 2% of cellular [³H]5-HT in the absence of drug, 20 ± 2.2% in the presence of jasplakinolide, and 31 ± 3% in the presence of latrunculin B.

Figure 9. — The effects of Rab27A-Q78L and MyRIP on secretion depend on the state of the actin cortex. (A) Rhodamine-phalloidin staining of the actin cortex of PC12 cells. Compared with control conditions (left), the thickness of actin is reduced upon latrunculin B treatment (middle) and increased by jasplakinolide (right). (B) The secretory activity of PC12 cells transfected with vectors encoding GFP (as control), Rab27A-Q78L, wild-type MyRIP (MyRIP 1–859), MyRIP-ΔCter (1–665), or granuphilin was measured as in Fig. 5 by means of the [³H]5-HT release assay. Cells were incubated without addition of drug (Std, black bars) or in the presence of 1 μM jasplakinolide (Jasp, open bars) or 5 μM latrunculin B (Lat B, gray bars) for 20 min before and during stimulation of secretion. The effect of the different proteins on secretion was expressed, after subtracting the basal release of [³H]5-HT in normal saline, as percentage of the stimulus-dependent secretory response of GFP-transfected cells measured under the same conditions (i.e., in the absence of drug, in the presence of jasp, or in the presence of lat B). Shown are the results (mean ± SEM) of several independent experiments (the number of experiments is shown above the bars). Latrunculin B significantly reduced (P < 0.001) the inhibitory effects of Rab27A-Q78L and MyRIP on secretion, whereas jasplakinolide increased that of MyRIP (P < 0.001). The net release of GFP-transfected cells was 19 ± 2% of cellular [³H]5-HT in the absence of drug, 20 ± 2.2% in the presence of jasplakinolide, and 31 ± 3% in the presence of latrunculin B.