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. 2009 Jun 15;20(12):2991–3002. doi: 10.1091/mbc.E08-10-1074

Figure 5.

Figure 5.

Steady-state Kv1.2 homeostasis is composed of constitutive cholesterol-dependent endocytosis and of constitutive recycling. (A) Application of the sterol-binding agent filipin (7.7 μM; 1 h) to HEK-K cells significantly increases steady-state surface Kv1.2 as detected by flow cytometry. Subsequent treatment with Y27632 (10 μM; 30 min) does not elicit a further increase (n = 32; *p < 0.01, **p < 0.001). (B) Filipin application does not prevent Kv1.2 endocytosis upon application of LPA (10 μM; 15 min) (n = 5; *p < 0.05, **p < 0.001). (C) Application of filipin (7.7 μM; 1 h) to HEK-K cells significantly increases surface Kv1.2 at both 37 and 16°C (n = 12; **p < 0.001). Inset, application of filipin at 16°C results in a significantly greater increase of surface Kv1.2 compared with 37°C control (n = 12; *p = 0.019). (D) Y27632 (10 μM; 30 min) does not elicit an increase of steady-state surface Kv1.2 at 16°C (n ≥10; **p < 0.001). Inset, percentage of increase in surface Kv1.2 elicited by Y27632 at 37 versus 16°C is significantly different (n ≥10, *p < 0.01). (E) Incubation at 16°C results in significantly reduced levels of steady-state surface Kv1.2 compared with 37°C control (n = 27; **p < 0.001).