Figure 6
Induction of Cav1 phosphorylation during C. neoformans infection. (A) HBMEC were infected with C. neoformans B-4500FO2 for different time periods, and then cell lysates (50 μg) were blotted and probed with an anti-phospho-Cav1 (Tyr-14) antibody. A time course study of Cav1 phosphorylation at 0, 5, 10, and 30 min is shown. β-actin was used as the loading control (second row). The ECL films (protein blots) were scanned by the software ImageMaster 2D Platinum 6.0 for quantitative and graphic analyses (lower panel). The total intensity of all bands is 100%. The relative intensity of the bands is indicated on the y-axis (n = 3). (B) Distribution of Pi-Cav1 during C. neoformans infection was examined by immunofluorescence microscopy. Untreated HBMEC were used as the control (1), or the HBMEC monolayer was incubated with 106 C. neoformans strain B-4500FO2 for (2) 5 min and (3) 30 min. Pi-Cav1 was stained with anti-phospho-Cav1 (Tyr-14) and second antibody-FITC conjugate (green). Nuclei were stained with DAPI (blue). Bar: 15 μm.