FIGURE 7.

Formed MYH9 aggregates is crucial for virion internalization to MARC-145 cells. (A) Co-localization in MARC-145 cells between PRRSV virions and MYH9. After exposure to PRRSV SD16 strain (MOI = 50) at 4°C for 2 h, MARC-145 cells were transferred to 37°C for 0, 5, 15, and 30 min. Cells were fixed, permeabilized and stained with anti-MYH9 pAb (Red) and 6D10 (mAb against PRRSV N protein, in house, Green) to visualize distribution of endogenous MYH9 and viral particles. Representative images are shown, representing one confocal z-section through the middle of the cells. Bars, 10 μm. (B) The co-localization of MYH9 and PRRSV described in (A) was assessed by determination of Manders’ overlap coefficient using Image-Pro Plus software. The Mean of Manders’ overlapcoefficient ±SD are representative of three individual enlarged pictures. (C) Native-PAGE for aggregated MYH9 and Western blot for internalized PRRSV virions in MARC-145 cells during PRRSV internalization. MARC-145 cells were incubated with SD16 at MOI = 50 at 4°C for 2 h following by shifting to 37°C, and then harvested at indicated time points (0, 5, 15, 30 min). Cells were lysed and subjected to native-PAGE and Western blot using anti-MYH9 pAb and anti-PRRSV N mAb (6D10). Values are normalized to uninfected MARC-145 cells and the fold of relative expression was indicated.