Figure 3. Actin regulatory proteins... | Rockefeller University Press

Figure 3.

$Figure 3. Actin regulatory proteins moesin and α-actinin-1 regulate the lateral mobility of ICAM-1. (A and B) Western blots showing levels of total and phosphorylated ERM proteins (A) and α-actinin1 in lysates from BMDCs matured with 100 ng/ml of LPS for 24 or 48 h (B). E, ezrin; M, moesin. (C and D) Immunofluorescence micrographs showing the distribution of F-actin and moesin with respect to cell surface ICAM-1 (C) or MHC II (D). Bottom panels show enlarged regions indicated by the white boxes. (E) Midplane of cell prepared as in C, demonstrating co-capping of ICAM-1 and moesin. (F) Western blot showing siRNA-mediated knockdown of either moesin (M), α-actinin 1 (A), or both proteins (M/A) in mature BMDCs. (G and H) Mature BMDCs treated with siRNA as in F were surface labeled with Fabs against MHCII or ICAM-1, and FRAP analysis was performed to determine the mobile fraction (G) and diffusion coefficient (H). Dots represent individual FRAP measurements (n = 143–285) pooled from three independent experiments. *, P < 0.01; ***, P < 0.0001. Bars, 10 µm.$

Actin regulatory proteins moesin and α-actinin-1 regulate the lateral mobility of ICAM-1. (A and B) Western blots showing levels of total and phosphorylated ERM proteins (A) and α-actinin1 in lysates from BMDCs matured with 100 ng/ml of LPS for 24 or 48 h (B). E, ezrin; M, moesin. (C and D) Immunofluorescence micrographs showing the distribution of F-actin and moesin with respect to cell surface ICAM-1 (C) or MHC II (D). Bottom panels show enlarged regions indicated by the white boxes. (E) Midplane of cell prepared as in C, demonstrating co-capping of ICAM-1 and moesin. (F) Western blot showing siRNA-mediated knockdown of either moesin (M), α-actinin 1 (A), or both proteins (M/A) in mature BMDCs. (G and H) Mature BMDCs treated with siRNA as in F were surface labeled with Fabs against MHCII or ICAM-1, and FRAP analysis was performed to determine the mobile fraction (G) and diffusion coefficient (H). Dots represent individual FRAP measurements (n = 143–285) pooled from three independent experiments. *, P < 0.01; ***, P < 0.0001. Bars, 10 µm.