Figure S3.
Ezrin is the only ERM family protein whose expression is reduced by RANKL treatment, and its expression is regulated at the transcriptional level. (A) Western blot analysis of cell lysates without or treated with RANKL for 60 h using anti-ezrin, anti-radixin, anti-moesin, and anti–β-actin antibodies. (B–D) Quantification of endogenous ezrin (B), radixin (C), and moesin (D) based on (A). Mean ± SEM, n = 6 experiments. P value obtained from two-tailed Student’s t test. ns: not significant; ****P < 0.0001. (E) Western blot analysis of cell lysates without or treated with RANKL and the proteasome inhibitor MG132 for 84 h using anti-ezrin and anti–β-actin antibodies. (F) Quantification of endogenous ezrin based on E. The values of each band were measured and normalized to cells not treated with RANKL and MG132. Mean ± SEM, n = 6 experiments. P value obtained from one-way ANOVA with Tukey’s test. ns: not significant; *P < 0.05; **P < 0.01. (G–J) qPCR analysis of the relative expression of Nfatc1 (G), Ezr (H), Rdx (I), and Msn (J). mRNA was extracted after 60 h of RANKL treatment and normalized to the expression of Gapdh mRNA. Mean ± SEM, n = 4 experiments. P value obtained from two-tailed Student’s t test. ns: not significant; *P < 0.05; ***P < 0.001. (K) Western blot analysis of Blimp1 knockdown cell lysates without or treated with RANKL for 84 h using anti-ezrin and anti–β-actin antibodies. (L) Quantification of endogenous ezrin based on K. The values of each band were measured and normalized to cells not treated with RANKL of si-Control group. Mean ± SEM, n = 7 experiments. P value obtained from one-way ANOVA with Tukey’s test. ns: not significant; *P < 0.05; ****P < 0.0001. (M) Representative western blot of siRNA knockdown cells with RANKL treatments for 70 h, using anti-ezrin, anti-radixin, anti-moesin, and anti–β-actin antibodies. (N–P) qPCR analysis of the relative expression of Fnbp1 (N) and Baiap2l1 (72 h, O; 84 h, P) mRNAs, normalized by that of Gapdh mRNA. Source data are available for this figure: SourceData FS3.

Ezrin is the only ERM family protein whose expression is reduced by RANKL treatment, and its expression is regulated at the transcriptional level. (A) Western blot analysis of cell lysates without or treated with RANKL for 60 h using anti-ezrin, anti-radixin, anti-moesin, and anti–β-actin antibodies. (B–D) Quantification of endogenous ezrin (B), radixin (C), and moesin (D) based on (A). Mean ± SEM, n = 6 experiments. P value obtained from two-tailed Student’s t test. ns: not significant; ****P < 0.0001. (E) Western blot analysis of cell lysates without or treated with RANKL and the proteasome inhibitor MG132 for 84 h using anti-ezrin and anti–β-actin antibodies. (F) Quantification of endogenous ezrin based on E. The values of each band were measured and normalized to cells not treated with RANKL and MG132. Mean ± SEM, n = 6 experiments. P value obtained from one-way ANOVA with Tukey’s test. ns: not significant; *P < 0.05; **P < 0.01. (G–J) qPCR analysis of the relative expression of Nfatc1 (G), Ezr (H), Rdx (I), and Msn (J). mRNA was extracted after 60 h of RANKL treatment and normalized to the expression of Gapdh mRNA. Mean ± SEM, n = 4 experiments. P value obtained from two-tailed Student’s t test. ns: not significant; *P < 0.05; ***P < 0.001. (K) Western blot analysis of Blimp1 knockdown cell lysates without or treated with RANKL for 84 h using anti-ezrin and anti–β-actin antibodies. (L) Quantification of endogenous ezrin based on K. The values of each band were measured and normalized to cells not treated with RANKL of si-Control group. Mean ± SEM, n = 7 experiments. P value obtained from one-way ANOVA with Tukey’s test. ns: not significant; *P < 0.05; ****P < 0.0001. (M) Representative western blot of siRNA knockdown cells with RANKL treatments for 70 h, using anti-ezrin, anti-radixin, anti-moesin, and anti–β-actin antibodies. (N–P) qPCR analysis of the relative expression of Fnbp1 (N) and Baiap2l1 (72 h, O; 84 h, P) mRNAs, normalized by that of Gapdh mRNA. Source data are available for this figure: SourceData FS3.

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