Fig. 4.

Effect of YBX1 in MC3T3-E1 cells on ATF4 mRNA stability. (A) RNA pulldown indicated YBX1 protein binding to ATF4 mRNA. (B) RIP demonstrated YBX1 protein binding to ATF4 mRNA. (C) Relative luciferase activity in YBX1 knockdown 293T and MC3T3-E1 cells transfected with ATF4-WT or ATF4-Mut. (D) ATF4 protein expression levels and quantification in shYBX1- or shNC-transfected MC3T3-E1 cells. (E) RT-qPCR evaluated the degradation of ATF4 mRNA after treatment with actinomycin D. Input: positive control. WT: wild type. Mut: mutant. As: antisense strand. n = 3. Data are shown as mean ± SD. * P < 0.05, ** P < 0.01, and *** P < 0.001. YBX1, Y-box binding protein 1; ATF4, activating transcription factor 4; RT-qPCR, quantitative real-time polymerase chain reaction; RIP, RNA immunoprecipitation