Stimulation of translation by RHA depends on the presence of collagen 5′ SL. (A) Schematic representation of the collagen/luciferase reporter genes. (Dotted line) Promoters; (black line) collagen 5′ UTR; (gray line) luciferase sequence. The transcription start site is marked by an arrow, and positions of the start and stop codons are indicated. (B) Immunoblot showing knockdown of RHA by siRNA. (C) Knockdown of RHA affects protein expression from the mRNA with 5′ SL. RHA-specific siRNA (RHA, black bars) and control siRNA (Sc, open bars) were transfected into HEK293 cells, followed by transfection of the reporter genes. (The two stars represent the significance p < 0.01.) Expression of luciferase protein was measured as the enzymatic activity and was normalized to the total protein in the extract. Luciferase mRNA was measured by real-time RT-PCR and was normalized to GAPDH mRNA. The ratio of luciferase protein to mRNA was plotted on the y-axis. The results of five independent experiments with error bars of ±1 SEM and the P-values are shown.
