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Figure 2 | BMC Genomics

Figure 2

From: A gene expression restriction network mediated by sense and antisense Alu sequences located on protein-coding messenger RNAs

Figure 2

Expression levels of Sens-alus and Ant-alus upon Alu perturbations. (A) An overview plot of Sens-alus, Ant-alus and genome-wide RNA levels across 6 different treatment conditions. HEK293 cells were transfected with (1) empty pDR2 vectors (pDR2; Hygromycin added); (2) antisense Alus with Hygromycin selection (pDR2-anti-Alu; Hygromycin added; week 0); (3) the same as (2) with Hygromycin removed subsequently (pDR2-anti-Alu; Hygromycin added→removed; week 1); (4) sense Alus with Hygromycin selection (pDR2-sense-Alu; Hygromycin added; week 0); (5) the same as (4) with Hygromycin removed subsequently (pDR2-sense-Alu; Hygromycin added→removed, week 1); and (6) the original HEK 293 cells (no treatment). Vertical bars, means of expression levels per various gene sets from triplicate experiments. Error bars, standard deviations. (B) GSEA plots for the suppression effects of Ant-alus (upper left) and Sens-alus (upper right) in response to antisense Alu transfections, as well as sense Alu transfections (lower left and right respectively). NES, Normalized enrichment score. (C) The heatmap of a collection of Ant-alu genes which showed significant suppression individually upon sense Alu transfection, using cells transfected by empty vectors as controls (P< 0.0005, FDR < 0.0321). Green color indicated suppression. (D) The heatmap of a collection of Sens-alu genes which showed significant suppression upon antiense Alus transfection (P< 0.0005, FDR < 0.0518). (E) A heatmap representation of protein expressions of randomly selected Ant-alu genes, quantified at different time points by western blotting, up to the 8th week after the removal of Hygromycin. Protein levels of cells transfected by empty vectors (pDR2) were presented as baselines. Numbers in the time axis indicated weeks after Hygromycin removal. The time-course profile of extrachromosomal expression of sense Alu RNA was also presented. (F) Protein levels of randomly selected Sens-alu genes quantified at different time points. The time-course profile of exogeneous antisense Alu RNA was presented.

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