Fig. 5: H19 might function by sponging let-7b targeting Casp3. | Cell Death & Disease

Fig. 5: H19 might function by sponging let-7b targeting Casp3.

From: Long non-coding RNA H19 contributes to apoptosis of hippocampal neurons by inhibiting let-7b in a rat model of temporal lobe epilepsy

Fig. 5

a Bioinformatics-predicted binding sites for eight let-7 subtypes (let-7a, let-7b, let-7c, let-7d, let-7e, let-7f, let-7g, and let-7i) in rat H19. Let-7-binding sites on H19 were predicted using a web-based program RNAhybrid (http://bibiserv.techfak.uni-bielefeld.de/rnahybrid/). Nucleotides of the miRNA let-7 seed region (positions 2–8) are marked in red. b The expression levels of let-7 family members (let-7a, let-7b, let-7d, let-7e, let-7g, and let-7i) in CA3 subfield of hippocampus from sham-operated and KA-induced epileptic rats at 1 day after surgery as determined using qPCR (n = 7–8). c, d qPCR quantification of H19 and let-7b levels in CA3 subfield of hippocampus from H19 overexpression (c) and H19 knockdown (d) rats with or without KA treatment for 7 days (n = 8). e Correlations according to Pearson coefficient between H19 and let-7b levels in surgically resected hippocampal samples from patients with TLE (n = 12). f Top, complementarity between let-7b seed sequence and the 3′-UTR position of Casp3 of rat predicted by a computational and bioinformatics-based approach using TargetScan. Mutant Casp3 (Casp3-Mut) was generated by mutating the putative let-7b-binding site (underlined). Watson–Crick complementarity was connected by “|”. Bottom, luciferase reporter gene assay for interactions between let-7b and its binding sites or mutated binding sites in the 3′-UTR of the Casp3 mRNA in HEK293 cells. The relative luciferase activity was normalized against the firefly luciferase values shown as mean ± s.e.m (n = 3). g, h qPCR quantification of let-7b and Casp3 mRNA levels in CA3 subfield of hippocampus from g H19 overexpression (n = 8) and h H19 knockdown (n = 4–5) rats with or without KA treatment for 7 days. The relative H19 levels were normalized to GAPDH and the relative expression levels of let-7 family members were normalized to U6. All data are shown as mean ± s.e.m. *p < 0.05, **p < 0.01, ***p < 0.001 versus sham groups (sham, NC+Veh, or Scr+Veh). #p < 0.05, ##p < 0.01, ###p < 0.001 versus NC+KA or Scr+KA group

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