Research Paper Volume 12, Issue 24 pp 25528—25546

miRNA-122-5p stimulates the proliferation and DNA synthesis and inhibits the early apoptosis of human spermatogonial stem cells by targeting CBL and competing with lncRNA CASC7

Figure 3. Identification and verification of the target CBL of MiRNA-122-5p in human SSC line. (A–E) Real-time qPCR revealed the relative levels of ZNF668 (A), PLAG1 (B), MEF2D (C), FOXP1 (D) and CBL (E) in human SSCs of miRNA-122-5p vector and control lentivirus. * indicated statistically significant differences (p< 0.05) in human SSC line between control lentivirus and miRNA-122-5p vector. (F) Western blots depicted CBL protein in human SSC line of miRNA-122-5p vector and control lentivirus. (G) The relative expression levels of CBL protein in human SSC line of miRNA-122-5p vector and control lentivirus after normalization to the signals of their loading control ACTB. * denoted statistically significant differences (p< 0.05) in human SSC line between control lentivirus and miRNA-122-5p vector. (H, I) Validation of the targeting of miRNA-122-5p to wild type CBL and mutated CBL by dual luciferase reporter assays. (J, K) Validation of the targeting of miRNA-122-5p to wild type ZNF668 and mutated ZNF668 by dual luciferase reporter assays. * implied statistically significant differences (p< 0.05) in human SSC line between miRNA-122-5p mimics and miRNA mimics control. (L) Schematic diagram illustrated the binding site of miRNA-122-5p to CBL mRNA.