Fig. 6: MACC1 silencing stabilizes KLF4 through the post-transcriptional level. | Cell Death Discovery

Fig. 6: MACC1 silencing stabilizes KLF4 through the post-transcriptional level.

From: MACC1 ablation suppresses the dedifferentiation process of non-CSCs in lung cancer through stabilizing KLF4

Fig. 6

A Schematic diagram of KLF4 promoter region including predicted binding sites for MACC1 was cloned into pGL3-basic plasmid, (B, C) and dual-luciferase assay was performed in H1299 and H2170 cells as described in Materials and Methods. Transcription activity was calculated as the ratio of Firefly luciferase activity (reporter) vs Renilla luciferase activity (control). Comparison was analyzed by two-tailed Student’s t-tests. ns=no significance, Error bars mean ± SD. D, G NCI-H1299 and NCI-H2170 cells were stably knockdown MACC1, actinomycin D (ActD, 5 μg/ml) was added for indicated time points (0, 2, 4 and 6 hours) before harvest. KLF4 mRNA levels were assessed by RT-PCR. MACC1 knockdown efficiencies were determined by RT–PCR analysis. E, H Relative fold of expression was calculated by normalizing mRNA level to that at 0 h (without ActD treatment) for both shNC and shMACC1 groups. Results were representative of three independent experiments. Error bars: mean ± SD. F, I Half-life of mRNA for each group was calculated and compared. The miR-25 activities were reconstituted by mimics. Relative mRNA abundance and protein level of KLF4 were examined by RT-qPCR assay and western blot., **P < 0.01, ***P < 0.001, AVOVA, ns=no significance, Error bars, mean ± SD.

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