Skip to main content
Fig. 6 | Journal of Translational Medicine

Fig. 6

From: SRSF9 promotes colorectal cancer progression via stabilizing DSN1 mRNA in an m6A-related manner

Fig. 6

Selective binding of SRSF9 to m6A-modified RNAs stabilizes DSN1 mRNA in an m6A-related manner in colorectal cancer. A RNA affinity assay using single-stranded RNA probes containing methylated (red) or unmethylated (green) adenosine. The consensus sequence is shown in bold type. Silver staining (left) and immunoblotting (right) showed selective pull-down of 26 kDa SRSF9 proteins from LOVO cell extracts. IB: Immunoblotting. B Gene-specific m6A qRT-PCR was used to detect enrichment of m6A modifications in DSN1 transcripts (***p < 0.001, Student’s t test). C Bioinformatic analysis of RNA modifications within the SRSF9 protein-binding sites of DSN1 mRNA. D Schematic diagram of the wild-type (WT) and mutant (MUT) firefly luciferase reporters. The A-T substitutions (red) were made within the m6A consensus sequence (green). Only the portion of the SRSF9-DSN1-binding sequence that contains the mutation sites is shown. E Relative luciferase activity of WT and MUT reporters in 293T cells ectopically expressing SRSF9 (****p < 0.0001, Student’s t test). F Relative luciferase activity of the WT reporter in cells cotransfected with the indicated amounts of the SRSF9 expression vectors (***p < 0.001; ****p < 0.0001, Student’s t test). G, H The expression of SRSF9 and METTL3 (G), DSN1 and METTL3 (H) was positively correlated in colon adenocarcinoma tumors in the GEPIA database. I, J Western blot assay showing the protein level of DSN1 in METTL3-deficient CRC cells upon overexpression of SRSF9 in CRC cells. KP The indicated stable cells were subjected to Western blotting (K, M and O). qRT-PCR assays were used to analyze the half-life of DSN1 mRNA in SRSF9 knockdown (L), SRSF9-overexpressing (N) and SRSF9-overexpressing with METTL3 interfered CRC cells after treatment with actinomycin D (normalized to 0 h). Each experiment was performed in triplicate independently

Back to article page