Fig. 6

tiRNA-Gly regulates alternative splicing of MAP4K4 mRNA via RBM17. A Alternative splicing induced by tiRNA-Gly is selected in K1 cells transfected with tiRNA-Gly by RNA-seq. 1622 significantly altered alternative splicing are detected, in which exons kipping (cassette) is 37.67%, alternative to 5′ splicing site (A5SS) is 6.78%, alternative to 3′ splicing site (A3SS) is 4.99%, alternative first exon (AltStart) is 18.19%, alternative end exon (AltEnd) is 18.74%, mutually exclusive events (MXE) is 5.12%, and intron retention (IR) is 8.51%. p < 0.05. B Five potential decreased alternative spliced genes were shown. Number means the location of exon. C The structure from exon 14 to exon 18 of long transcript NM_1242560.1 and truncated transcript NM_4834.4 of MAP4K4 mRNA. D Knockdown of RBM17 blocked the increase in the truncated variant (NM_4834.4) of MAP4K4 mRNA induced by tiRNA-Gly. E Expression of MAP4K4 mRNA in K1 cells transfected with two transcripts of MAP4K4 by qRT-PCR. F-G Truncated transcript NM_4834.4 has stronger effect compared with long transcript NM_1242560.1 in K1 cells in CCK-8 assay and migration assay. Scale bar = 100 μm. H Phosphorylation level of proteins in ERK and JNK pathways is higher in K1 cells transfected with truncated transcript NM_4834.4 compared with long transcript NM_1242560.1 by western blot. I The expression of total proteins and phosphorylated proteins involved in ERK and JNK pathways in xenografted cancer. Phosphorylation level of those proteins is reduced in xenografted cancer of si-tiRNA-Gly injected group (S) compared with those in normal saline group (N), while the level of total proteins does not change. J Working model. tiRNA-Gly binds the UHM domain of RBM17 and facilitates its translocation, giving rise to increased RBM17 protein, which induces alternative splicing of target genes in PTC cells. Values are triple replicated and displayed in mean ± SD. **p < 0.01, ***p < 0.001