RBM10 regulates AS of ECM and cytoskeletal transcripts. (A) Western blot of RBM10 in isogenic human thyroid cancer cell lines. (B) Experimental strategy to identify molecular mechanisms of thyroid cancer growth and metastases in RBM10-deficient cells; illustration by Biorender. (C) Average changes in aberrant splicing events in RBM10 deficient detected by high-depth RNAseq in the RBM10 isogenic cell lines; SE—cassette exon inclusion; RI—retained intron; A3SS and A5SS—most intron-proximal isoform for competing 3′ or 5′ splice sites. (D) GO using ranked differentially spliced genes by GORILLA showing enrichment in pathways involved in cell adhesion and cytoskeleton reorganization. (E) Volcano plot showing differentially expressed alternative spliced isoforms detected by Partek Flow RNAseq alt-splicing analysis in the combined isogenic PE121410 and KTC1 cells. Arrows point to representative isoforms of the indicated genes. Black arrows: ECM/cytoskeletal modifying transcripts. Blue arrows: known RBM10 AS genes. (F) RT-PCR showing indicated exon inclusion isoforms of VCL, CD44, and TNC in PE121410 and KTC1 cells and their decrease with RBM10 expression. Constitutive isoforms and GAPDH were used as controls. Red—cassette exon; green—constitutive exon; and dotted arrow—primer placement. (G) Relative ratio of the indicated inclusion isoforms of VCL, CD44, and TNC determined by qRT-PCR in PE121410 and KTC1 cells ± RBM10. The data represent mean with SEM of atleast three replicates; statistical difference was analyzed using unpaired t test and is indicated as **P < 0.01 and ***P < 0.0001. (H) Western blot of VCL and β-actin in the indicated cells. Arrow points to VCL and MVCL. A3SS: 3′ splice sites and A5SS: 5′ splice sites. Source data are available for this figure: SourceData F2.