Fig 1: Intronic pPA events occur following the large internal exons of additional TSGs and correlate with reduced large internal exon m6A levels. (A) Diagrams showing the exon/intron arrangement of the full-length LATS1 gene and a truncated variant. The large internal exon is colored blue. (B) Detection of full-length LATS1 mRNA isoforms (lengths vary depending on 3' UTR PAS selection) as well as a truncated LATS1pPA mRNA isoform corresponding to intronic pPA downstream of exon 4 in the MDA-MB-231 and MCF10A cell lines by 3' RACE. Approximate molecular mass markers are indicated in kb. (C) Immunoblot of LATS1 full-length and pPA-truncated products in the indicated cell lines. The membrane from Fig. 1D was stripped and re-probed with an anti-LATS1 antibody. Immunoblot of ß-actin is included to show loading, approximate molecular mass markers are indicated in kDa, and the relative levels of full-length and pPA-truncated LATS1 proteins were normalized to ß-actin levels. (D) Domains and functional regions of the encoded LATS1 full-length and pPA-truncated proteins. (E) Diagrams showing the exon/intron arrangement of the full-length BRCA1 gene and a truncated variant. The large internal exon is colored blue. (F) Immunoblots of BRCA1-p220 and BRCA1-IRIS proteins in the indicated cell lines. Immunoblot of ß-actin is included to show loading, approximate molecular mass markers are indicated in kDa, and the relative levels of full-length and pPA-truncated BRCA1 proteins were normalized to ß-actin levels. (G) Domains and functional regions of the encoded gene products, BRCA1-p220 and BRCA1-IRIS. (H) m6A levels at the m6A consensus sites of LATS1, relative to a distal LATS1 exonic segment (exons 2–3), as determined by m6A RIP-qPCR in MCF10A cells (n = 3 m6A RIP replicates). (I) m6A levels at the m6A consensus sites of BRCA1, relative to a distal BRCA1 exonic segment (exons 2–3), as determined by m6A RIP-qPCR in MCF10A cells (n = 3 m6A RIP replicates). (J–M) Relative m6A levels at the indicated m6A consensus sites of LATS1 (J,L) and BRCA1 (K,M) large internal exons, as determined by m6A RIP-qPCR in the indicated cell lines (n = 3 m6A RIP replicates). Data in (H–J) are presented as mean ± SEM. **p = 0.01, ***p = 0.001 (two-tailed Student’s t-tests).