Sites of transcription initiation drive mRNA isoform selection [Human_3seq]

The generation of distinct messenger RNA isoforms through alternative RNA processing influences the expression and function of genes, often in a cell-type specific manner. Here, we assess the regulatory relationships between transcription initiation, alternative splicing, and 3' end site selection. Applying multiple long-read-sequencing approaches to obtain an assembly accurately representing even the longest transcripts from end to end, we quantify mRNA isoform choice in Drosophila and human tissues, including the transcriptionally complex nervous system. We find that in Drosophila brains as well as in human cerebral organoids, 3' end site choice is globally influenced by the site of transcription initiation. “Dominant promoters”, characterized by specific epigenetic signatures including p300/CBP binding, impose a transcriptional constraint to define splice and polyadenylation variants. In vivo deletion or overexpression of dominant promoters as well as CBP/p300 loss disrupted the 3' end expression landscape. Our study demonstrates the crucial impact of TSS choice on the regulation of transcript diversity and tissue identity. Overall design: 3'end sequencing of RNA from 60 days-old iPSC-derived cerebral organoids