Uncovering the dynamics and consequences of RNA isoform changes during neuronal differentiation

Static gene expression programs have been extensively characterized in stem cells and mature human cells. However, the dynamics of RNA isoform changes upon cell-state-transitions during cell differentiation, the determinants and functional consequences have largely remained unclear. Here, we established an improved model for human neurogenesis in vitro that is amenable for systems-wide analyses of gene expression. Our multi-omics analysis reveals that the pronounced alterations in cell morphology correlate strongly with widespread changes in RNA isoform expression. Our approach identifies thousands of new RNA isoforms that are expressed at distinct differentiation stages. RNA isoforms mainly arise from exon skipping and the alternative usage of transcription start and polyadenylation sites during human neurogenesis. The transcript isoform changes can remodel the identity and functions of protein isoforms. Finally, our study identifies a set of RNA binding proteins as a potential determinant of differentiation stage-specific global isoform changes. This work supports the view of regulated isoform changes that underlie state-transitions during neurogenesis. Synopsis Multi-omics analysis of a newly established human neuronal cell differentiation model reveals widespread dynamic changes in RNA isoform expression, their functional consequences and potential determinants, providing evidence that they underlie cell-state-transitions during neurogenesis. Dynamic changes in RNA and protein levels are strongly correlated during all stages of neuronal differentiation. Nanopore sequencing (ONT-seq) during human neurogenesis reveals 12,019 non-annotated RNA isoforms, a large number of which are differentially expressed during differentiation. 70% of new RNA isoforms result from the use of alternative transcription start sites (TSSs) or polyadenylation (pA) sites and exon skipping. RNA isoform changes underlie protein isoform changes during human neurogenesis as revealed by integrating ONT-seq, RNA-seq and proteomics time course data. RNA motif enrichment, RNA-seq and available CLIP-seq data uncover a set of RNA binding proteins (RBPs) as potential determinants of differentiation stage-specific global isoform changes. Multi-omics analysis of a newly established human neuronal cell differentiation model reveals widespread dynamic changes in RNA isoform expression, their functional consequences and potential determinants, providing evidence that they underlie cell-state-t