RESUMO
The distribution, dynamics, and function of RNA structures in human development are under-explored. Here, we systematically assayed RNA structural dynamics and their relationship with gene expression, translation, and decay during human neurogenesis. We observed that the human ESC transcriptome is globally more structurally accessible than differentiated cells and undergoes extensive RNA structure changes, particularly in the 3' UTR. Additionally, RNA structure changes during differentiation are associated with translation and decay. We observed that RBP and miRNA binding is associated with RNA structural changes during early neuronal differentiation, and splicing is associated during later neuronal differentiation. Furthermore, our analysis suggests that RBPs are major factors in structure remodeling and co-regulate additional RBPs and miRNAs through structure. We demonstrated an example of this by showing that PUM2-induced structure changes on LIN28A enable miR-30 binding. This study deepens our understanding of the widespread and complex role of RNA-based gene regulation during human development.
Assuntos
Redes Reguladoras de Genes , Estudo de Associação Genômica Ampla , Neurogênese , Neurônios/metabolismo , Transcrição Gênica , Regiões 3' não Traduzidas , Diferenciação Celular , Análise por Conglomerados , Técnicas Genéticas , Células HEK293 , Humanos , MicroRNAs/metabolismo , Modelos Estatísticos , Neurônios/fisiologia , Conformação de Ácido Nucleico , RNA/análise , Splicing de RNA , Proteínas de Ligação a RNA/metabolismo , Especificidade por Substrato , Biologia de Sistemas , TranscriptomaRESUMO
Current methods for determining RNA structure with short-read sequencing cannot capture most differences between distinct transcript isoforms. Here we present RNA structure analysis using nanopore sequencing (PORE-cupine), which combines structure probing using chemical modifications with direct long-read RNA sequencing and machine learning to detect secondary structures in cellular RNAs. PORE-cupine also captures global structural features, such as RNA-binding-protein binding sites and reactivity differences at single-nucleotide variants. We show that shared sequences in different transcript isoforms of the same gene can fold into different structures, highlighting the importance of long-read sequencing for obtaining phase information. We also demonstrate that structural differences between transcript isoforms of the same gene lead to differences in translation efficiency. By revealing isoform-specific RNA structure, PORE-cupine will deepen understanding of the role of structures in controlling gene regulation.
