Impaired stem cell differentiation and somatic cell reprogramming in DIDO3 mutants with altered RNA processing and increased R-loop levels

• DOI: 10.1038/s41419-021-03906-2
• Publication Date: 2021-06-21T15:03:51Z
• Authors: Agnes Fütterer; Amaia Talavera-Gu...; Tirso Pons; Jesús de Celis; Julio Gutiérrez; Verónica Domíngue...; Carlos Martínez-A
• Keywords: RNA Splicing; Mice, Transgenic; Article; DEAD-box RNA Helicases; Mice; 03 medical and health sciences; Animals; Cellular Reprogramming Techniques; Cells, Cultured; 0303 health sciences; QH573-671; Gene Expression Regulation, Developmental; Cell Differentiation; Mouse Embryonic Stem Cells; Fibroblasts; Cellular Reprogramming; DNA-Binding Proteins; Mice, Inbred C57BL; Phenotype; Transcription Termination, Genetic; Mutation; R-Loop Structures; Cytology; DNA Damage; Transcription Factors

AbstractEmbryonic stem cell (ESC) differentiation and somatic cell reprogramming are biological processes governed by antagonistic expression or repression of a largely common set of genes. Accurate regulation of gene expression is thus essential for both processes, and alterations in RNA processing are predicted to negatively affect both. We show that truncation of the DIDO gene alters RNA splicing and transcription termination in ESC and mouse embryo fibroblasts (MEF), which affects genes involved in both differentiation and reprogramming. We combined transcriptomic, protein interaction, and cellular studies to identify the underlying molecular mechanism. We found that DIDO3 interacts with the helicase DHX9, which is involved in R-loop processing and transcription termination, and that DIDO3-exon16 deletion increases nuclear R-loop content and causes DNA replication stress. Overall, these defects result in failure of ESC to differentiate and of MEF to be reprogrammed. MEF immortalization restored impaired reprogramming capacity. We conclude that DIDO3 has essential functions in ESC differentiation and somatic cell reprogramming by supporting accurate RNA metabolism, with its exon16-encoded domain playing the main role.