Genotype-integrated single-cell transcriptome analysis of a myelodysplastic neoplasm case reveals the role of DDX41 p.R525H

DEAD-box helicase 41 ( DDX41 ) is a key gene in germline-predisposed myeloid neoplasms, where pathogenic germline variants often lead to disease after acquiring a somatic mutation in trans. We conducted single-cell RNA sequencing combined with genotyping of the p.R525 site in a myelodysplastic neoplasm (MDS) case with germline p.R339C and somatic p.R525H variants, which enabled us to compare p.R525H cells with germline variant-only cells. p.R525H acquisition selectively induced G2/M arrest in colony-forming unit-erythroid cells along with R-loop elevation, thus impairing erythropoiesis via DNA damage accumulation. In hematopoietic stem cell (HSC) and myeloid progenitor populations, gene expression profiles were largely similar between p.R525H-positive and -negative cells; however, HSC frequencies were lower and granulocyte-monocyte progenitor frequencies were higher in p.R525H cells. Ligand-receptor and transcriptional regulation analyses suggested non-cell-autonomous influence from p.R525H cells on germline variant-only cells, driving convergence toward a shared expression profile and revealing an intricate interplay that shapes the MDS phenotype.