The Effect of DDX41 Mutations on Hematopoiesis During Aging

Myelodysplastic Syndromes (MDS) are a group of bone marrow failure disorders caused by clonal expansion of hematopoietic stem cells (HSC) that fail to produce mature blood cells of sufficient quality and quantity. The disease-causing HSC bear acquired mutations that confer a selective advantage compared to other HSC but are detrimental to hematopoiesis. Typically, the mutations confer increased proliferation and survival upon HSC and their progeny, creating a hypercellular bone marrow with increased proportions of immature myeloid cells. In contrast, mutations in DDX41, an essential RNA helicase, cause reduced proliferation and survival of hematopoietic progenitors and yet contribute to about 4% of MDS cases. DDX41-mutated MDS most often occurs in individuals with inherited heterozygous mutations in the gene, 50-70% of which are truncating (frameshift or loss of translation start) and are thus considered loss-of-function. The other 30-50% of these are missense mutations, whose effect on protein function is largely unknown. The most common acquired mutation occurring in these patients is a second-hit mutation affecting the other allele of DDX41, typically causing the amino acid change R525H. Unique features of DDX41-mutated MDS include a hypocellular bone marrow, few co-mutations, and relatively slower disease progression. Our published and preliminary data indicate that the most common combination of DDX41 mutations observed in patients (truncating/R525H) causes a profound defect in hematopoietic progenitor cell proliferation and survival. Remarkably, our patient sequencing studies demonstrate that HSC bearing biallelic DDX41 mutations clonally expand and dominate the HSC pool, accounting for over 90% of HSCs in 11 out of 11 patients analyzed but only 5-25% of total bone marrow cells. These data and our published mouse models indicate that biallelic DDX41 mutations are favored in HSC but strongly detrimental to progenitor cells. Mechanistically, we found that DDX41 is required for ribosome biogenesis through its function in splicing at small nucleolar RNA (snoRNA) genes. Thus, biallelic DDX41 mutations confer reduced protein synthesis, which is a cause of the progenitor cell viability defect. HSC maintain a lower protein synthesis rate than progenitors, even when cycling, likely for protection from proteotoxic stress, which contributes to aging-associated decline of HSC and other tissue-specific stem cells. We hypothesize that biallelic DDX41 mutations are positively selected in aging HSC pools due to a reduction in proteotoxic stress. In the case of germline missense mutations, this requires dominant negative effects by the acquired R525H mutation to cause reduced protein synthesis and the associated stem cell expansion and progenitor cell defect. To test these hypotheses, we propose to determine the effect of combined germline and acquired missense DDX41 mutations on HSC function through analysis of MDS patient samples and mouse models, and then we propose to determine if proteotoxic stress is the driver of the clonal advantage of biallelic DDX41-mutated HSC in aging bone marrow through analysis of genetically and temporally precise mouse models of the disease. Project Number: 1R01AG093208-01A1 | Fiscal Year: 2026 | NIH Institute/Center: National Institute on Aging (NIA) | Principal Investigator: Timothy Chlon | Institution: CINCINNATI CHILDRENS HOSP MED CTR, CINCINNATI, OH | Award Amount: $645,245 | Activity Code: R01 | Study Section: Hemostasis, Thrombosis, Blood Cells and Transfusion Study Section[HTBT] View on NIH RePORTER: https://reporter.nih.gov/project-details/11292446