Characterizing DNA Methylation Dynamics in Trisomy 21-Associated Hematopoiesis and Leukemia - PROJECT SUMMARY DNA methylation (5mC) is a critical epigenetic mechanism in hematopoiesis. Down syndrome (DS), or trisomy 21 (T21), predisposes individuals to unique conditions of hematopoietic malignancies, such as the preleukemic condition Transient Abnormal Myelopoiesis (TAM) and Myeloid Leukemia associated with Down Syndrome (ML- DS). To enhance our understanding of the role of 5mC in these conditions, our project aims to map 5mC dynamics in conjunction with chromatin accessibility and RNA expression at single-cell resolution across these critical stages of DS hematopoiesis. Aim 1 focuses on the fetal stage, employing our innovative single-cell multi- omic sequencing technology, SHARE-ME-seq, to explore highly enriched hematopoietic stem cells (HSCs) in T21 fetal liver. By capturing over 1 million single-cell profiles, this aim will profile samples from ten donors between 16-23 weeks of gestation, allowing us to capture the epigenetic landscape during the crucial period of HSC expansion. The data generated will enable us to establish a detailed molecular timeline, revealing how 5mC influences HSC self-renewal and predisposition towards preleukemia. Aim 2 extends this study to postnatal stages, where we will examine primary patient samples from TAM and ML-DS stages. Here, we will apply the same SHARE-ME-seq technology to discern how specific 5mC patterns evolve from the preleukemic TAM stage to full-blown ML-DS. By integrating concurrent analyses of chromatin accessibility and gene expression, this aim will characterize the role of epigenetic modifications in driving the progression of DS-associated leukemia. Additionally, histone modification profiling through CUT&Tag will be conducted to identify active, repressed, or poised enhancers and promoters, providing deeper insights into the chromatin dynamics and lineage priming that contribute to leukemogenesis in DS. These comprehensive and innovative approaches are designed to uncover the interactions between DNA methylation, chromatin accessibility, and gene expression in DS- associated hematopoiesis. By elucidating these mechanisms, our study aims to identify new therapeutic targets and improve intervention strategies for hematologic malignancies in DS, ultimately enhancing the prognosis and quality of life for individuals affected by this condition. The data generated will serve as a high-quality reference, informing future research and therapeutic development tailored to DS individuals.
