Leveraging quantitative assessments of mitochondrial DNA sequence variation for inferences of clonal dynamics of human hematopoiesis

• Applicant: Dr. Leif Ludwig
• Subject Area: Hematology, Oncology ; General Genetics and Functional Genome Biology
• Term: since 2020
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 438931857

Hematopoiesis has served as a paradigm for understanding stem cell biology, providing insights into stem cell maintenance and differentiation, the occurrence of cell fate decisions and how such processes may be pathologically deregulated. While advances in single cell genomics technologies and functional assays have further redefined concepts about hematopoietic development and differentiation, we only have a modest understanding of in vivo hematopoietic cell population dynamics in humans. While natural somatic mutations provide a means to assess the output of individual cells, current approaches remain challenging and costly, leaving open fundamental questions about hematopoietic stem cell frequency, functionality, longevity, dynamics and underlying regulatory mechanisms. Recently, we have developed novel conceptual and technical approaches enabling high-throughput single cell-based detection of mitochondrial DNA (mtDNA) sequence variation for inferences about the clonal composition of complex populations, further providing concomitant information about cellular states (e.g. accessible chromatin or transcriptome) of the same single cells. In this regard, this proposal has two complementing major aims. First, a fundamental objective of this proposal is to leverage somatic mtDNA mutations as natural clonal markers of individual hematopoietic stem and progenitor cells (HSPCs) to study their dynamic contributions in vivo. Specifically, we will study the process of hematopoietic reconstitution following allogeneic stem cell transplantation as a curative approach to treat patients with leukemia. In this setting, we will longitudinally probe HSPC clonal contributions at different phases of reconstitution and assess how cellular states may regulate such activity or cell-type specific biases. Simultaneously, our approach enables sensitive “single-cell level” detection of potentially remaining/ recurrent or newly emerging leukemic clones, providing a complementary means to monitor and study epigenetic and leukemic evolution across stages of disease progression and relapse. Second, we will concern ourselves with systematically assessing the rate of evolution of somatic mtDNA mutations that have further been intimately tied to the process of age-related hematopoietic dysfunction. Specifically, we will apply single cell genomic and cell biological approaches to link individual cell genotype and epigenetic assessments to phenotypic deficiencies in the context of dysfunctional hematopoiesis. Together, with a more sophisticated knowledge about these properties, we expect not only to obtain a significantly improved understanding of the cellular dynamics underlying homeostatic and diseased hematopoiesis, but will conceivably also be able to better predict and direct HSPC behavior to facilitate their extended maintenance/ expansion for basic research purposes as well as the development of targeted interventions, such as for clinical cellular replacement therapies.

• DFG Programme: Independent Junior Research Groups