Quantitative association between gene expression and blood cell production of individual hematopoietic stem cells in mice.

Individual hematopoietic stem cells (HSCs) produce different amounts of blood cells upon transplantation. Taking advantage of the intercellular variation, we developed an experimental and bioinformatic approach to evaluating the quantitative association between gene expression and blood cell production across individual HSCs. We found that most genes associated with blood production exhibit the association only at some levels of blood production. By mapping gene expression with blood production, we identified four distinct patterns of their quantitative association. Some genes consistently correlate with blood production over a range of levels or across all levels, and these genes are found to regulate lymphoid but not myeloid production. Other genes exhibit one or more clear peaks of association. Genes with overlapping peaks are found to be coexpressed in other tissues and share similar molecular functions and regulatory motifs. By dissecting intercellular variations, our findings revealed four quantitative association patterns that reflect distinct dose-response molecular mechanisms modulating the blood cell production of HSCs.

RNA splicing factor Rbm25 underlies heterogeneous preleukemic clonal expansion in mice.

Clonal expansion sets the stage for cancer genesis by allowing for the accumulation of molecular alterations. Although genetic mutations such as Tet2 that induce clonal expansion and malignancy have been identified, these mutations are also frequently found in healthy individuals. Here, we tracked preleukemic clonal expansion using genetic barcoding in an inducible Tet2 knockout mouse model and found that only a small fraction of hematopoietic stem cells (HSCs) expanded excessively upon Tet2 knockout. These overexpanded HSCs expressed significantly lower levels of genes associated with leukemia and RNA splicing than nonoverexpanded Tet2 knockout HSCs. Knocking down Rbm25, an identified RNA splicing factor, accelerated the expansion of Tet2-knockout hematopoietic cells in vitro and in vivo. Our data suggest that mutations of an epigenetic factor Tet2 induce variability in the expression of an RNA splicing factor Rbm25, which subsequently drives heterogeneous preleukemic clonal expansion. This heterogeneous clonal expansion could contribute to the variable disease risks across individuals.

Lineage tracking to reveal the fate of hematopoietic stem cells influenced by Flk2- multipotent progenitors after transplantation.

After transplantation, hematopoietic stem cells (HSCs) sustain blood cell regeneration throughout the patient's life. Recent studies suggest that several types of mature blood cells provide feedback signals to regulate HSC fate. However, the potential feedback effect of hematopoietic progenitor cells has not been characterized to date. The present investigation demonstrated that multipotent progenitors (MPPs) promoted T cell production of HSCs when both cell types were cotransplanted in mice. Using genetic barcodes to track individual HSCs in mice, we found that the increased T cell production by HSCs was associated with the combined effects of altered lineage bias and clonal expansion during HSC differentiation. We showed that MPP and HSC co-transplantation promoted the multilineage differentiation of HSCs in the short term while preserving lymphoid-specialized HSC differentiation in the long term. Our findings indicate that MPPs derived from HSCs regulate the fate of HSCs after bone marrow transplantation.

Deciphering intratumoral heterogeneity using integrated clonal tracking and single-cell transcriptome analyses.

Cellular heterogeneity is a major cause of treatment resistance in cancer. Despite recent advances in single-cell genomic and transcriptomic sequencing, it remains difficult to relate measured molecular profiles to the cellular activities underlying cancer. Here, we present an integrated experimental system that connects single cell gene expression to heterogeneous cancer cell growth, metastasis, and treatment response. Our system integrates single cell transcriptome profiling with DNA barcode based clonal tracking in patient-derived xenograft models. We show that leukemia cells exhibiting unique gene expression respond to different chemotherapies in distinct but consistent manners across multiple mice. In addition, we uncover a form of leukemia expansion that is spatially confined to the bone marrow of single anatomical sites and driven by cells with distinct gene expression. Our integrated experimental system can interrogate the molecular and cellular basis of the intratumoral heterogeneity underlying disease progression and treatment resistance.