To elucidate the transcriptional landscape that regulates human lymphoid commitment during postnatal life, we used RNA sequencing to assemble the long non-coding transcriptome across human bone marrow and thymic progenitors spanning the earliest stages of B and T lymphoid specification. Over 3000 novel long non-coding RNA genes (lncRNAs) were revealed through the analysis of these rare populations. Lymphoid commitment was characterized by lncRNA expression patterns that were highly stage-specific and more lineage-specific than protein coding patterns. Protein-coding genes co-expressed with neighboring lncRNA genes were enriched for ontologies related to lymphoid differentiation. The exquisite cell-type specificity of global lncRNA expression patterns independently revealed new developmental relationships between the earliest progenitors in the human bone marrow and thymus.
The expression of CD10 has long been used to define human lymphoid commitment. We report a unique lymphoid-primed population in human bone marrow that was generated from hematopoietic stem cells (HSCs) before the onset of CD10 expression and B cell commitment. This subset was identified by high expression of the homing molecule L-selectin (CD62L). CD10−CD62Lhi progenitors possessed full lymphoid and monocytic potential, but lacked erythroid potential. Gene expression profiling placed the CD10−CD62Lhi population at an intermediate stage of differentiation between HSCs and lineage-negative (Lin−) CD34+CD10+ progenitors. L-selectin was expressed on immature thymocytes and its ligands were expressed at the cortico-medullary junction, suggesting a possible role in thymic homing. These studies identify the earliest stage of lymphoid priming in human bone marrow.
Highlights d CD34 + human thymic progenitors present a spectrum of specification and commitment states d Earliest progenitors are CD7and exhibit stem-cell-like and T-primed transcriptomes d Loss of B-potential precedes that of myeloid and NK potentials during T-commitment d A CD34 + subpopulation of cells is primed for the plasmacytoid dendritic lineage
The initial stages of T-cell differentiation are characterized by a progressive commitment to the T-cell lineage, a process that involves the loss of alternative (myelo-erythroid, NK, B) lineage potentials. Aberrant differentiation during these stages can result in T-cell acute lymphoblastic leukemia (T-ALL). However, the mechanisms regulating the initial stages of human T-cell differentiation are obscure. Through loss of function studies we showed BCL11B, a transcription factor recurrently mutated T-ALL, is essential for T-lineage commitment, particularly the repression of NK and myeloid potentials and the induction of T-lineage genes, during the initial stages of human T-cell differentiation. In gain of function studies, BCL11B inhibited growth of and induced a T-lineage transcriptional program in T-ALL cells. We found previously unknown differentiation stage-specific DNA binding of BCL11B at multiple T-lineage genes; target genes showed BCL11B dependent expression, suggesting a transcriptional activator role for BCL11B at these genes. Transcriptional analyses revealed differences in the regulatory actions of BCL11B between human and murine thymopoiesis. Our studies show BCL11B is a key regulator of the initial stages of human T-cell differentiation and delineate the BCL11B transcriptional program, enabling the dissection of the underpinnings of normal T-cell differentiation and providing a resource for understanding dysregulations in T-ALL.
During the last five decades, elegant mouse models of hematopoiesis have yielded most of the seminal insights into this complex biological system of self-renewal and lineage commitment. More recent advances in assays to measure human stem and progenitor cells as well as high resolution RNA profiling have revealed that although the basic roadmap of blood development is generally conserved across mammals, evolutionary pressures have generated many differences between the species that have important biological and translational implications. To enhance the utility of the mouse as a model organism, it is more important than ever that research data are presented with regard to how they might be influenced by the species of origin as well as the developmental source of the hematopoietic tissue.
A system that allows manipulation of the human thymic microenvironment is needed both to elucidate the extrinsic mechanisms that control human thymopoiesis, and to develop potential cell therapies for thymic insufficiency. In this report, we developed an implantable thymic microenvironment composed of two human thymic stroma populations critical for thymopoiesis; thymic epithelial cells (TECs) and thymic mesenchyme (TM). TECs and TM from postnatal human thymi were cultured in specific conditions, allowing cell expansion and manipulation of gene expression, prior to re-aggregation into a functional thymic unit. Human CD34+ hematopoietic stem and progenitor cells (HSPC) differentiated into T cells in the aggregates in vitro and in vivo following inguinal implantation of aggregates in immune deficient mice. Cord blood HSPC previously engrafted into murine bone marrow, migrated to implants and differentiated into human T cells with a broad T cell receptor repertoire. Furthermore, lentiviral-mediated expression of vascular endothelial growth factor in TM enhanced implant size and function, and significantly increased thymocyte production. These results demonstrate an in vivo system for the generation of T cells from human HSPC, and represent the first model to allow manipulation of gene expression and cell composition in the microenvironment of the human thymus.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.