Search

The differentiation of hematopoietic stem cells into cells of the immune system has been studied extensively in mammals, but the transcriptional circuitry that controls it is still only partially understood. Here, the Immunological Genome Project gene-expression profiles across mouse immune lineages allowed us to systematically analyze these circuits. To analyze this data set we developed Ontogenet, an algorithm for reconstructing lineage-specific regulation from gene-expression profiles across lineages. Using Ontogenet, we found differentiation stage-specific regulators of mouse hematopoiesis and identified many known hematopoietic regulators and 175 previously unknown candidate regulators, as well as their target genes and the cell types in which they act. Among the previously unknown regulators, we emphasize the role of ETV5 in the differentiation of γδ T cells. As the transcriptional programs of human and mouse cells are highly conserved, it is likely that many lessons learned from the mouse model apply to humans.

The differentiation of αβT cells from thymic precursors is a complex process essential for adaptive immunity. Here we exploited the breadth of expression data sets from the Immunological Genome Project to analyze how the differentiation of thymic precursors gives rise to mature T cell transcriptomes. We found that early T cell commitment was driven by unexpectedly gradual changes. In contrast, transit through the CD4(+)CD8(+) stage involved a global shutdown of housekeeping genes that is rare among cells of the immune system and correlated tightly with expression of the transcription factor c-Myc. Selection driven by major histocompatibility complex (MHC) molecules promoted a large-scale transcriptional reactivation. We identified distinct signatures that marked cells destined for positive selection versus apoptotic deletion. Differences in the expression of unexpectedly few genes accompanied commitment to the CD4(+) or CD8(+) lineage, a similarity that carried through to peripheral T cells and their activation, demonstrated by mass cytometry phosphoproteomics. The transcripts newly identified as encoding candidate mediators of key transitions help define the 'known unknowns' of thymocyte differentiation.

Although much progress has been made in the understanding of the ontogeny and function of dendritic cells (DCs), the transcriptional regulation of the lineage commitment and functional specialization of DCs in vivo remains poorly understood. We made a comprehensive comparative analysis of CD8(+), CD103(+), CD11b(+) and plasmacytoid DC subsets, as well as macrophage DC precursors and common DC precursors, across the entire immune system. Here we characterized candidate transcriptional activators involved in the commitment of myeloid progenitor cells to the DC lineage and predicted regulators of DC functional diversity in tissues. We identified a molecular signature that distinguished tissue DCs from macrophages. We also identified a transcriptional program expressed specifically during the steady-state migration of tissue DCs to the draining lymph nodes that may control tolerance to self tissue antigens.

Fibroblastic reticular cells (FRCs) and lymphatic endothelial cells (LECs) are non-hematopoietic stromal cells of lymphoid organs. They influence the migration and homeostasis of naïve T cells; however, their influence on activated T cells remains undescribed. Here we report that FRCs and LECs inhibited T cell proliferation through regulated nitric oxide synthase-2(NOS2)-dependent mechanism. Moreover, in vivo NOS2 expression by FRCs and LECs controlled the size of the activated T cell pool. Mechanistically, the expression of NOS2 and production of nitric oxide (NO) paralleled the activation of T cells and required IFNGR1 signaling in FRCs and LECs, and was augmented by TNF and direct contact with activated T cells. Comparative transcriptomic analysis revealed that various adhesion molecules and multiple components of cytokine, complement and JAK-STAT signaling pathways are upregulated in FRCs in the presence of activated T cells. Based on these data, current efforts aim to identify the molecules involved in the direct stromal cell-T cell interaction influencing NO production by FRCs. This negative regulatory feedback mechanism is likely of great relevance to regulation of immune responses either to prevent excessive T cell expansion or to avoid structural damage within LNs caused by the expanding T cell pool.