Single-cell multi-omic analysis of thymocyte development reveals drivers of CD4/CD8 lineage commitment

Abstract

CD4 and CD8 T cells play critical roles in the mammalian immune system. While their development within the thymus from the CD4+CD8+ stage has been widely studied as a model of lineage commitment, the underlying mechanism remains unclear. To deconstruct this process, we apply CITE-seq, measuring the transcriptome and over 100 surface proteins in thymocytes from wild-type and lineage-restricted mice. We jointly analyze the paired measurements to build a comprehensive timeline of RNA and protein expression in each lineage, supporting a sequential model of lineage determination in which both lineages go through an initial phase of CD4 lineage audition, which is followed by divergence and specification of CD8 lineage cells. We identify early differences implicating T cell receptor signaling via calcineurin-NFAT in driving CD4 lineage commitment. Pharmacological inhibition validates the requirement of calcineurin- NFAT for CD4, but not CD8, lineage development, providing insight into the CD4/CD8 commitment mechanism.

A spatial human thymus cell atlas mapped to a continuous tissue axis

“…These differences were seen in both Visium and IBEX data, demonstrating how multimodal data sets can be employed to derive cross-validated interpretations regarding a cell’s maturation state, its local environment, and its position in a tissue. Our findings directly align with two recent studies that suggest a slower maturation of CD8 lineage cells 83,85 . Moreover, our observation that co-receptor reversal in CD8 lineage cells coincides with the cortical retention window further indicates that CD8SP thymocytes may require more time to initiate the lineage-specific differentiation programme, which in turn could impact the timing of intercompartmental cell migration.…”

“…This CR was upregulated in early phase 3 for CD4 lineage cells but only in late phase 3 for the CD8 lineage, coinciding with the main medullary migration window of each lineage ( Figure 6F ). This delay in CCR7/ CD197 expression during CD8 differentiation matches observations in a recent mouse CITE-seq study 85 and points to a potential role of this receptor in the timing of the cortico-medullary transition.…”

SEC-seq: Association of molecular signatures with antibody secretion in thousands of single human plasma cells

“…A parallel study is also exploring the transcriptome that underlies secretion of high levels of vascular endothelial growth factor by mesenchymal stromal cells (unpublished work), which can elucidate features associated with therapeutic sub-populations of cells. We demonstrate here that the technique is compatible with standard CITE-seq 18 reagents to create workflows that link surface proteins, secreted proteins, and single-cell transcriptomes 19 . More broadly, nanovials can link surface markers and secreted proteins in viable cells selected using multicolor FACS.…”

Mapping the temporal transcriptional landscape of human white and brown adipogenesis using single-nuclei RNA-seq

“…Next, dynamically regulated genes were identified by binning nuclei in the pseudotemporal space (Fig. S2A and S2B, see Methods, Steier et al, 2021). We reasoned that there would be significant deviation in expression of dynamically regulated genes from either the initial or final stages of adipogenesis, and hence performed differential expression testing for each bin against the first and last pseudotemporal bins (logFC >1 and FDR < 0.05) to identify such genes.…”