Cooking up T cells

Comment on La Motte-Mohs et al, page 1431, and Lehar et al, page 1440

The Notch ligands Delta-1 and Jagged-1 differentially induce T-cell development in culture.

Two articles in this issue address how Notch ligands induce T-cell development. Mammalian Notch ligands comprise 2 families, serrate-like (Jagged1, Jagged2) and Delta-like (Dll1, 3, and 4). Both families encode transmembrane proteins whose extracellular domain contains epidermal growth factor (EGF)–like repeats and an N-terminal DSL domain (for Delta, Serrate, Lag2) that binds Notch receptors and short, poorly conserved intracellular domains that have important but poorly understood functions. Notch receptor binding by ligand initiates a series of proteolytic cleavages in the receptor, causing release of the intracellular domain of Notch from the plasma membrane and translocation into the nucleus, where it creates a short-lived complex that activates transcription.

Specificity of Notch ligand–receptor interactions is poorly understood. In some assays, such as inhibition of myocyte development, Delta and Jagged function equivalently. In vivo, Delta and Jagged often show overlapping expression, providing limited insight into specificity. Knockout studies demonstrate that Notch ligands are not equivalent, at least for marginal zone B-cell development. These results show that Dll1 is the important partner for Notch2 in marginal zone B-cell development, as conditional deletion of either leads to loss of these B cells.¹ 

The precise Notch ligands in T-cell development are less certain. Notch1 signals are uniquely required among the 4 receptors for both T-cell commitment from a multipotent progenitor and proper development to the double-positive (DP) stage.²  Conditional deletion of either Jagged1 or Dll1 leaves T-cell development unaffected, suggesting that neither uniquely signals T-cell development and that conditional deletion of multiple family members will be necessary.^1,3  A major breakthrough in identifying Notch ligands relevant to T-cell lymphopoiesis was provided by establishing stromal cell cultures that recapitulate many aspects of T-cell development.⁴  Using either OP9 or S17 stromal cells engineered to express Dll1, Schmitt and Zuniga-Pflucker⁵  and Jaleco et al⁶  succeeded in establishing a cell culture–based assay that recapitulates T-cell development, a process previously thought to require intact thymic organ cultures. In the absence of Delta-induced Notch signals, these cultures generate B cells. OP9-Dl1 cells are particularly useful as they efficiently generate DP and even mature single-positive CD8 cells from murine hematopoietic progenitors derived from embryonic stem cells, fetal liver, and adult bone marrow. In addition, these cultures have provided important mechanistic insights into murine T-cell development. Jaleco et al⁶  also showed that CD34⁺ human cord blood cells were capable, albeit inefficiently, of generating DP T cells on S17-Dl1 cells. In this issue, La Motte-Mohs and colleagues extend the previous studies by showing that culture of human cord blood on OP9-Dl1 cells leads to efficient T-cell commitment, expansion, and generation of T-cell receptor αβ (TCRαβ)–expressing cells. Whether the latter are functionally mature remains to be determined. Similar results were obtained using human adult bone marrow progenitors.⁷  Thus, it should now be possible to obtain a detailed understanding of early human T-cell development and use this information to manipulate and expand T-cell progenitors for therapeutic purposes.

The ability of OP9-Dl1 cells to induce T-cell development raises the question of whether Jagged ligands have the equivalent ability. In this issue, Lehar and colleagues engineered OP9 cells to express increased levels of Jagged1 (OP9-Jag1). As Jagged1, unlike Dll1, is expressed on parental OP9 cells, this appears to be a quantitative rather than qualitative change. In contrast to Dll1, OP9-Jag1 cells failed to induce T-cell development and induced only weak expansion of committed T-cell progenitors. Nevertheless, Jagged had some effects, as thymic DN1 cells, a heterogeneous population, were unable to form B cells when cultured on OP9-Jag1, as opposed to OP9 control cells. This study and previous work from Jaleco et al⁶  demonstrate that Jagged and Delta signals are not equivalent in the context of T-cell development. Whether these differences are quantitative or qualitative is unknown. For example, Fringe-induced Notch modification could lead to Jagged insensitivity. Alternatively, ligand density may play a role, a parameter that has not yet been assessed.

Although much remains to be learned about the mechanism of Notch-induced T-cell development, the ability to efficiently generate human T cells in culture holds great therapeutic promise, such as in improving T-cell generation after myeloablative therapy. The culture systems described to date are an excellent start and understanding the functions of the ligands in producing specific signals will be important. This hope must be tempered with caution, as it is not yet known whether the T cells derived from these cultures will have an increased propensity to cause autoimmune disease or leukemia. Nevertheless, the advent of these culture systems should allow rapid progress in addressing these issues and translating these findings to the clinic. ▪