Regulatory gene network circuits underlying T cell development from multipotent progenitors

Abstract

Regulatory gene circuits enable stem and progenitor cells to detect and process developmental signals and make irreversible fate commitment decisions. To gain insight into the gene circuits underlying T cell fate decision making in progenitor cells, we generated an updated T-lymphocyte developmental gene regulatory network from genes and connections found in the literature. This reconstruction allowed us to identify candidate regulatory gene circuit elements underlying T cell fate decision making. Here, we examine the roles of these circuits in facilitating different aspects of the decision making process, and discuss experiments to further probe their structure and function.

Figure 1

Stages of T-cell specification and commitment. Double negative (CD4-CD8-DN) developmental stages before T-cell fate commitment (Phase 1), and immediately after commitment (Phase 2) are shown. Straight arrows represent signal-dependent developmental transitions between successive stages, and the curved arrow represents cytokine-dependent proliferation in the Phase 1 stage. Gray dashed lines represent the commitment checkpoint (left), and two TCR-dependent developmental checkpoints (right). [all figures are original]

Figure 2

Regulatory circuit motifs. A) Coherent feedforward loop. B) Negative feedback loop (top). C) Incoherent feedforward loop. D) Positive feedback loop, consisting of a cycle of positive connections (top), or a cycle of repressive connections (bottom). In the former circuit, X and Y correspond to regulators of the same fate; in the latter circuit, X and Z are regulators of alternate fates that are expressed in a mutually exclusive manner (bottom).

Figure 3

Strategies used for inferring regulatory gene connections. A) Natural developmental progression, showing developmental stages of interest. B) In vivo conditional or germline deletion experiments. Upper: ideal in vivo stage-specific gene deletion experiment, where development before the stage of interest is not compromised and compensatory mechanisms do not come into play. Lower: Complication of in vivo germline or conditional deletion experiments, if normal developmental sequence is deranged long before the stages of interest. Gene expression pattern in cell state generated as a result of altered developmental progression (yellow circle) is not strictly comparable with control. C) In vitro T-cell differentiation experiment. Short-term effects of stage-specific environmental signal or gene perturbations (gain or loss of function) remain comparable with controls. Altered cell state (yellow circle), in some cases transformation to a different lineage, can be generated due to prolonged effects of the applied perturbations. Gene expression in such cases may not simply reflect gain or loss of function of the targeted regulator. Dosage effects (not shown) can also yield developmentally inappropriate responses.

Figure 4

Reconstruction of the T-Cell Developmental Gene Regulatory Network. Regulatory network encompasses Notch signaling (blue), IL-7 signaling (dark green), pre-TCR signaling (gray), T-cell regulatory genes (light green), and stem and alternate fate regulatory genes (red). T-cell regulatory genes sharply up-regulated prior to commitment are enclosed in a yellow box. Genes and connections active during the earlier T-cell developmental stages are placed to the left, whereas genes and connections active during later stages are placed to the right. The prevailing connections between the genes in different groups and their signs are summarized in the inset. An online version of this network is available at http://www.its.caltech.edu/~tcellgrn/.

Figure 5

Regulatory Gene Circuits Underlying T-Cell Fate Decision Making. A) Coherent feedforward loops for detection of persistent Notch signaling. Shown here are coherent feedforward loops mediating inputs from Notch signaling to Bcl11b (left), as well as to CD25, pTa and Rag1 (right). B) Incoherent feedforward loops and negative feedback loops for detection of IL-7 signal level changes. Shown here is a negative feedback loop regulating expression of IL-7Ra (top), and an incoherent feedforward loop regulating expression of Tcf7 and Lef1 (bottom). C) Positive feedback loops for T-cell fate commitment and exclusion of alternate fates. Shown here are a series of positive feedback loops involving mutual repression between PU.1 and the T-cell regulators TCF-1, Bcl11b, E2A/HEB and Gfi1. Note that TCF-1, Bcl11b, PU.1, HEBAlt and E2A also receive inputs from Notch and/or IL-7 signaling and may hence mediate engagement of the positive feedback loops in response to upstream signals.