Research

Background and Rationale

Hematopoietic Stem Cells (HSCs) can give rise to a large variety of cells.

We are interested in the process by which HSCs differentiate into T cells. This process occurs after uncommitted blood precursor cells enter the thymus, and requires environmental signals provided by the thymus.  Fate selection may take up to 10 cell cycles to be completed, and occurs through steps in which at least four developmental stages can be distinguished by cell-surface phenotype. By convention, cells at these stages are called DN1 – DN4. Cells enter the thymus as multipotent progenitors but become progressively narrowed in their developmental potential, reaching commitment to the T cell lineage before the DN3 stage. Only afterwards do most of them acquire their immunological recognition specificities. Once they are committed to the T cell fate, they preserve a core of T cell gene expression through all later T cell development and through all the complex responses that mature T cells make during immune responses.

The key questions are: how does this transformation come about, at the level of gene regulation? Then, how can T cell fate choice become irreversible, yet compatible with extensive self-renewal over the whole life of the organism?

The one environmental signal that is rate-limiting and indispensable for T-cell specification is Notch-Delta signaling.  Notch signals are required throughout the DN stages of T cell development and are guaranteed by expression of the Notch ligand Delta-like 4 in the thymic stroma. Notch-Delta interaction has been used as the basis of powerful in vitro differentiation systems that enable us to dissect the mechanisms involved in the T cell fate choice with great experimental access and flexibility.

Notch signals by themselves can decide many disparate fate choices in embryology, so that in order to instruct cells to become T cells specifically they must be collaborating with particular cell-intrinsic transcription factors. Genetic research based on gene knockouts in mice has led to the identification of several key transcriptional regulators for T cell development.  All of these regulators are required to generate committed T-cell precursors.  However, most of these regulators are also required in different combinations or levels of activity for other cell fates.  There is no single "master regulator" of T-cell differentiation. The right combinations of factors, working the right coordination, are needed to define the T cell path.

The slowness of T cell lineage commitment in response to Notch signaling is striking. Our research has shown that to reach this point, the T-cell program has to overcome two kinds of restraints. One is the presence of inaccessible, repressive chromatin around some key T-cell regulatory genes that can only be gradually converted to open chromatin. The other is the ongoing operation of a stem-like multipotency regulatory program in the cells that first enter the thymus, a program that actively delays full entry into T cell development. Thus, a complex regulatory network must connect the drivers and braking systems of differentiation as cells enter the T cell pathway.