Scientific Report 2005

Immunology

Regulators of T-Cell Development and Lymphocyte Function

J. Kaye, P. Aliahmad, O. Garijo, P. Han, C. Krieg

Precursor cells in the thymus undergo a complex developmental program before seeding peripheral lymphoid organs as mature T lymphocytes. Developmental checkpoints in the thymus, termed β-selection, positive selection, and negative selection, narrow the repertoire of T-cell antigen specificities to those that are not overtly autoreactive but maintain weak reactivity against self-MHC-peptide complexes. We are interested in the mechanisms that determine the fate of developing T cells and the control of gene expression during these developmental processes. Our identification of a ell-surface protein that is upregulated on developing thymocytes also led to studies on regulation of the immune response and the potential of this protein as a novel therapeutic target.

A Nuclear Protein Involved in Regulation of Thymocyte Selection

We identified thymocyte selection–associated high mobility group (HMG) box protein (TOX) several years ago. Members of the HMG box protein superfamily share one or more copies of a sequence-related and structurally related DNA-binding domain that can recognize distorted DNA structures and modify chromatin by bending DNA. In general, HMG box proteins function as architectural factors that regulate gene expression by promoting formation of transcriptional complexes or by acting as components of chromatin-remodeling complexes.

We found that TOX belongs to a small subfamily of evolutionarily conserved proteins whose members share almost identical HMG box sequences. The HMG box sequence in TOX can recognize distorted DNA but is a relatively poor bender of DNA, because of the lack of a critical internal wedge residue.

Expression of TOX in the thymus is tightly regulated. The protein is expressed in early thymocyte progenitors and then transiently upregulated during β-selection and positive selection. Using transgenic animals that express wild-type TOX or mutants of the protein, we investigated the role of this nuclear factor in positive selection and the associated commitment to the CD4⁺ or CD8⁺ T-cell lineages. Our data indicate that expression of TOX is sufficient to initiate the differentiation of immature thymocytes to the CD8⁺ T-cell lineage, even in the absence of signals mediated by T-cell antigen receptors. Both the DNA-binding domain and the N-terminal domain of TOX are required for this in vivo activity. Interestingly, expression of TOX coupled with IL-7–mediated signals appears to be sufficient to induce both the development and parital maturation of CD8⁺ lineage T cells.

Signaling through the serine/threonine phosphatase calcineurin is required for positive selection of thymocytes. We found that expression of the gene for TOX is regulated by this signaling pathway. These studies led to a model in which upregulation of TOX is a critical component of cell-lineage decisions during positive selection. We are producing mice that lack the gene for TOX to shed further light on the role of TOX in vivo.

A Cell-Surface Lymphocyte and Antigen-
Presenting Cell Protein with Regulatory Function

The functional outcome of engagement of the T-cell antigen receptor is modulated by secondary signals, which can have costimulatory or coinhibitory functions. We isolated a gene that encodes a cell-surface protein of the immunoglobulin superfamily, now designated BTLA (B- and T-lymphocyte attenuator), that is upregulated during positive selection and that is expressed by mature lymphocytes and antigen-presenting cells. Evidence indicates that this protein can act as a negative regulator of lymphocyte activation. We produced mice that lack the gene for BTLA and panels of monoclonal antibodies specific for BTLA to analyze the in vivo function of this protein. One of our monoclonal antibodies acts as an agonist for this coinhibitory molecule, thereby inhibiting facets of T-cell activation, including T-cell proliferation and IL-2 secretion. We are testing the ability of this antibody to modulate immune responses in several model systems.

Publications

Kreig, C., Han, P., Stone, R., Goularte, O.D., Kaye, J. Functional analysis of BTLA engagement of CD4⁺ and CD8⁺ T cells. J. Immunol., in press.

Parinaz, A., Kaye, J. Commitment issues: linking positive selection signals and lineage diversification in the thymus. Immunol. Rev., in press.