Distinct contributions of CCR4 and CCR7 to central tolerance

T cells are integral to the adaptive immune response against foreign pathogens and tumor cells. They respond to foreign substances, termed antigens, that can elicit immune responses. After becoming activated due to recognition of antigens presented by professional antigen-presenting cells (APCs), T cells can mount multiple effector functions, such as cytotoxic activities, which eliminate infected cells and tumor cells, and helper activities that assist other immune cells’ responsiveness. Memory T cells, which differentiate upon antigen encounter, respond rapidly to subsequent antigen exposure and provide long-lasting protection against pathogens. While one T cell can recognize few antigens, T cells collectively can recognize and respond to a very diverse array of antigens. This unique property by which T cells recognize and respond to a myriad of antigens stems from the rearrangement of their T cell receptor (TCR) genes during development. The highly random TCR rearrangement during development gives each T cell a unique TCR that can recognize a specific antigen, and collectively, the entire T cell repertoire is able to respond to a wide range of antigens. However, autoreactive T cells, which express TCRs that recognizes normal antigens found in the host, could potentially arise from this TCR rearrangement process, posing a potential risk of mounting an autoimmune reaction and damaging the host’s healthy tissues and organs if released to the periphery. The immune system has multiple tolerance mechanisms in place to prevent autoimmunity, and one such tolerance mechanism, termed central tolerance, works by inducing apoptosis of developing autoreactive T cells or by diverting them to the suppressive regulatory T cell (Treg) lineage. T cell development takes place in the thymus. Developing T cells (thymocytes) migrate through the thymic cortex and the medulla at different developmental stages, interacting with and receiving differentiation, proliferation, and selection cues from different thymic stromal cells. After being positively selected for correctly rearranged TCRs that are able to bind self-peptide:MHC complexes expressed by the host, thymocytes migrate from the cortex into the medulla, where they interrogate medullary stromal cells including medullary thymic epithelial cells, dendritic cells (DCs), and B cells. These medullary stromal cells express, process, and present a broad range of self-antigens. Thymocytes that react strongly to the presented self-antigens undergo apoptosis, a process referred to as “negative selection”, or they differentiate into Tregs, thereby removing autoreactive T cells from the naïve T cell repertoire and establishing central tolerance. Chemokine receptor signaling contributes to the orchestrated movement and localization of thymocytes, and their interaction with thymic stromal cells at different development stages. Chemokine receptors are G-protein coupled receptors that respond to a family of small cytokines termed chemokines. Thymocytes upregulate different chemokine receptors at various development stages and receive chemokine cues from different thymic stromal cells. Chemokine receptor signaling plays an important role in thymocytes’ directional movement, speed changes and interactions with thymic stromal cells. Some thymic stromal cells also express chemokine receptors, which may facilitate their own survival, differentiation, and localization within the thymus. The mechanism by which chemokine receptor signaling contributes to the establishment of central tolerance is not fully elucidated. In this dissertation, I will report on the distinct roles of the chemokine receptors CCR4 and CCR7 in the establishment of central tolerance in the thymus. While the role of CCR7 in central tolerance has been relatively well-characterized, the contribution of CCR4 to this process is not well understood. Both CCR4 and CCR7 are expressed by thymocytes after positive selection, raising the question of whether these two chemokine receptors have distinct or redundant impacts on medullary accumulation and tolerance of post-positive selection thymocytes. In Chapter 2, I investigate the roles of CCR4 and CCR7 expression on thymocyte negative selection. Our data reveal a striking disconnect between CCR7 expression and function on intermediate post-positive selection thymocyte subsets, arguing against the consensus in the field that CCR7 expression decisively locates thymocytes to the medulla. Instead, we find that thymocytes express CCR4 within hours of positive selection signaling and swiftly migrate into the thymic medulla in a CCR4-dependent manner, days before CCR7 expression begins. Additionally, CCR4 and CCR7 promote medullary accumulation of immature versus mature post-positive selection thymocyte subsets, and contribute to early versus late phases of negative selection, respectively. The contribution of CCR4 to central tolerance against activated APCs is then highlighted. Collectively, this study suggests a layered model of central tolerance in which CCR4 directs medullary migration and negative selection of immature post-positive selection thymocytes, and CCR7 directs medullary migration and negative selection of more mature post-positive selection thymocytes. The exact mechanism by which chemokine receptors regulate Treg generation in the thymus is not fully understood. In Chapter 3, I investigate the impact of CCR7 expression on thymic dendritic cells (DCs) on the thymic Treg compartment. An increase in Treg generation in thymus was discovered in Ccr7 [superscript -/-] mice. Surprisingly, CCR7 deficiency in the thymic DC compartment, rather than on the thymocytes, contributed to the increased Treg generation in Ccr7 [superscript -/-] mice. CCR7 is expressed by MHCII [superscript hi] thymic conventional DCs (cDCs) of both the Sirpα⁻ cDC1 and Sirpα⁺ cDC2 lineages. Ccr7 deficiency does not alter the cell-intrinsic potential of different DC subsets to induce Treg; however, CCR7 deficiency results in increased apoptosis of MHCII [superscript hi] Sirpα⁻ DCs and a relative increase of MHCII [superscript lo] Sirpα⁺ DCs, which possess the greatest potential to induce Tregs of all thymic DC subsets. This study reveals that CCR7 deficiency alters the composition of the thymic DC compartment, thus dysregulating the induction of Treg. Together, my studies further elucidate the contribution of chemokine receptor signaling to the maintenance of central tolerance in the thymus and provide additional insights into the understanding of mechanisms underlying autoimmune diseases.