PP6 Controls T Cell Development and Homeostasis by Negatively Regulating Distal TCR Signaling

Abstract

T cell development and homeostasis are both regulated by TCR signals. Protein phosphorylation and dephosphorylation, which are catalyzed by protein kinases and phosphatases, respectively, serve as important switches controlling multiple downstream pathways triggered by TCR recognition of Ags. It has been well documented that protein tyrosine phosphatases are involved in negative regulation of proximal TCR signaling. However, how TCR signals are terminated or attenuated in the distal TCR signaling pathways is largely unknown. We investigated the function of Ser/Thr protein phosphatase (PP) 6 in TCR signaling. T cell lineage-specific ablation of PP6 in mice resulted in enhanced thymic positive and negative selection, and preferential expansion of fetal-derived, IL-17–producing Vγ6Vδ1+ T cells. Both PP6-deficient peripheral CD4+ helper and CD8+ cytolytic cells could not maintain a naive state and became fast-proliferating and short-lived effector cells. PP6 deficiency led to profound hyperactivation of multiple distal TCR signaling molecules, including MAPKs, AKT, and NF-κB. Our studies demonstrate that PP6 acts as a critical negative regulator, not only controlling both αβ and γδ lineage development, but also maintaining naive T cell homeostasis by preventing their premature activation before Ag stimulation.

Functions of protein phosphatase-6 in NF-κB signaling and in lymphocytes

“…However, e x vivo PMA/ionomycin stimulation of splenocytes from SAPS1-deficient mice resulted in a greater proportion of IL-4-producing CD4 + T cells when compared with corresponding controls. This response was similar in lymphocytes with either PP6c or SAPS1 knockout [46 and unpublished data]. Furthermore, in vitro differentiation assays demonstrate that SAPS1-deficient CD4 T cells favor differentiation into IL-4-producing cells but not differentiation into interferon-γ-producing cells.…”

“…Ppp6c homozygous null mutations are early embryonic lethal [45], necessitating use of conditional knockout approaches to study the role of PP6 in vivo . Ye et al [46] engineered mice that are homozygous for a LoxP-targeted allele on chromosome 2, which were crossed with Lck-Cre or CD4-Cre mice to drive selective deletion of the gene for PP6 catalytic subunit (PP6c) at early or intermediate stages of thymocyte development, respectively. In an alternative approach, we have produced mice that are homozygous for a LoxP-targeted Ppp6r1 allele (encoding SAPS1) on chromosome 7, which were crossed with Sox2-Cre to delete the SAPS1 regulatory subunit in all adult tissues.…”

“…Interestingly, the increased number of DN cells was due to an increase in the number of γ/δ-lineage thymocytes. More detailed analysis revealed significantly increased use of γv1, γv5 and, particularly, γv6 variable region gene segments in PP6c-deficient γδ-thymocytes [46]. Subsequent experiments demonstrated that the increased number of γδ-thymocytes arose from ongoing expansion of fetal-derived γv6/δv1 producing thymocytes rather than increased production of γδ-thymocytes from bone marrow-derived hematopoietic stem cells.…”

“…To get around this problem, Ye and colleagues used Pp6c f/f CD4-cre mice [46]. These mice initiate deletion in the DP compartment and were found to contain a larger peripheral CD4 + and CD8 + splenic T cell compartment than Pp6c f/f lck-cre mice, although the number of CD4 + and CD8 + splenic T cells was still reduced by ∼70 and 60%, respectively.…”

“…However, both CD4 + and CD8 + T cells had a faster turnover rate and were prone to apoptotic cell death. In addition to disrupted homeostasis of CD4 + and CD8 + T cells, there was a notable increase in the frequency and number of interferon-γ producing CD4 + and CD8 + effector T cells in Pp6c f/f CD4-cre mice as well as a modest increase in the number of IL-4-producing CD4 T cells induced ex vivo by PMA/ionomycin [46]. Thus, loss of PP6c during thymocyte development results in significant defects in thymic output, seemingly as a result of increased negative selection, as well as hyperactivation, increased proliferation and apoptotic cell death in the splenic CD4 + and CD8 + T cell compartments.…”