Date Published: January 26, 2010
Publisher: Public Library of Science
Author(s): Jianfang Hu, Qian Qi, Avery August, Derya Unutmaz. http://doi.org/10.1371/journal.pone.0008891
Abstract: T cell development is critically dependent on both the environment and signals delivered by the T cell Receptor (TCR). The Tec family kinase Itk has been suggested to be an amplifier of signals emanating from the TCR and the loss of Itk partially affects most stages of thymopoiesis. Loss of Itk also differentially affects the development of conventional vs. non-conventional or innate memory phenotype T cells. Here, we examine whether these lineage choices are affected by a combination of TCR affinity and Itk by analyzing mice lacking Itk and carrying two TCR transgenes with differing affinities, OT-II and DO11.10. Our results show that developing thymocytes receive a gradient of signals, DO11.10>OT-II>DO11.10/Itk−/−>OT-II/Itk−/−. We also show that the development of CD4+ T cells is controlled by TCR signaling via Itk, which regulates the expression of the transcription factor, Th-POK, an enforcement factor for CD4 commitment. This results in a reduction in CD4+ T cell development, and an increase in the development of MHC class II restricted TCR transgenic CD8+ T cells that resemble non-conventional or innate memory phenotype CD8 T cells. This alteration accompanies increased expression of Runx3 and its target genes Eomesodermin, Granzyme B and Perforin in Itk null OT-II CD4+ thymocytes. All together, these data suggest that Itk plays an important role in CD4/CD8 commitment by regulating signal thresholds for the lineage commitment. Our data also suggest that the lower level of TCR signaling that occurs with a low affinity TCR in the absence of Itk can redirect some MHC class II restricted CD4+ T cell to class II-restricted CD8+ innate memory phenotype T cells.
Partial Text: Mature T cell development takes place in the thymus and is critically dependent on signals through the TCR. During T cell development, TCR signals generated by interaction with major histocompatibility complex class (MHC)-II peptide complexes are required for differentiation of CD4+ T cells, while TCR signals generated by interaction with class I MHC-peptide complexes are required for differentiation of CD8+ T cells. This process, referred as CD4 and CD8 commitment, is a major developmental process after positive and negative selection. While TCR signals are important for this lineage, they are not completely clarified , , . High signaling activity generated by the tyrosine kinase Lck or MAP kinases ERK1/2 enhances CD4 Single Positive (SP) development , , while low activity of Lck, ZAP70 or ERK1/2 leads to CD8 SP development , , . These findings support the idea that attenuating TCR signaling could redirect thymocytes with class II restricted TCRs from CD4 to the CD8 lineage, while enhanced signaling could redirect thymocytes with class I specific TCRs from CD8 to CD4 lineage. Other evidence indicates that the duration of signaling as well as the number of TCRs triggered are key factors in determining CD4/CD8 T cell fate decision , . A role for TCR affinity has not been implicated in these events, but may play a role dependent on whether TCR affinity affects these implicated signaling events.
This study was designed to examine whether CD4/CD8 lineage choices are affected by the reduced TCR signals in the absence of Itk. We analyzed Itk−/− mice crossed to mice carrying two TCRs reactive against ovalbumin, the low affinity MHC class II-restricted transgenic TCR (OT-II) and the higher affinity (50 fold) MHC class II-restricted transgenic TCR (DO11.10). Our results show that dependent on the affinity of the TCR, Itk affects the development of naïve CD4+ T cells. Our results also suggest that this occurs via Itk mediated (direct or indirect) regulation of Th-POK expression, suggesting that the strength of the TCR signal modulates Th-POK expression and thus CD4+ T cell development. Finally, our results suggest that by modulating Th-POK expression, Itk derived signals affect the expression of CD8 patterning genes such as Runx3, which regulates the expression of Eomesodermin, Granzyme B and Perforin in naïve or conventional CD4SP thymocytes, altering the potential response of these T cells. Finally, our results suggest that at significantly reduced TCR affinity, “non conventional” CD8+ T cells begin to develop.