ERK Signaling Controls Innate-like CD8+ T Cell Differentiation via the ELK4 (SAP-1) and ELK1 Transcription Factors.

In mouse thymocyte development, signaling by the TCR through the ERK pathway is required for positive selection of conventional naive T cells. The Ets transcription factor ELK4 (SAP-1), an ERK-regulated cofactor of the SRF transcription factor, plays an important role in positive selection by activating immediate-early genes such as the Egr transcription factor family. The role of ELK4-SRF signaling in development of other T cell types dependent on ERK signaling has been unclear. In this article, we show that ELK4, and its close relative ELK1, act cell autonomously in the thymus to control the generation of innate-like αß CD8+ T cells with memory-like characteristics. Mice lacking ELK4 and ELK1 develop increased numbers of innate-like αß CD8+ T cells, which populate the periphery. These cells develop cell autonomously rather than through expansion of PLZF+ thymocytes and concomitantly increased IL-4 signaling. Their development is associated with reduced TCR-mediated activation of ELK4-SRF target genes and can be partially suppressed by overexpression of the ELK4-SRF target gene EGR2. Consistent with this, partial inhibition of ERK signaling in peripheral CD8+T cells promotes the generation of cells with innate-like characteristics. These data establish that low-level ERK signaling through ELK4 (and ELK1) promotes innate-like αß CD8+ T cell differentiation, tuning conventional versus innate-like development.

Ternary complex factors SAP-1 and Elk-1, but not net, are functionally equivalent in thymocyte development.

The ternary complex factors (TCFs; SAP-1, Elk-1, and Net) are serum response factor cofactors that share many functional properties and are coexpressed in many tissues. SAP-1, the predominant thymus TCF, is required for thymocyte positive selection. In this study, we assessed whether the different TCFs are functionally equivalent. Elk-1 deletion, but not the hypomorphic Net(delta) mutation, exacerbated the SAP-1 positive selection phenotype, but triply deficient thymocytes were no more defective than SAP-1(-/-) Elk-1(-/-) cells. Inactivation of the other TCFs did not affect SAP-1-independent processes, including beta-selection, regulatory T cell selection, and negative selection, although reduced marginal zone B cells were observed in SAP-1(-/-) Elk-1(-/-) animals. Ectopic expression of Elk-1, but not Net, rescued positive selection of SAP-1(-/-) thymocytes; thus, SAP-1 and Elk-1 are functionally equivalent in this system, and the SAP-1 null selection phenotype reflects only its high expression in the thymus. Array analysis of TCR-stimulated double-positive cells identified SAP-1-dependent inducible genes whose transcription was further impaired in SAP-1(-/-) Elk-1(-/-) cells; thus, these genes, which include Egr-1 and Egr-2, represent candidate mediators of positive selection. Chromatin immunoprecipitation revealed subtly different promoter targeting between the different TCFs. Ectopic expression of Egr-1 restored positive selection in SAP-1 null thymocytes, establishing it (and possibly other Egr family members) as the major effector for ERK-SAP-1 signaling in thymocyte positive selection.

Raf signaling but not the ERK effector SAP-1 is required for regulatory T cell development.

Regulatory T cells (T(reg)) play an important role in immune regulation. Their development in the thymus requires TCR activation and recognition of peptide-MHC, although the downstream signals controlling commitment to the lineage are unclear. To compare the requirements for positive selection and T(reg) development, we studied knockout and transgenic mice defective in Raf signaling and the ERK effector SRF accessory protein 1 (SAP-1), a member of the ternary complex factor family of Ets domain transcription factors. Although SAP-1 deficient mice display a severe defect in thymocyte positive selection, T(reg) development was unimpaired as assessed by expression of Foxp3 and the activation markers CD25, GITR, CTLA4, and CD103 in the CD4(+) cell population. In contrast, inhibition of Raf signaling by the interfering dominant negative Raf derivative reduced both Foxp3(+) and Foxp3(-) CD4(+) populations. In SAP-1-deficient CD4(+)CD25(+) T(reg) cells, TCR crosslinking efficiently induced ERK activation, but transcriptional induction of the immediate early gene Egr-1 was impaired. Nevertheless, neither deletion of SAP-1 nor expression of a dominant negative Raf derivative affected the ability of CD4(+)CD25(+) T(reg) cells to suppress CD4(+)CD25(-) cell proliferation in vitro. Finally the suppressive activity of CD4(+)CD25(+) T(reg) cells lacking SAP-1 in an in vivo colitis model was not significantly impaired. The signaling requirements for development of T(reg) cells in the thymus are thus distinct from those required for "conventional" T cell positive selection, and ERK signaling to SAP-1 is not required for the suppressive activity of T(reg) cells.