PKB Rescues Calcineurin/NFAT-Induced Arrest of Rag Expression and Pre-T Cell Differentiation

Abstract

Protein kinase B (PKB), an Ag receptor activated serine-threonine kinase, controls various cellular processes including proliferation and survival. However, PKB function in thymocyte development is still unclear. We report PKB as an important negative regulator of the calcineurin (CN)-regulated transcription factor NFAT in early T cell differentiation. Expression of a hyperactive version of CN induces a profound block at the CD25+CD44− double-negative (DN) 3 stage of T cell development. We correlate this arrest with up-regulation of Bcl-2, CD2, CD5, and CD27 proteins and constitutive activation of NFAT but a severe impairment of Rag1, Rag2, and intracellular TCR-β as well as intracellular TCR-γδ protein expression. Intriguingly, simultaneous expression of active myristoylated PKB inhibits nuclear NFAT activity, restores Rag activity, and enables DN3 cells to undergo normal differentiation and expansion. A correlation between the loss of NFAT activity and Rag1 and Rag2 expression is also found in myristoylated PKB-induced CD4+ lymphoma cells. Furthermore, ectopic expression of NFAT inhibits Rag2 promoter activity in EL4 cells, and in vivo binding of NFATc1 to the Rag1 and Rag2 promoter and cis-acting transcription regulatory elements is verified by chromatin immunoprecipitation analysis. The regulation of CN/NFAT signaling by PKB may thus control receptor regulated changes in Rag expression and constitute a signaling pathway important for differentiation processes in the thymus and periphery.

Constitutive Akt1 signals attenuate B‐cell receptor signaling and proliferation, but enhance B‐cell migration and effector function

“…In response to IL-7 signals, STAT5 regulates the survival and proliferation of B-cell progenitors and plays an important role in the generation of B-cell leukemia and lymphoma [36,42]. Notably, both BCR and IL-7 signals regulate the expression of Rag proteins via the PI3K/Akt/FoxO pathway and thus influence early B-cell development [43][44][45][46][47][48][49][50][51]. Therefore, the block of B-cell differentiation in Akt1 +/+ Tg mice is presumably linked to altered Rag expression and impaired recombination of Ig genes.…”

Critical roles of the PI3K/Akt signaling pathway in T cell development

“…Thymocytes that express a constitutively active form of calcineurin, thus maintaining NF-AT nuclear localization, are blocked at the DN3 stage of development. Constitutive NF-AT activation also causes decreased Rag1 protein expression, and thymocytes subsequently develop into DP cells without detectable intracellular TCRβ chain [55]. Transgenic expression of myristoylated Akt rescues the calcineurin induced block and restores expression of Rag proteins and the TCRβ chain in DP thymocytes.…”

“…Constitutively active Akt can modulate Rag expression during T cell development, possibly through NF-AT (nuclear factor of activated T cells) [55]. Thymocytes that express a constitutively active form of calcineurin, thus maintaining NF-AT nuclear localization, are blocked at the DN3 stage of development.…”

Epigenetic Changes and Suppression of the Nuclear Factor of Activated T Cell 1 (NFATC1) Promoter in Human Lymphomas with Defects in Immunoreceptor Signaling

“…22 The NFATc2-induced motility and invasion of breast cancer cells can be blocked by Akt/PKB signals, 23 which were shown to suppress the activation of NFATcs in thymocytes and peripheral T cells. 24,25 Taken together, these data indicate that alterations in calcineurin/NFATc signaling play an important role in human cancerogenesis, and because of their immunoreceptor-dependent activation, aberrant NFATc activity affects particularly the generation and maintenance of human lymphomas.…”