IL-2 Family Cytokines and their Receptors-- Molecular Regulation via GABP

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The interleukin-2 receptor and related cytokine/cytokine receptor systems are being studied to understand critical components of the T cell immune response in normal and neoplastic cells. Following T-cell activation, IL-2 and IL-2 receptors are induced; the magnitude and duration of the T-cell immune response is controlled by the amount of IL-2 produced, the levels of receptors expressed, and the time course of these events. Three chains of the IL-2 receptor exist, IL-2Ra, IL-2Rb, and gc, with IL-2Ra and IL-2Rb being significantly regulated at the level of transcription. gc is a shared chain also used by the receptors for IL-4, IL-7, IL-9, IL-15, and IL-21, and is the protein that is mutated in XSCID. We have focused primarily on the types of signals induced by some of these cytokines, and the basis for regulation of these cytokines and their receptors. Previously, we demonstrated that IL-7 receptor alpha chain expression is negatively regulated by IL-2. Although some information was available regarding the basis for regulation of the IL-7 receptor in B cells, essentially nothing was known in T cells. In B cells, an Ets family transcription factor, PU.1, is critical for control of the gene. We previously discovered that the same Ets binding site that binds PU.1 in B cells is also essential for IL-7Ra expression in T cells, but that in T cells, the critical factor is GA binding protein (GABP). Additionally, mice with diminished levels of the GABP alpha chain were generated by a gene-trap methodology. Embryos with diminished expression of GABPa exhibited markedly decreased numbers of thymocytes and lower IL-7Ra expression on those cells. In the previous reporting period, we observed that GABP was also a critical regulator of B cell development, acting in part by regulating Pax5 and Pax5 target genes and that GABP and PU.1 control maturation stage-specific regulation of megakaryopoiesis. In the current year, we reported that GABP regulates expression of the gene encoding a serine-threonine kinase, denoted KIS, that phosphorylates p27Kip1, a cyclin-dependent kinase inhibitor, and is important for cell migration and cell cycle progression. Interestingly, knocking down GABPa expression by siRNA in vascular smooth muscle cells diminished KIS gene expression and reduced cell migration. Moreover, after serum stimulation, KIS gene expression was diminished in the GABPa KO cells, with decreased cells in S-phase. Furthermore, after vascular injury in vivo, mice heterozygous for GABPa had diminished KIS expression within arterial lesions. These results overall underscored an important role for GABPa for KIS expression and function. GABP comprises both alpha and beta chains, with the beta chain having both long and short isoforms. The L form can form alpha-beta heterotetramers whereas the S form cannot. To learn more about the function of GABPb, we selectively targeted the long form that was believed to be essential for GABP action and generated GABPb-L form KO mice that still retained expression of the GABPb-S form. Unexpectedly, rather than having a severe phenotype, there was no defect in lymphoid development, with normal T and B cell development and normal mature T and B cell responses to various stimuli. In contrast, targeting both the L and S forms resulted in early embryonic lethality. These data indicate that contrary to prior belief, GABPb-S is also functional rather than serving as a dominant negative. We also studied IL-7Ra expression in CD8+ T cells. Virus-specific CD8 T cells typically diminish levels of IL-7Ra and are short lived, but some escape and become memory precursor effector cells. We found that GABPa was required for IL-7Ra expression in memory precursor effector cells whereas Gfi-I was needed for stable IL-7Ra repression in effector T cells. Thus, expression of IL-7Ra is regulated by opposing functions of GABP and Gfi-1, confirming our own earlier reports on GABP and identifying Gfi-1 as being a counterbalancing factor. Overall, the above findings enhance our understanding of mechanisms by which the common cytokine receptor family of cytokines control gene expression to regulate the immune response.