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

人们正在研究白介素2受体和相关的细胞因子/细胞因子受体系统，以了解正常细胞和肿瘤细胞中T细胞免疫反应的关键成分。T细胞激活后，IL-2和IL-2受体被诱导；T细胞免疫反应的大小和持续时间由IL-2产生的量、受体的表达水平和这些事件的时间进程控制。IL-2受体存在三条链，即IL-2Ra、IL-2Rb和GC，其中IL-2Ra和IL-2Rb在转录水平上受到显著调控。GC是IL-4、IL-7、IL-9、IL-15和IL-21受体的共享链，是XSCID中突变的蛋白质。我们主要关注其中一些细胞因子诱导的信号类型，以及这些细胞因子及其受体调节的基础。 在此之前，我们证明了IL-7受体α链的表达受IL-2的负性调节。尽管有一些关于调节B细胞中IL-7受体的基础的信息，但基本上对T细胞一无所知。在B细胞中，ETS家族转录因子PU.1对该基因的控制至关重要。我们先前发现，在B细胞中与PU1结合的相同的ETS结合位点也是T细胞表达IL-7ra所必需的，但在T细胞中，关键因素是GA结合蛋白(GABP)。此外，GABPα链水平降低的小鼠是通过基因陷阱方法产生的。GABPA表达减弱的胚胎胸腺细胞数量明显减少，IL-7ra表达降低。在之前的报告期间，我们观察到GABP也是B细胞发育的关键调节因子，部分通过调节Pax5和Pax5靶基因发挥作用，GABP和PU.1控制成熟阶段对巨核细胞生成的特异性调节。 在最近的一年里，我们报道了GABP调节编码丝氨酸苏氨酸激酶的基因的表达，KIS是一种细胞周期蛋白依赖的激酶抑制物p27Kip1的磷酸化，对细胞迁移和细胞周期进程是重要的。有趣的是，通过siRNA下调血管平滑肌细胞中GABPA的表达会减少KIS基因的表达，并减少细胞的迁移。此外，经血清刺激后，GABPA KO细胞中Kis基因表达减弱，细胞处于S期。此外，体内血管损伤后，GABPA杂合子小鼠的动脉损伤中KIS表达减少。这些结果总体上强调了GABPA在KIS表达和功能中的重要作用。 GABP由α链和β链组成，β链既有长的异构体，也有短的异构体。L形式可以形成α-β异构体，而S形式不能。为了进一步了解GABP的功能，我们选择性地以GABP作用所必需的长形式为靶点，并从仍然保留GABP-S形式表达的KO小鼠中产生了GABP-S形式。出乎意料的是，没有严重的表型，淋巴发育没有缺陷，T和B细胞发育正常，成熟的T和B细胞对各种刺激的反应也正常。相比之下，针对L和S两种类型的病毒会导致早期胚胎死亡。这些数据表明，与先前的看法相反，GABP-S也是功能性的，而不是作为显性否定。 我们还研究了IL-7ra在CD8T细胞中的表达。病毒特异性的CD8T细胞通常会降低IL-7ra的水平，而且寿命很短，但一些细胞会逃脱，成为记忆前体效应细胞。我们发现GABPA是记忆前体效应细胞表达IL-7ra所必需的，而GFI-I是稳定抑制效应T细胞IL-7ra所必需的。因此，IL-7ra的表达受GABP和GFI-1相反功能的调节，证实了我们早期关于GABP的报道，并确认GFI-1是一个平衡因子。 总体而言，上述发现加深了我们对常见的细胞因子受体家族控制基因表达以调节免疫反应的机制的理解。

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.