Role of GATA-3 During CD4/CD8 Lineage Commitment

描述(由申请者提供)：在这项资助中，我们提出了一些实验，旨在详细了解转录因子GATA-3在胸腺中CD4/CD8谱系承诺过程中的作用。我们最近发现，GATA-3在发育成CD4谱系的未成熟胸腺细胞中差异表达，并且通过改变胸腺中的GATA-3活性，我们可以偏向谱系承诺。我们现在将： 分析TCR诱导胸腺GATA-3上调的机制，使用功能获得/功能丧失的遗传学方法来确定在正常DP胸腺细胞中参与这一过程的信号转导途径。 使用重组胎儿胸腺器官培养(RFTOC)、慢病毒转基因和不同的GATA-3突变体来定位其在CD4/CD8谱系承诺过程中所需的GATA-3结构区域。 分析GATA-3与两个转录因子Egr-2和LMO-4的相互作用，我们已经确定这两个转录因子在全基因组筛选的CD4谱系中间体中优先表达。 这些研究将使我们更好地了解控制CD4/CD8谱系承诺的机制，更广泛地说，将提供关于在胸腺中建立细胞命运决定和发育程序的遗传网络的线索。

DESCRIPTION (provided by applicant): In this grant we propose a number of experiments aimed at understanding in detail the role of the transcription factor GATA-3 during CD4/CD8 lineage commitment in the thymus. We have recently shown that GATA-3 is differentially expressed in immature thymocytes that develop into the CD4 lineage, and that by modifying GATA-3 activity in the thymus we can bias lineage commitment. We will now: Dissect the mechanisms that regulate TCR-induced GATA-3 upregulation in the thymus, using a gain-of function/loss-of-function genetic approach to identify the signal transduction pathways involved in this process in normal DP thymocytes. Use reaggregate fetal thymic organ culture (rFTOC), lentiviral transgenesis and different GATA-3 mutants to map the structural regions of GATA-3 required for its effect during CD4/CD8 lineage commitment. Analyze the interactions of GATA-3 with two transcription factors, Egr-2 and Lmo-4, that we have identified as preferentially expressed in CD4 lineage intermediates in a genome-wide screen. These studies should provide us with a better understanding of the mechanisms that control CD4/CD8 lineage commitment, and, more widely, will provide clues about the genetic networks that establish cell fate determination and developmental programs in the thymus.