DNA METHYLATION AND T CELL CYTOKINE GENE EXPRESSION

最近几个实验室的研究结果表明，T细胞中细胞因子基因表达的差异是通过细胞因子基因染色质结构和甲基化的差异来表观遗传的。然而，DNA甲基化在体内这一过程中的作用尚未得到直接测试。我们已经有了在CD4+CD8+ (DP)阶段的T细胞发育过程中选择性和有效地切除主要DNA甲基转移酶基因（Dnmtl）的小鼠（CD4 Cre Dnmt 21ox小鼠）。T细胞在这些小鼠体内的发育基本上是正常的。我们打算利用这些小鼠来确定DNA甲基化在细胞因子基因表达调控中的作用。具体来说，我们将解决以下假设：首先，缺乏Dnmt 1的初始CD4和CD8 T细胞在初始激活后比来自对照小鼠的初始T细胞更容易产生效应细胞因子（需要更少的细胞分裂）。其次，与对照T细胞相比，Dnmt-/- T细胞表达的细胞因子模式的差异将不那么明显。第三，Dnmt的缺失将导致细胞因子基因位点周围的染色质结构更加开放。第四，来自CD4 Cre Dnmt 21ox小鼠的CD4和CD8 T细胞将无法发展和维持一种可遗传的、极化的1型与2型细胞因子基因表达模式。最后，CD4 Cre Dnmt 21ox小鼠更倾向于双等位基因而不是单等位基因表达细胞因子基因。

Recent findings from several laboratories suggest that differential cytokine gene expression in T cells is epigenetically enforced through differences in cytokine gene chromatin structure and methylation. However, the role of DNA methylation in this process in vivo has not been tested directly. We have available mice in which the major DNA methyltransferase gene (Dnmtl) is selectively and efficiently ablated during T cell development at the CD4+CD8+ (DP) stage (CD4 Cre Dnmt 21ox mice). T cell development in these mice is essentially normal. We propose to use these mice to determine the role of DNA methylation in the regulation of cytokine gene expression. Specifically, we will address the following hypotheses: First, naive CD4 and CD8 T cells lacking Dnmt 1 will produce effector cytokines more readily (requiring fewer cell divisions) after primary activation than naive T cells from control mice. Second, differences in the pattern of cytokines expressed by Dnmt-/- T cells will be less evident than in control T cells. Third, the loss of Dnmt will result in a more open chromatin structure surrounding cytokine gene loci. Fourth, CD4 and CD8 T cells from CD4 Cre Dnmt 21ox mice will be unable to develop and maintain a heritable, polarized type 1 vs. type 2 pattern of cytokine gene expression. Lastly, CD4 Cre Dnmt 21ox mice will favor biallelic over monoallelic expression of cytokine genes.