Tonic Signaling Controls Gene Expression in T Lymphocytes. Mus musculus

To determine the role of tonic signals through the adapter LAT in controling helper T cell function, we performed microarray analysis of splenic CD4+ T cells from mice that are WT for LAT, are deficient in LAT (KO), and have a point-mutated version of LAT that abrogates PLCg1 binding (Y136F). These mouse models allow for inducible perturbation or deletion of LAT (using floxed alleles and the Cre-ER system), so we compared gene expression after both 1 and 4 weeks of tamoxifen treatment. We identified a cluster of genes that are downregulated when LAT is perturbed, and these genes are also putative targets of the histone deacetylase HDAC7. We explored this mechanistic link between LAT, HDAC7, and genes in this cluster in CD4+ T cells Overall design: Mice that are CRE ER+ and either LAT WT, LAT f/- (KO), or LAT Y136F/- were treated with tamoxifen for 1 or 4 weeks to inducibly delete or point-mutate LAT. Naïve CD44low CD4+ T cells were sorted from mouse spleens from three mice per genotype and analyzed by Agilent Mouse GE 4x44k arrays.

为明确衔接蛋白LAT（adapter LAT）介导的强直信号在辅助T细胞功能调控中的作用，我们针对三类小鼠的脾脏CD4+ T细胞开展了全基因组微阵列分析：LAT野生型（WT）小鼠、LAT基因敲除型（KO）小鼠，以及携带可阻断磷脂酶Cγ1（PLCγ1）结合的LAT点突变体（Y136F）的小鼠。本研究采用的小鼠模型可通过他莫昔芬诱导实现LAT的靶向扰动或敲除（借助floxed等位基因与Cre-ER系统），因此我们分别比较了他莫昔芬处理1周与4周后的基因表达谱差异。我们筛选得到一个基因簇，其成员在LAT受扰动时表达下调，且该基因簇同时被鉴定为组蛋白去乙酰化酶HDAC7的潜在靶标。我们进一步在CD4+ T细胞中探究了LAT、HDAC7与该基因簇之间的调控机制关联。实验设计概述：携带Cre-ER+且基因型分别为LAT野生型、LAT f/-（KO）或LAT Y136F/-的小鼠，经他莫昔芬处理1周或4周，以诱导性敲除或点突变LAT。每一种基因型选取3只小鼠的脾脏，分选初始型CD44low CD4+ T细胞，采用Agilent Mouse GE 4x44k小鼠基因表达芯片进行检测分析。