Hedgehog signaling in T cell differentiation

Despite Hedgehog’s influence on T-cell activation and proliferation, the transcriptional targets of Gli2 in lymphocytes are not known. We therefore examined the Hedgehog-dependent transcriptional response of resting and early-stage activated T-cells in order to define their transcriptional response to Hedgehog pathway activation. We have established transgenic models where transcription of target genes by Gli2 is either constitutively activated or repressed in cells of the T-lineage. Gli2 has an N-terminal repressor domain and a C-terminal activator domain. Lck-Gli2ΔN2 (Gli2A) mice carry a transgene encoding a truncated form of Gli2 that acts as a permanent transcriptional activator of Hh target genes. Conversely, Lck-Gli2ΔC2 (Gli2R) mice express a repressor of Gli2-dependent transcription, which by binding to Gli binding sites inhibits physiological Hh-dependent transcription, and hence Hh signal transduction, in the cell. These transgenes are driven by the Lck promoter and are only expressed in T-lineage cells Purified fresh, resting CD4 cells (unstimulated) and from CD4 cells stimulated with anti-CD3/CD28 for 6h (stimulated) were analysed in order to obtain transcriptional profiles before and during the early stages of T-cell activation.

尽管刺猬信号通路（Hedgehog, Hh）对T细胞活化与增殖具有调控作用，但淋巴细胞中Gli2的转录靶标尚未明确。为此，我们针对静息态与早期活化T细胞的刺猬信号依赖性转录应答开展研究，以期明确其对刺猬信号通路活化的转录响应模式。我们已构建转基因模型，可在T细胞谱系细胞中实现Gli2介导的靶基因转录的组成型激活或抑制。Gli2兼具N端抑制结构域与C端激活结构域。Lck-Gli2ΔN2（Gli2A）小鼠携带的转基因编码截短型Gli2，可作为Hh靶基因的永久性转录激活因子。与之相反，Lck-Gli2ΔC2（Gli2R）小鼠表达Gli2依赖性转录的抑制因子，该因子通过结合Gli结合位点，抑制生理状态下的Hh依赖性转录，进而阻断细胞内的Hh信号转导通路。上述转基因均由Lck启动子驱动，仅在T细胞谱系细胞中特异性表达。我们对纯化得到的新鲜静息CD4阳性T细胞（未受刺激），以及经抗CD3/抗CD28共刺激6小时的CD4阳性T细胞（受刺激）进行转录组分析，以获取T细胞活化早期阶段及其之前的转录谱数据。