The Hedgehog-regulated transcription factors GLI1 and GLI2 play overlapping roles in development and disease; however, the mechanisms underlying their interplay remain elusive. We report for the first time that GLI1 and GLI2 physically and functionally interact in cancer cells. GLI1 and GLI2 were shown to co-immunoprecipitate in PANC1 pancreatic cancer cells and RMS13 rhabdomyosarcoma cells. Mapping analysis demonstrated that the zinc finger domains of both proteins are required for their heteromerization. RNAi knockdown of either GLI1 or GLI2 inhibited expression of many well-characterized GLI target genes (BCL2, MYCN, PTCH2, IL7 and CCND1) in PANC1 cells, whereas PTCH1 expression was only inhibited by GLI1 depletion. qPCR screening of a large set of putative canonical and non-canonical Hedgehog/GLI targets identified further genes (e.g. E2F1, BMP1, CDK2) strongly down-regulated by GLI1 and/or GLI2 depletion in PANC1 cells, and demonstrated that ANO1, AQP1 and SOCS1 are up-regulated by knockdown of either GLI1 or GLI2. Chromatin immunoprecipitation showed that GLI1 and GLI2 occupied the same regions at the BCL2, MYCN and CCND1 promoters. Furthermore, depletion of GLI1 inhibited GLI2 occupancy at these promoters, suggesting that GLI1/GLI2 interaction is required for the recruitment of GLI2 to these sites. Together, these findings indicate that GLI1 and GLI2 co-ordinately regulate the transcription of some genes, and provide mechanistic insight into the roles of GLI proteins in carcinogenesis.

中文翻译：

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需要GLI1 / GLI2功能相互作用来控制刺猬/ GLI目标基因表达。

刺猬调节的转录因子GLI1和GLI2在发育和疾病中起着重叠的作用。但是，它们相互影响的机制仍然难以捉摸。我们首次报告GLI1和GLI2在癌细胞中发生物理和功能相互作用。已显示GLI1和GLI2在PANC1胰腺癌细胞和RMS13横纹肌肉瘤细胞中共免疫沉淀。定位分析表明，这两种蛋白的锌指结构域是其杂聚化所必需的。RNAi敲低GLI1或GLI2抑制了PANC1细胞中许多良好表征的GLI靶基因（BCL2，MYCN，PTCH2，IL7和CCND1）的表达，而PTCH1的表达仅受到GLI1耗竭的抑制。qPCR筛选了大量推定的典型和非典型Hedgehog / GLI靶标，从而鉴定出其他基因（例如E2F1，BMP1，CDK2）在PANC1细胞中被GLI1和/或GLI2消耗强烈下调，并证明ANO1，AQP1和SOCS1被GLI1或GLI2的敲低上调。染色质免疫沉淀显示，GLI1和GLI2在BCL2，MYCN和CCND1启动子处占据相同区域。此外，GLI1的耗竭抑制了这些启动子上的GLI2占用，这表明将GLI2募集到这些位点需要GLI1 / GLI2相互作用。总之，这些发现表明，GLI1和GLI2协同调节某些基因的转录，并提供了有关GLI蛋白在致癌作用中的机制的见解。染色质免疫沉淀显示，GLI1和GLI2在BCL2，MYCN和CCND1启动子处占据相同区域。此外，GLI1的耗竭抑制了这些启动子上的GLI2占用，这表明将GLI2募集到这些位点需要GLI1 / GLI2相互作用。总之，这些发现表明，GLI1和GLI2协同调节某些基因的转录，并提供了有关GLI蛋白在致癌作用中的机制的见解。染色质免疫沉淀显示，GLI1和GLI2在BCL2，MYCN和CCND1启动子处占据相同区域。此外，GLI1的耗竭抑制了这些启动子上的GLI2占用，这表明GLI1 / GLI2相互作用是将GLI2募集到这些位点所必需的。总之，这些发现表明，GLI1和GLI2协同调节某些基因的转录，并提供了有关GLI蛋白在致癌作用中的机制的见解。