Institute of Brain Science, School of Medicine, National Yang-Ming University, Taipei 112, Taiwan.
Taiwan International Graduate Program (TIGP) in Molecular Medicine, National Yang-Ming University and Academia Sinica, Taipei 112, Taiwan.
J Cell Sci. 2018 Dec 17;131(24):jcs221218. doi: 10.1242/jcs.221218.
The primary cilium is a tiny cell protrusion known to transduce key extracellular signals, including those of the sonic hedgehog pathway, which activates Gli transcription factors for various cellular functions. To understand the significance of the Gli2 transcription factor in fibroblasts, we establish a Gli2-knockout NIH3T3 cell line by CRISPR/Cas9 technology. Surprisingly, NIH3T3 fibroblasts lacking Gli2 expression through gene knockout or RNA interference possess longer primary cilia after stimulation of ciliogenesis by serum starvation. This lengthening of primary cilia is associated with enhanced autophagy-mediated Ofd1 degradation, and can be reversed by pharmacological and genetic inhibition of autophagy. Meanwhile, flow cytometry reveals that Gli2 NIH3T3 fibroblasts exhibit a delay in cell cycle re-entry after serum re-stimulation. Ablation of their primary cilia through Kif3a knockdown rescues the delay in cell cycle re-entry. These results suggest that Gli2 plays an unexpected role in cell cycle re-entry through an autophagy-mediated regulation on ciliary length in fibroblasts.
初级纤毛是一种微小的细胞突起，已知可以转导关键的细胞外信号，包括 sonic hedgehog 通路的信号，该通路激活 Gli 转录因子以实现各种细胞功能。为了了解 Gli2 转录因子在成纤维细胞中的意义，我们通过 CRISPR/Cas9 技术建立了 Gli2 敲除 NIH3T3 细胞系。令人惊讶的是，通过基因敲除或 RNA 干扰使 NIH3T3 成纤维细胞缺乏 Gli2 表达后，在用血清饥饿刺激纤毛发生时，初级纤毛会变长。这种初级纤毛的延长与增强的自噬介导的 Ofd1 降解有关，并且可以通过自噬的药理学和遗传学抑制来逆转。同时，流式细胞术显示 Gli2 NIH3T3 成纤维细胞在血清再刺激后细胞周期重新进入时出现延迟。通过 Kif3a 敲低去除它们的初级纤毛可以挽救细胞周期重新进入的延迟。这些结果表明，Gli2 通过自噬介导的对成纤维细胞中纤毛长度的调节，在细胞周期重新进入中发挥了意想不到的作用。
