Gli2 modulates cell cycle re-entry through autophagy-mediated regulation on the length of primary cilia.

The primary cilium is a tiny cell protrusion known to transduce key extracellular signals, including the sonic hedgehog pathway, which activates Gli transcription factors for various cellular functions. To understand the significance of 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 by Kif3a knockdown rescues the delay in cell cycle re-entry. These results suggest that Gli2 plays an unexpected role for cell cycle re-entry through autophagy-mediated regulation on ciliary length in fibroblasts.