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

Abstract

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.

Impairment in dynein-mediated nuclear translocation by BICD2 C-terminal truncation leads to neuronal migration defect and human brain malformation

“…HeLa and U2OS cells were plated on 12-mm-diameter glass coverslips coated with poly-D-lysine (Sigma Aldrich) for cell staining [21,25]. HA-BicD2 WT or other mutant constructs were transfected into cells for 24 h. For identifying the interaction between Nesprin-2 and BicD2, cells were synchronized in G0 phase by serum starvation for 24 h. The detailed steps for cell staining were performed as described previously [22].…”

“…Signal of DAPI was excited by a 405-nm laser. Analysis of images were carried out with Zen software (Zeiss) of ImageJ (NIH) [21,25].…”

Primary Cilium in Cancer Hallmarks

“…Col 1 was found to be promote cilia growth by repressing HDAC6-mediated autophagy, thus indicating a negative role for autophagy in ciliogenesis [142]. Finally, the GLI2 transcription factor, one of the effectors of Hedgehog signaling, was found to control cell cycle re-entry and cilia length in an autophagy-dependent manner [143]. NIH3T3 fibroblasts lacking Gli2 expression showed longer primary cilia, and increased autophagy, with a reduction in the Ofd1 protein levels in serum-free media, along with a delay in cell cycle re-entry.…”

The Autophagy Regulator p62 Controls PTEN-Dependent Ciliogenesis

“…Early approaches examined whether the two processes affected each other under same stimulus, with research now focusing on identifying specific mediators. Recent studies have observed multiple Atg proteins near primary cilia and have shown that ciliary components are controlled by autophagy (Boehlke et al, 2010;Kim et al, 2015;Shin et al, 2015a,b;Wang et al, 2015;Orhon et al, 2016;Takacs and Proikas-Cezanne, 2016;Hsiao et al, 2018;Liu et al, 2018;Xu et al, 2018). Another candidate to modulate the interconnection between autophagy and ciliogenesis was AMP-activated protein kinase (AMPK).…”