2. Academic Validation
  3. Cholesterylation of Smoothened is a calcium-accelerated autoreaction involving an intramolecular ester intermediate

Cholesterylation of Smoothened is a calcium-accelerated autoreaction involving an intramolecular ester intermediate

  • Cell Res. 2022 Mar;32(3):288-301. doi: 10.1038/s41422-022-00622-0.
Ao Hu  # 1,  Jing-Zan Zhang  # 2,  Jie Wang 2,  Chen-Chen Li 2,  Meng Yuan 1,  Gang Deng 1,  Zi-Cun Lin 1,  Zhi-Ping Qiu 1,  Hu-Yue Liu 1,  Xian-Wei Wang 2 3,  Peng-Cheng Wei 1,  Xiao He 2 4,  Xiaolu Zhao 5,  Wen-Wei Qiu 6,  Bao-Liang Song 7
Affiliations

Affiliations

  • 1 The Institute for Advanced Studies, Hubei Key Laboratory of Cell Homeostasis, College of Life Sciences, TaiKang Center for Life and Medical Sciences, TaiKang Medical School, Wuhan University, Wuhan, Hubei, China.
  • 2 Shanghai Engineering Research Center of Molecular Therapeutics and New Drug Development, School of Chemistry and Molecular Engineering, East China Normal University, Shanghai, China.
  • 3 College of Science, Zhejiang University of Technology, Hangzhou, Zhejiang, China.
  • 4 NYU-ECNU Center for Computational Chemistry at NYU Shanghai, Shanghai, China.
  • 5 The Institute for Advanced Studies, Hubei Key Laboratory of Cell Homeostasis, College of Life Sciences, TaiKang Center for Life and Medical Sciences, TaiKang Medical School, Wuhan University, Wuhan, Hubei, China. [email protected].
  • 6 Shanghai Engineering Research Center of Molecular Therapeutics and New Drug Development, School of Chemistry and Molecular Engineering, East China Normal University, Shanghai, China. [email protected].
  • 7 The Institute for Advanced Studies, Hubei Key Laboratory of Cell Homeostasis, College of Life Sciences, TaiKang Center for Life and Medical Sciences, TaiKang Medical School, Wuhan University, Wuhan, Hubei, China. [email protected].
  • # Contributed equally.
PMID: 35121857 DOI: 10.1038/s41422-022-00622-0
Abstract

Hedgehog (Hh) is a morphogen that binds to its receptor Patched 1 and activates Smoothened (Smo), thereby governing embryonic development and postnatal tissue homeostasis. Cholesterol can bind and covalently conjugate to the luminal cysteine-rich domain (CRD) of human Smo at the D95 residue (D99 in mouse). The reaction mechanism and biological function of Smo cholesterylation have not been elucidated. Here, we show that the SMO-CRD undergoes auto-cholesterylation which is boosted by calcium and involves an intramolecular ester intermediate. In cells, Hh stimulation elevates local calcium concentration in the SMO-localized endosomes through store-operated calcium entry. In addition, we identify the signaling-incompetent Smo D95E mutation, and the D95E mutant Smo can bind Cholesterol but cannot be modified or activated by Cholesterol. The homozygous SmoD99E/D99E knockin mice are embryonic lethal with severe developmental delay, demonstrating that cholesterylation of CRD is required for full-length Smo activation. Our work reveals the unique autocatalytic mechanism of Smo cholesterylation and an unprecedented role of calcium in Hh signaling.

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