Recently, Prof. Song Baoliang, academician of Chinese Academy of Sciences and Assistant Professor Zhao Xiaolu, both from College of Life Sciences, Wuhan University, and Prof. Qiu Wenwei from School of Chemistry and Molecular Engineering, East China Normal University published a paper on Cell Research, which demonstrated that cholesterylation of SMO was an auto processing event enhanced by calcium ion, and revealed that calcium ion played an important role in Hedgehog signal transduction pathway and proved the significance of cholesterylation in SMO activation.
The paper is entitled Cholesterylation of Smoothened is a calcium-accelerated autoreaction involving an intramolecular ester intermediate, with Dr. Hu Ao from the Institute for Advanced Studies, Wuhan University and Zhang Jingzan, a PhD student from School of Chemistry and Molecular Engineering, East China Normal University as the co-first authors and Song Baoliang, Zhao Xiaolu and Qiu Wenwei as corresponding authors.
The Hedgehog (Hh) signaling pathway is a pathway from drosophila to mammals, whose signal transduction is mainly conducted by three proteins—secretory protein Hh, Hh receptor Patched1 (PTCH1) and Smoothened (SMO), a G-protein coupled receptor (GPCR). Hh signaling pathway plays an important role in governing embryonic development and postnatal tissue homeostasis whereas misactivated Hh signaling leads to certain cancers. When extracellular Hh potency increases, Hh will bind to PTCH1 and relieve the suppression on SMO, translocating SMO in intracellular vesicles to the primary cilium and SMO further transducting extracellular signals to intracellular areas to activate glioma-associated oncogene homolog (GLI) and other transcription factors.
Song Baoliang and Qiu Wenwei firstly reported that SMO protein was covalently modified by cholesterol in 2017. SMO protein forms ester bond through its extracellular the 95th aspartic acid residue (Asp95 or D95) in cysteine-richdomain (CRD) and 3β-hydroxyl in cholesterol. However, it remains unclear the chemical reaction and regulation mechanism of covalent modification of SMO by cholesterol.
Later, it was found that when cells were stimulated by Hh signaling, calcium ion concentration in SMO-localized endosomes would dramatically increase. Further filtration found that Hh signaling could mediate extracellular calcium ion influx through activating STIM-ORAI1 calcium ion channel.
To study the important role of cholesterylation for SMO activation, based on SMO structure, researchers established a SMO (D95E) mutant, which could still bind cholesterol but be not modified by cholesterol. For cells, the D95E mutant of SMO would dramatically decrease the activity in downstream Hh signaling pathway. The homozygous SmoD99E knock-in mice were embryonic lethal with severe developmental delay, demonstrating that lack of cholesterylation would totally suppress SMO activation.
This research received grants from the plan of Dynamic Modifications of Biomacromolecules and Chemical Interventions by the National Natural Science Foundation of China.
Link to the paper：
https://doi.org/10.1038/s41422-022-00622-0
