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Assembly and gating mechanism of native AMPA receptors from the cerebellum

Xiaojing Li^1,2,3,† , Renjie Li^1,2,3,† , Yiqing Wei^1,2,3,† , Jiexin Chen^4,† , Jiaojiao Zhao^5,† , Jun Zhao^6,† , Wei Wang⁴ , Na Li⁴ , Lili Wang⁴ , Tuo Hu⁷ , Yanli Dong⁸ , Yongping Zhu⁹ , Chao Wei² , Long Li² , Wei Zhang^5,* , Zhuo Huang^4,10,* , Yan Zhao^1,2,*

¹Key Laboratory of Biomacromolecules (CAS), National Laboratory of Biomacromolecules, CAS Center for Excellence in Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing, China
²State Key Laboratory of Cognitive Science and Mental Health, Institute of Biophysics, Chinese Academy of Sciences, Beijing, China
³College of Life Sciences, University of Chinese Academy of Sciences, Beijing, China
⁴State Key Laboratory of Natural and Biomimetic Drugs, Department of Molecular and Cellular Pharmacology, School of Pharmaceutical Sciences, Peking University Health Science Center, Beijing, China
⁵Department of Pharmacology of Chinese Materia Medica, Institution of Chinese Integrative Medicine, The Key Laboratory of Tranquilizing TCM, Hebei Provincial Administration of Traditional Chinese Medicine, The Key Laboratory of Neural and Vascular Biology, Ministry of Education, Hebei Medical University, Shijiazhuang, Hebei, China
⁶Peking University Institute of Advanced Agricultural Sciences, Shandong Laboratory of Advanced Agricultural Sciences at Weifang, Weifang, Shandong, China
⁷State Key Laboratory of Natural Medicines, School of Pharmacy, China Pharmaceutical University, Nanjing, Jiangsu, China
⁸School of Biological Sciences and Technology, Beijing Forestry University, Beijing, China
⁹State Key Laboratory for Quality Ensurance and Sustainable Use of Dao-di Herbs, Artemisinin Research Center, and Institute of Chinese Materia Medica, China Academy of Chinese Medical Sciences, Beijing, China
¹⁰IDG/ McGovern Institute for Brain Research, Peking University, Beijing, China
^†These authors contributed equally: Xiaojing Li, Renjie Li, Yiqing Wei, Jiexin Chen, Jiaojiao Zhao, Jun Zhao
* Correspondence: Wei Zhang(weizhang@hebmu.edu.cn)Zhuo Huang(huangz@hsc.pku.edu.cn)Yan Zhao(zhaoy@ibp.ac.cn)

AMPA receptors (AMPARs) mediate the majority of fast excitatory synaptic transmission throughout the central nervous system. Calcium-permeable AMPARs and GluA4-containing receptors are critical for cerebellar functions, such as motor learning, associative memory, auditory processing, and synaptic plasticity. In contrast to the well-characterized, predominantly GluA2-containing AMPARs of the hippocampus and cortex, cerebellar AMPARs contain a higher proportion of GluA4 and remain poorly understood. Here, we generated a highly GluA4-specific antibody. Using this antibody in combination with antibodies specifically recognizing GluA1 and GluA2, we purified native AMPARs and determined the subunit compositions of both calcium-impermeable and calcium-permeable native AMPARs in the cerebellum. The isolated cerebellar AMPARs that contained both GluA1 and GluA4 were calcium-permeable, with GluA4 occupying mainly the B/D positions, GluA1 occupying the A/C positions, and the complex associated primarily with cornichon 3 (CNIH3). We determined the structures of the complex in distinct functional states, including the resting, active, and desensitized states, and characterized the conformational transitions that underlie its activity. During desensitization, the receptor adopts a pseudo-4-fold configuration of the ligand-binding domain layer, which may be important for its functional properties. This study provides a blueprint for the subunit compositions of AMPARs in the cerebellum and clarifies the gating mechanism of the calcium-permeable native AMPARA1A4–CNIH3 complex, providing significant insight into AMPAR-mediated synaptic transmission in the cerebellum.

https://doi.org/10.1038/s41422-026-01234-8

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