On December 30, 2019, the research group of Prof. WANG Chao have revealed the molecular mechanism by which the Ankyrin-G/Ndel1 complex in the axon initial segment (AIS) of neurons maintains the axonal polarity through regulating cargo sorting. The achievement was obtained via multiple study methods of biochemistry, structural biology, chemical biology and molecular neurocell biology. The result, their another study about Ankyrin protein, was published online on PNAS, following their work on the identification of the peptide regulating cell autophagy published in Nat ChemBiol in 2018.
The nervous system is the most important functional regulation system in the human body. As the basic structure and functional unit of the nervous system, neurons are highly polarized cells. Typical neurons are composed of dendrites which receive signals, soma, and axons that transmit signals. Neuron polarity is maintained owing to the diverse morphologies and unique physiologic functions of different dendrites, soma and axons which are composed of variable substances.
AIS is a specific membrane structure region where the near end of axon is close to the cell body. It is rich in voltage-gated sodium channels (Nav channel), voltage-gated potassium channels (KCNQ2/3) and cell adhesion molecules (Nfasc, NrCAM, etc.), as well as a variety of scaffold proteins and cytoskeleton proteins, so is of great significance for neurons to integrate signals from soma and dendrites, and regulate and generate action potentials. As the most important protein complex organizer in AIS, the neural scaffold protein Ankyrin-G (AnkG) is a hallmark protein of AIS. AnkG anchors a variety of ion channels and cell adhesion molecules to the cell membrane, thereby ensuring the integrity and stability of the structure and function of AIS. At the meantime, the special status of the AIS allows AnkG to also play a role as a specific gatekeeper by selectively regulating the proteins and specific organelles that enter the axon, therefore maintaining the axon-dendritic polarity of neurons. However, the molecular basis for this selective regulation of substances in and out of axons had remained elusive.
The structure of AnkB/Ndel1 bind
In this study, Prof. WANG Chao's research group determined the direct interaction between AnkG and the dynein regulator protein Ndel1 through in vitro biochemical experiments, and used ITC for quantitative binding capacity testing. By comparing the dissociation constant of the binding between the two proteins, the assembly of the complex was analyzed systematically at the biochemical level. Then they found that AnkG's homolog, AnkB, could also form a complex with Ndel1, and analyzed the high-resolution crystal structure of the AnkB/Ndel1 complex, which helped to illustrat the molecular mechanism of binding between AnkG/AnkB and Ndel1. In the following cell biology studies on the AnkG/Ndel1 complex in mouse hippocampal neurons, they found that the AnkG/Ndel1 complex can selectively prevent the cargo that does not initially belong to the axon entering at AIS, and thus maintaining the polarity of the neurons. Furthermore, cargo sorting can be disrupted if the AnkG/Ndel1 complex formation is blocked. This research work provides a structural basis for understanding the organizational structure of the AIS and the maintenance of axonal polarity. Besides, it offers a theoretical basis for the study of the pathogenesis of human mental system diseases caused by corresponding gene mutations.
YE Jin, a Ph.D student of the School of Life Sciences of the University of Science and Technology of China, and Prof. LI Jianchao of the South China University of Technology School of Medicine are the co-first authors of the paper, with Prof. WANG Chao from the School of Life Sciences of the University of Science and Technology of China be the communication author. This work was also greatly assisted by Academician ZHANG Mingjie's research group of the Hong Kong University of Science and Technology and the Prof. ZHANG Yan's research group of the School of Life Sciences of Peking University. This research was supported by the Ministry of Science and Technology's key R & D plan, the National Natural Science Foundation of China, the Hefei Material Science and Technology Center orientation training project, the basic scientific research business expenses of the central university, and the first action of the Chinese Academy of Sciences.