https://doi.org/10.4081/ejh.2023.3594

Phosphorylation mutation impairs the promoting effect of spastin on neurite outgrowth without affecting its microtubule severing ability

Vol. 67 No. 1 (2023)
Submitted: 4 November 2022
Accepted: 27 December 2022
Published: 12 January 2023
Spastin, phosphorylation, hippocampal neurons, COS7, microtubule severing, neurite outgrowth
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Authors

Yunlong Zhang
Department of Orthopaedics, The First Affiliated Hospital of Jinan University, Guangzhou, China.
Xin He
https://orcid.org/0000-0002-0861-2170
Clinical Laboratory Center, The First Affiliated Hospital of Jinan University, Guangzhou, China.
Jianyu Zou
Department of Orthopaedics, The First Affiliated Hospital of Jinan University, Guangzhou, China.
Jie Yang
Department of Orthopaedics, The First Affiliated Hospital of Jinan University, Guangzhou, China.
Ao Ma
https://orcid.org/0000-0002-3032-8918
Minghui Tan
tanminghui@jnu.edu.cn
https://orcid.org/0000-0001-6260-4115
Department of Orthopaedics, The First Affiliated Hospital of Jinan University, Guangzhou, China.

Spastin, a microtubule-severing enzyme, is known to be important for neurite outgrowth. However, the role of spastin post-translational modification, particularly its phosphorylation regulation in neuronal outgrowth, remains unclear. This study aimed to investigate the effects of eliminating spastin phosphorylation on the neurite outgrowth of rat hippocampal neurons. To accomplish this, we constructed a spastin mutant with eleven potential phosphorylation sites mutated to alanine. The phosphorylation levels of the wildtype spastin (WT) and the mutant (11A) were then detected using Phos-tag SDS-PAGE. The spastin constructs were transfected into COS7 cells for the observation of microtubule severing, and into rat hippocampal neurons for the detection of neuronal outgrowth. The results showed that compared to the spastin WT, the phosphorylation levels were significantly reduced in the spastin 11A mutant. The spastin mutant 11A impaired its ability to promote neurite length, branching, and complexity in hippocampal neurons, but did not affect its ability to sever microtubules in COS7 cells. In conclusion, the data suggest that mutations at multiple phosphorylation sites of spastin do not impair its microtubule cleavage ability in COS7 cells, but reduce its ability to promote neurite outgrowth in rat hippocampal neurons.

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Ethics Approval

All animal experiments were conducted in accordance with the Regulations of the People's Republic of China on the Administration of Laboratory Animals and were reviewed and approved by the Ethics Committee of Jinan University

Supporting Agencies

National Natural Science Foundation of China

How to Cite

Zhang, Y., He, X., Zou, J., Yang, J., Ma, A., & Tan, M. (2023). Phosphorylation mutation impairs the promoting effect of spastin on neurite outgrowth without affecting its microtubule severing ability. European Journal of Histochemistry, 67(1). https://doi.org/10.4081/ejh.2023.3594
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