一种光控神经退行性相关蛋白p62聚集的研究模型构建

Objective: To Construct a light control model of ALS/FTD (amyotrophic lateral sclerosis/frontotemporal dementia)-associated protein p62, and achieve manipulation of p62 phase transition in multiple cell lines. Methods: PB1 domain of p62 was replaced with CRY2-mCherry by homologous recombination. pCMV-CRY2-mCherry plasmid template were linearized and the p62 Δ1-122 sequence were amplified using PCR. Homologous recombination reactions were performed after purification and the product were sequenced to verify the successful of pCMV-CRY2-mCherry-p62 Δ1-122（opto-p62）construct. Opto-p62 was expressed in HEK 293 cells and SH-SY5Y cells, and were stimulated by blue light to form light-induced p62 bodies. The properties and factors affecting phase transition ability of opto-p62 were investigated.Results: The p62 bodies were liquid-like. Opto-p62 overexpressed in HEK 293 cells and SH-SY5Y cells can undergo phase transition to form light-induced p62 bodies under blue light stimulation at a wavelength of 488 nm. Opto-p62 phase transition ability was positively correlated with protein expression intensity and blue light intensity, and the light-induced p62 bodies can recruit p62-related proteins.