Open AccessDifferent fusion tags affect the activity of ubiquitin overexpression on spastin protein stability
Author(s) -
Jianyu Zou,
Zhen Cai,
Zhi Liu,
Yubin Liang,
Guowei Zhang,
Jie Yang,
Yunlong Zhang,
Hongsheng Lin,
Minghui Tan
Publication year - 2021
Publication title -
european journal of histochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.754
H-Index - 42
eISSN - 2038-8306
pISSN - 1121-760X
DOI - 10.4081/ejh.2021.3352
Subject(s) - hereditary spastic paraplegia , ubiquitin , microbiology and biotechnology , mcherry , neurite , fusion protein , microtubule , chemistry , green fluorescent protein , biology , in vitro , biochemistry , phenotype , gene , recombinant dna
Spastin is one of the proteins which lead to hereditary spastic paraplegia (HSP), whose dysfunction towards microtubule severing and membrane transporting is critically important. The present study is to elucidate the mechanisms of the protein stability regulation of spastin. The ubiquitin encoding plasmids are transfected into COS-7 cells with different fusion tags including Green Fluorescent Protein (GFP), mCherry and Flag. The expression level of spastin was detected, microtubule severing activity and neurite outgrowth were quantified. The data showed that ubiquitin overexpression significantly induced the decreased expression of spastin, suppressed the activity of microtubule severing in COS-7 cells and inhibited the promoting effect on neurite outgrowth in cultured hippocampal neurons. Furthermore, when modulating the overexpression experiments of ubiquitin, it was found that relatively small tag like Flag, but not large tags such as GFP or mCherry fused with ubiquitin, retained the activity on spastin stability. The present study investigated the effects of small/large tags addition to ubiquitin and the novel mechanisms of post-transcriptional modifications of spastin on regulating neurite outgrowth, in the attempt to experimentally elucidate the mechanisms that control the level or stability of spastin in hereditary spastic paraplegia.