A novel tool for monitoring endogenous alpha-synuclein transcription by NanoLuciferase tag insertion at the 3' end using CRISPR-Cas9 genome editing technique

被引：17

作者：

Basu, Sambuddha ^{[1
]}

Adams, Levi ^{[1
]}

Guhathakurta, Subhrangshu ^{[1
]}

Kim, Yoon-Seong ^{[1
,2
]}

机构：

[1] Univ Cent Florida, Coll Med, Burnett Sch Biomed Sci, Div Neurosci, 6900 Lake Nona Blvd, Orlando, FL 32827 USA

[2] Kyunghee Univ Med Coll, Seoul, South Korea

来源：

SCIENTIFIC REPORTS | 2017年 / 7卷

关键词：

HISTONE DEACETYLASE INHIBITORS; LUCIFERASE REPORTER; DNA METHYLATION; MOLECULAR-MECHANISMS; GENE; EXPRESSION; 5-AZA-2'-DEOXYCYTIDINE; DUPLICATION; NEURONS;

D O I：

10.1038/srep45883

中图分类号：

O [数理科学和化学]; P [天文学、地球科学]; Q [生物科学]; N [自然科学总论];

学科分类号：

07 ; 0710 ; 09 ;

摘要：

alpha-synuclein (alpha-SYN) is a major pathologic contributor to Parkinson's disease (PD). Multiplication of alpha-SYN encoding gene (SNCA) is correlated with early onset of the disease underlining the significance of its transcriptional regulation. Thus, monitoring endogenous transcription of SNCA is of utmost importance to understand PD pathology. We developed a stable cell line expressing alpha-SYN endogenously tagged with NanoLuc luciferase reporter using CRISPR/Cas9-mediated genome editing. This allows efficient measurement of transcriptional activity of alpha-SYN in its native epigenetic landscape which is not achievable using exogenous transfection-based luciferase reporter assays. The NanoLuc activity faithfully monitored the transcriptional regulation of SNCA following treatment with different drugs known to regulate alpha-SYN expression; while exogenous promoter-reporter assays failed to reproduce the similar outcomes. To our knowledge, this is the first report showing endogenous monitoring of alpha-SYN transcription, thus making it an efficient drug screening tool that can be used for therapeutic intervention in PD.

引用

页数：11

共 37 条

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