德隆
生物
清脆的
细胞生物学
Cas9
生殖系
秀丽隐杆线虫
功能(生物学)
拟南芥
遗传学
突变体
计算生物学
基因
泛素
泛素连接酶
作者
Liangyu Zhang,Jordan D. Ward,Ze Cheng,Abby F. Dernburg
出处
期刊:Development
[The Company of Biologists]
日期:2015-01-01
被引量:572
摘要
Experimental manipulation of protein abundance in living cells or organisms is an essential strategy for investigation of biological regulatory mechanisms. While powerful techniques for protein expression have been developed in C. elegans, existing tools for conditional disruption of protein function are far more limited. To address this, we have adapted the auxin-inducible degradation (AID) system discovered in plants to enable conditional protein depletion in C. elegans. We report that expression of a modified Arabidopsis TIR1 F-box protein mediates robust auxin-dependent depletion of degron-tagged targets. We document the effectiveness of this system for depletion of nuclear and cytoplasmic proteins in diverse somatic and germline tissues throughout development. Target proteins were depleted in as little as 20-30 minutes, and their expression could be reestablished upon auxin removal. We have engineered strains expressing TIR1 under the control of various promoter and 3' UTR sequences to drive tissue-specific or temporally regulated expression. The degron tag can be efficiently introduced by CRISPR/Cas9-based genome editing. We have harnessed this system to explore the roles of dynamically expressed nuclear hormone receptors in molting, and to analyze meiosis-specific roles for proteins required for germline proliferation. Together, our results demonstrate that the AID system provides a powerful new tool for spatiotemporal regulation and analysis of protein function in a metazoan model organism.
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