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在人类细胞中使用菌体抗CRISPR蛋白质降解Cas9

Zuriah Meacham¹, Luisa Arake de Tacca¹, Joseph Bondy-Denomy²

¹Acrigen Biosciences, Inc., Alameda, California, United States of America.

|December 8, 2023

概括

在人类细胞中,像AcrIIA1这样的抗CRISPR (Acr) 蛋白质会降解CRISPR-Cas核酶,如Cas9. 这一发现为精确的基因编辑疗法提供了新的工具.

科学领域:

分子生物学分子生物学
遗传学是一个遗传学.
微生物学微生物学

背景情况:

菌体利用抗CRISPR (Acr) 蛋白质使细菌的CRISPR-Cas免疫系统无法成功复制.
在ACR蛋白中,使用多种机制来抑制CRISPR-Cas系统.
已知Streptococcus pyogenes Cas9 (SpyCas9) 抑制剂AcrIIA1被各种菌体和等离子体编码.

研究的目的:

为了研究人类细胞中CRISPR-Cas系统AcrIIA1抑制的机制.
为了确定AcrIIA1是否可以诱导Cas9核酶的降解.
探索AcrIIA1介导的Cas9降解的治疗潜力.

主要方法:

进行了人类细胞培养实验,观察AcrIIA1与SpyCas9和金黄色葡萄球菌Cas9 (SauCas9) 的相互作用.
用AcrIIA1的变异来确定负责Cas9降解的关键相互作用域.
在存在 AcrIIA1.1 的情况下,评估了 CRISPR-Cas9 核酶活性.

主要成果:

观察到AcrIIA1诱导人体细胞中SpyCas9和SauCas9的降解,这是Acr蛋白的新发现.
突变破坏了AcrIIA1和Cas9之间的物理相互作用,取消了降解效应.
这表明,Cas9降解的机制取决于直接的蛋白质与蛋白质相互作用.

结论:

通过直接相互作用,AcrIIA1在人类细胞中积极降解Cas9核酶.
由AcrIIA1对Cas9的有针对性的降解为调节基因编辑活动提供了一种新的策略.
AcrIIA1的特性可以使CRISPR-Cas蛋白解向仿真体 (PROTACs) 的发展成为更安全的基因编辑应用.

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