RT Journal Article
SR Electronic
T1 Intrinsic RNA targeting constrains the utility of CRISPR-Cas13 systems
JF bioRxiv
FD Cold Spring Harbor Laboratory
SP 2022.05.14.491940
DO 10.1101/2022.05.14.491940
A1 Zexu Li
A1 Zihan Li
A1 Xiaolong Cheng
A1 Xiaofeng Wang
A1 Shixin Ma
A1 Shengnan Wang
A1 Zhiyan Lu
A1 Han Zhang
A1 Wenchang Zhao
A1 Zhisong Chen
A1 Yingjia Yao
A1 Lumen Chao
A1 Wei Li
A1 Teng Fei
YR 2022
UL http://biorxiv.org/content/early/2022/05/14/2022.05.14.491940.abstract
AB CRISPR-Cas13 systems have been adapted as versatile toolkits for RNA-related applications. Here we systematically evaluate the performance of several popular Cas13 family effectors (Cas13a, Cas13b and Cas13d) under lentiviral vectors and reveal surprisingly differential defects and characteristics of these systems. Using RNA immunoprecipitation sequencing, transcriptome profiling, biochemistry analysis, high-throughput CRISPR-Cas13 screening and machine learning approaches, we determine that each Cas13 system has its intrinsic RNA targets in mammalian cells. Viral process-related host genes can be targeted by Cas13 and affect production of fertile lentiviral particles, thereby restricting the utility of lentiviral Cas13 systems. Multiple RNase activities of Cas13 are involved in endogenous RNA targeting. Unlike target-induced nonspecific collateral effect, intrinsic RNA cleavage can be specific, target-independent and dynamically tuned by varied states of Cas13 nucleases. Our work provides guidance on appropriate use of lentiviral Cas13 systems and further raises cautions about intrinsic RNA targeting during Cas13-based basic and therapeutic applications.Competing Interest StatementW.L. is a paid consultant to Tavros Therapeutics, Inc. Others declared no competing interests.