Genome-wide Perturbation Analysis Screening System for Exosomes-Related Genes Based
Release time：
2024-07-12
Source：
https://doi.org/10.1101/2023.09.07.556640
Author：Lu Lu1#, Jianxin Yin1#, Ning Chen1#, Xiaofei Guo1#, Chunle Han1 , Miao Wang1 , Huanqing Du1 , Huifang Li1 , Xingang Pan1 , Mengya Gao1 , Na Wang1 , Dongli Qi1 , Jianhong Wang1 , Fengwei Dong1 ，Tianshi Li2 , Xiaohu Ge1*
Release date：2023-09-08
ABSTRACT：As reported previously, exosomes have significant impacts on the physiological and pathological state in vivo. Exosomes have been extensively studied as a drug delivery carrier and some exosomes drugs have been undergoing clinical research. The mechanisms of exosome production, transport, and secretion remain to be studied in depth. Thus, here, we developed a novel CRISPR UMI-based gene screening system, with which we can reveal the exosomes biogenesis, transport, and uptake mechanisms on a genome-wide scale. This system consists of two parts: one is a gene knockout component; the other part is a unique molecular identifiers (UMI) labeling component t hat can label the exosomes produced by the knockout cells, in which each sgRNA corresponds to a specific UMI. In this way, by detecting the UMI loaded in the exosomes, we can trace the knockout gene. In this study, we first verified the function of each component using plasmids and lentiviruses respectively. Next, we simulated the infection of cells with lentiviral libraries using a single lentivirus to validate the functionality of the screening system. Finally, we constructed a CRISPR-UMI-based library targeting 15 genes (genes with clear effects on exosomes biogenesis) to further validate the reliability of the screening system. The development of this screening system is of indispensable importance for the in-depth study of the mechanisms of exosome production and secretion, as well as for the improvement of exosomes production and the
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