Identification of Cables1 as a critical host factor that promotes ALV-J replication via genome-wide CRISPR/Cas9 gene knockout screening

作者：Peng Liu, Jinghua Jiang, Yuntong Chen, Fei Gao, Suyan Wang, Mengmeng Yu, Yongzhen Liu, Ru Guo, Li Zhang, Zhuangzhuang Xu, Caiying Wang, Xiaole Qi, Yanping Zhang, Hongyu Cui, Yulu Duan, Sen Wu, Yulong Gao · 发表于：Journal of Biological Chemistry · 年份：2024 · DOI：10.1016/j.jbc.2024.107804 · 被引用次数：7 · 研究领域：Viral Infections and Immunology Research、CRISPR and Genetic Engineering、Cytomegalovirus and herpesvirus research

Avian leukosis virus subgroup J (ALV-J), a member of the genus Alpharetrovirus, possesses a small genome and exploits a vast array of host factors during its replication cycle. To identify host factors required for ALV-J replication and potentially guide the development of key therapeutic targets for ALV-J prevention, we employed a chicken genome-wide CRISPR/Cas9 knockout library to screen host factors involved in ALV-J infection within DF-1 cells. This screening revealed 42 host factors critical for ALV-J infection. Subsequent knockout assays showed that the absence of the genes encoding cycle-regulatory proteins, namely, Cables1, CDK1, and DHFR, significantly inhibited ALV-J replication. Notably, Cables1 knockout cell lines displayed the most pronounced inhibitory effect. Conversely, overexpression assays confirmed that Cables1 significantly promotes ALV-J replication. Immunoprecipitation assays further indicated that Cables1 specifically interacts with the viral protein p15 (viral protease) among all ALV-J proteins, enhancing ALV-J p15 polyubiquitination. Additionally, we identified 26 lysine residues of ALV-J p15 as key sites for ubiquitination, and their replacement with arginine attenuated the replication ability of ALV-J in both in vitro and in vivo assays. This study demonstrates that Cables1 is a critical replication-dependent host factor of ALV-J by enhancing p15 ubiquitination and thereby promoting viral replication. Overall, these findings contribute to a deeper u...