Rodriguez-Irizarry Valerie J, Maples Robert W, Pfeiffer Julie K
Department of Microbiology, University of Texas Southwestern Medical Center, Dallas, TX 75390.
bioRxiv. 2025 Feb 26:2025.02.25.640062. doi: 10.1101/2025.02.25.640062.
Viruses encounter a range of selective pressures, but inefficiencies during replication can be masked. To uncover factors that limit viral replication, we used forward genetics to enrich for a murine norovirus (MNV) mutant with faster replication. We sequentially harvested the earliest progeny in cultured cells and identified a single amino acid change in the viral NS3 protein, K40R, that was sufficient to enhance replication speed. We found that the NS3-K40R virus induced earlier cell death and viral egress compared with wild-type virus. Mechanistically, NS3-K40R protein disrupted membranes more efficiently than wild-type NS3 protein, potentially contributing to increased mitochondrial dysfunction and cell death. Mice infected with NS3-K40R virus had increased titers, suggesting that increasing egress did not reduce fitness . Overall, by using a forward genetic approach, we identified a previously unknown inefficiency in norovirus egress and provide new insights into selective pressures that influence viral replication and evolution.
病毒会面临一系列选择压力，但复制过程中的低效率可能会被掩盖。为了揭示限制病毒复制的因素，我们采用正向遗传学方法富集复制速度更快的小鼠诺如病毒（MNV）突变体。我们在培养细胞中依次收获最早产生的子代，并在病毒NS3蛋白中鉴定出一个单一氨基酸变化，即K40R，这足以提高复制速度。我们发现，与野生型病毒相比，NS3-K40R病毒诱导细胞死亡和病毒释放的时间更早。从机制上讲，NS3-K40R蛋白比野生型NS3蛋白更有效地破坏细胞膜，这可能导致线粒体功能障碍和细胞死亡增加。感染NS3-K40R病毒的小鼠病毒滴度升高，这表明增加病毒释放并没有降低适应性。总体而言，通过正向遗传学方法，我们发现了诺如病毒释放过程中一个以前未知的低效率现象，并为影响病毒复制和进化的选择压力提供了新的见解。
