Department of Microbiology & Immunology, College of Medicine and Health Sciences (CMHS), United Arab Emirates University (UAEU), Al Ain, United Arab Emirates.
Université de Strasbourg, CNRS, Architecture et Réactivité de l'ARN, UPR 9002, Strasbourg, France.
J Mol Biol. 2021 May 14;433(10):166923. doi: 10.1016/j.jmb.2021.166923. Epub 2021 Mar 11.
How retroviral Gag proteins recognize the packaging signals (Psi) on their genomic RNA (gRNA) is a key question that we addressed here using Mason-Pfizer monkey virus (MPMV) as a model system by combining band-shift assays and footprinting experiments. Our data show that Pr78 selects gRNA against spliced viral RNA by simultaneously binding to two single stranded loops on the MPMV Psi RNA: (1) a large purine loop (ssPurines), and (2) a loop which partially overlaps with a mostly base-paired purine repeat (bpPurines) and extends into a GU-rich binding motif. Importantly, this second Gag binding site is located immediately downstream of the major splice donor (mSD) and is thus absent from the spliced viral RNAs. Identifying elements crucial for MPMV gRNA packaging should help in understanding not only the mechanism of virion assembly by retroviruses, but also facilitate construction of safer retroviral vectors for human gene therapy.
逆转录病毒 Gag 蛋白如何识别其基因组 RNA（gRNA）上的包装信号（Psi）是一个关键问题，我们使用猕猴泡沫病毒（MPMV）作为模型系统，通过结合迁移率变动分析和足迹实验来解决这个问题。我们的数据表明，Pr78 通过同时结合 MPMV Psi RNA 上的两个单链环来选择 gRNA 对抗剪接的病毒 RNA：（1）一个大嘌呤环（ssPurines），和（2）一个与大部分碱基配对的嘌呤重复部分重叠并延伸到富含 GU 的结合基序的环。重要的是，第二个 Gag 结合位点位于主要剪接受体（mSD）的下游，因此不存在于剪接的病毒 RNA 中。鉴定对 MPMV gRNA 包装至关重要的元件不仅有助于理解逆转录病毒衣壳组装的机制，还有助于构建用于人类基因治疗的更安全的逆转录病毒载体。
