| dc.description.abstract | Single-stranded RNA viruses (ssRNAs) are a group of contagious microbes that have compact structures, containing a protein capsid and a genomic RNA (gRNA) packaged inside. Genome packaging is an essential part of the viral infectious cycle, yet this step is not fully understood. To know the packaging of ssRNA viruses, having a gRNA structure is needed. In this work, we combined cryo-electron microscopy (cryoEM) and molecule modeling to determine the structures of packaged gRNAs. Compared to the capsid, the gRNA is structurally flexible, limiting the resolution of its structure solved by cryo-EM. To efficiently refine RNA structures into middleresolution cryo-EM maps, we developed the Hierarchical Natural Move Monte Carlo (HNMMC) simulation, which takes advantage of collective motions for groups of nucleic acid residues to reduce the computational resources and also preserve atomic details in the models. After successfully refining several cryo-EM models, we used HNMMC simulation to model atomic details and flexibility for two distinct conformations of the gRNA packaged inside the bacteriophage MS2. We also presented the complete gRNA model of the bacteriophage Qβ revealing that the 3’ untranslated region binds to the maturation protein and the 4,127 nucleotides fold domain-by-domain. 33 operator-like RNA stem-loops serving as package signals are located, suggesting a pathway for the assembly of Qβ virion. Besides, from overexpressing the Qβ capsid proteins or through the infection by wild-type Qβ phages, we have discovered various forms of the virus-like particles (VLPs), including the regular T=3 icosahedral, larger T=4 icosahedral, prolate, oblate forms, and a small prolate form elongated along the 3-fold axis. Overexpressing the shorter RNA fragments increased the percentages of the smaller VLPs, which may be sufficient to encapsidate a shorter RNA. With the help of cryo-EM and molecule modeling, the structure of a packaged gRNA can be solved and the interaction between the capsid protein and the RNA can also be revealed. These techniques provide a new direction to study the viral RNA packaging in structural virology. Understanding the packaging of ssRNA viruses has an implication of using them for biotechnology, such as RNA perseverance and delivery. | en |
