Fig. 4: Comparison of SARS-CoV-2’s structure in virions and host cells.

a Schematic diagram showing the life cycle of SARS-CoV-2. b Single-stranded nucleotides (n = 8269) identified by vRIC-seq in virions have higher SHAPE-MaP reactivities than base-paired nucleotides (n = 19055) in cells. P-value was determined by two-tailed, unpaired Student’s t-test. The center line of the box plot represents the median, the box borders represent the first (Q1) and third (Q3) quartiles, and the whiskers are the most extreme data points within 1.5× the interquartile range (from Q1 to Q3). c Venn diagram showing the overlap between duplexes revealed in virions and cells. d The percentage of in-virion interactions supported by the COMRADES chimeric reads is decreased along with the spanning distance. The control datasets are randomly selected pairwise loci (repeated 1000 times, n = 1000). Data are mean ± s.e.m., ***P < 0.001, one-sided permutation test. e The positive predictive value (PPV) of the in-virion duplexes compared to the in-cell predicted duplexes along the SARS-CoV-2 genome in sliding 1 kb windows. The dashed line indicates the average percentage. f In-virion (top) and in-cell (bottom) duplexes in the FSE surrounded region. Arc lines colored in gray indicate base pairs spanning more than 500 nt, arc lines in red indicate base pairs shared by duplexes in virions and cells, while arc lines in blue indicate base pairs specific in virions or cells. g Diagram the pseudoknot structure of FSE. The dashed lines represent base pairs in Stem 2. Different colors in bases stand for the strength of vRIC signals. The scale is shown at the right bottom. An alternative duplex can form at the gray shadowed regions. h vRIC-seq data preferably supports an elongated duplex in the FSE region. i In-virion (top) and in-cell (bottom) duplexes in 3′ UTR surrounded region. j Dual-Luciferase Reporters for functional characterization of the alternative duplex. Before the slippery site, a 33 nt sequence (orange) can form a duplex with the Stem 1 (green) of FSE-PK. In-frame is denoted as “0 frame”, while “−1 frame” stands for programmed −1 ribosomal frameshift (−1 PRF). The dashed orange lines stand for the deleted 33 nt sequence. Ctrl control, WT wild type, Del deletion, Rluc renilla luciferase, Fluc firefly luciferase. k The alternative duplex can stimulate −1 PRF activity. Data are mean ± s.e.m., P-value was determined by a two-tailed, unpaired Student’s t-test (n = 6 biologically independent experiments). Source data are provided as a Source Data file.