Fig. 1: HSV-1 infections generate Z-RNA.
From: Host cell Z-RNAs activate ZBP1 during virus infections
a,c,e, Z-NA accrual in HSV-1-infected (F strain; multiplicity of infection (MOI) = 5) (gB+) Flag–ZBP1 MEFs. a, Time course of Z-NA formation. c,e, MEFs were exposed to the indicated nucleases after fixation (c) or were treated with ActD 1 h post-infection (h.p.i.) (e), before fixation at 9 h.p.i. b,d,f, The fluorescence intensity (arbitrary units, a.u.) of the Z-NA signal in a, c and e, respectively. g, Cell death kinetics of empty vector (Vec) or Flag–ZBP1 MEFs (Flag–ZBP1) infected with HSV-1 ICP6 RHIMmut virus (MOI = 5) in the presence or absence of ActD. h, The genomic distribution of viral RNAs enriched in anti-Flag RIP–seq from HSV-1 infected Flag–ZBP1 MEFs. The outer and inner sectors represent genomic annotation and scaled fold-change values on the forward and reverse strands, respectively. The black caps denote the 5′ ends. i, The estimated proportion of canonical and uncharacterized viral RNA (vRNA) in anti-Flag RIP–seq data. j,k, Exemplar Z-RNA forming viral transcripts for canonical vRNA (j) and uncharacterized vRNA (k). qPCR analysis of the indicated viral transcripts after Z22 RIP analysis of WT HSV-1-infected Flag–ZBP1 MEFs (left). Putative Z-RNA-forming regions within exemplar transcripts (right). Each arc denotes a base-pairing event and red arcs show Z-prone stems (enumerated above the MFE Circos plots). Exemplar Z-RNAs (black arrowheads) and potential Zα-binding sites (pink boxes) are detailed at the bottom. l, qPCR analysis of viral RNAs after Flag RIP from either Flag–ZBP1 or Flag–ZBP1(ΔZα) mutant MEFs infected with HSV-1. Data are mean ± s.d. n = 30 cells per group (b, d and f), and n = 4 (g and i) and n = 3 (j–l) biologically independent samples. Statistical analysis was performed using one-way analysis of variance (ANOVA) with Dunnett’s multiple-comparison test (d, f and l), two-way ANOVA (g) and two-tailed unpaired t-tests with Welch’s correction (j and k). ***P < 0.0005 (except for in d, f, g and l, for which P < 0.0001). Data are representative of at least two (a, c and e) or three (g) independent experiments. Scale bars, 10 μm (a, c and e (right)) and 20 μm (e (left and middle)).
