Fig. 6: HSV-1 infection regulates pre-tRNA expression and Ψ installation.

a Total read count and read length distribution per DRAP3R run using RA flowcells with RNA004 chemistry and a minimum read length cutoff of 20 nt. b Sequence reads were classified as Pol II transcribed (green) or Pol III transcribed with the latter subdivided into pre-tRNAs (dark blue), primary Pol III genes (orange), and pseudogenes (red), the latter defined by their annotation in Gencode v45. Short reads that could not be unambiguously assigned to a specific gene are shown in beige while reads that did not overlap with any existing annotation are shown in gold. Viral reads, derived from EBV present in CRO-AP5 cells are shown in purple (c–d) Volcano plots showing significantly (padj < 0.05) differentially regulated pre-tRNAs (dark blue), Pol II RNAs (green) and Pol III genes (orange) at (c) 6 hpi and (d) 12 hpi relative to mock-infected cells (n  =  2 biological replicates per condition / timepoint). Differential transcript expression was determined using DeSeq2 with shrunken LFC obtained using the adaptive shrinkage estimator (ashr) and adjusted p values derived using the Benjamini–Hochberg (BH) procedure. e–f Scatter plot comparing Ψ stoichiometries on pre-tRNAs shows decreasing correlation (Spearman’s r²) between uninfected and HSV-1 infected (e—6 hpi, f—12 hpi) ARPE-19 cells (n  =  2 biological replicates per condition / timepoint). Underlying data are additionally represented as bar and whisker plots (center =  median, box = 25th–75th percentiles, whiskers = values within 1.5 × IQR, outliers plotted individually) with p values calculated using Student’s paired T test (two-sided).