Fig. 5: Accumulation of histone mRNAs in cells lacking RNases P and MRP.

A qPCR analysis of indicated histone mRNAs in control or dTAG-treated C40 cells (n = 3 biological replicates). Mean ± SD. *, p < 0.05; **, p < 0.01 (two-tailed Student’s t test). Exact p-values in the Source Data file. B qPCR of histone mRNAs in Dox-inducible C40 cells treated with DMSO or dTAG, with or without Dox (n = 3 biological replicates). Mean ± SD. *, p < 0.05; **, p < 0.01 (two-tailed Student’s t test). Exact p-values in the Source Data file. See also Fig. 2F, G and Supplementary Fig. 2D. C Representative images of control and dTAG-treated C40 cells stained for DNA (blue) and histone H3 and H4 mRNAs (red). Arrows point to cytoplasmic accumulations of histone mRNAs. Scale bar, 20 μm. D Northern analysis of histone mRNAs in C40 cells treated with DMSO or dTAG for 24 h. SNORD3A serves as a loading control (n = 3). The H2A probes recognize multiple histone genes in the H2A cluster. E Schematic of a histone mRNP with conserved 3′ stemloop bound by SLBP and ERI1 (top). ERI1 trims 1–2 nucleotides, which are restored by uridylation (green and orange U’s), producing three distinct 3′ ends (bottom). Created in BioRender. Murn, J. (2025) https://BioRender.com/1dxr6z2. F EnD-seq of H2AC13 mRNA from C40 cells after dTAG treatment. x-axis: last templated nucleotide; y-axis: normalized read counts at each position (CPM, counts per million). Colors indicate nontemplated tail lengths. The sequence below each plot indicates processed histone H2AC13 mRNA 3′ end formed in the nucleus; two nucleotides are subsequently trimmed to yield cytoplasmic histone mRNA (n = 3). G Heatmap summarizing EnD-seq data. Fold changes in nontemplated 3′ terminal nucleotide content for dTAG-treated (3 h and 24 h) versus DMSO-treated control (0 h) conditions are shown for each analyzed histone mRNA. Nontemplated nucleotides were > 99% uridines. Source data are provided as a Source Data file.