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. 2022 Jan 20;50(3):1583–1600. doi: 10.1093/nar/gkac001

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© The Author(s) 2022. Published by Oxford University Press on behalf of Nucleic Acids Research.

This is an Open Access article distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0/), which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited.

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Figure 5. — 3′ end trimming in EXOSC3 depletion conditions. (A) Metagene profiles of regions around SA sites of intron-hosted snoRNAs, showing log₂FC in single nucleotide resolution between pA⁺ and pA^+/− 3′ end sequencing data of newly synthesized RNA upon RNAi- or 6 h AID-mediated ZCCHC8 and EXOSC3 depletion relative to siEGFP or unedited HeLa:TIR1, 0 h IAA controls. Regions are displayed as in Figure 4C. (B) Genome browser views as in Figure 3D, but showing regions around selected SAs of snoRNA-hosting introns. Shown data are from AID- (0 and 6 h) or RNAi-mediated ZCCHC8 and EXOSC3 depletion conditions. (C) Trimming assay based on RT-qPCR analysis of selected amplicons (gray areas and ‘qPCR amplicon’ annotations in genome browser view) within 3′ extended snoRNAs. RT-qPCR analyses were conducted on total RNA isolated from EXOSC3-mAID (purple) and ZCCHC8-mAID (green) cells treated with siRNAs against EGFP, EXOSC10 or PARN and followed by 0 h (−) or 6 h (+) exposure to IAA. Data were normalized to Rplp0 mRNA (RPO) and displayed as the trimming factor, obtained from the ratio of distal/proximal amplicon values. Differences within RNAi treatment groups, comparing +/− IAA samples, were determined by a two-way ANOVA test [*P_adj ≤ 0.05, **P_adj ≤ 0.01, ***P_adj ≤ 0.001, ^****P_adj ≤ 0.0001, nonsignificant (ns) = P_adj > 0.05].