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. Author manuscript; available in PMC: 2020 Sep 19.
Published in final edited form as: Mol Cell. 2019 Sep 4;75(6):1218–1228.e6. doi: 10.1016/j.molcel.2019.07.027

Figure 3. Analysis of ribosomes during 2–5AMD.

Figure 3.

(A) RNase L cleavage positions in 18S and 28S rRNA found by RtcB RNA-seq. The Y-axis provides a unified metric of read abundance and cleavage induction strength. (B) Mapping the RNase L sites onto 3D structures of 18S and 28S rRNAs. Structures are colored by cleavage strength as defined in (A). (C) Status of rRNA and translation activity of purified ribosomes obtained from rabbit reticulocyte lysate (RRL) or A549 cells. A titration with purified ribosomes was done in the presence of 50 ng capped luciferase mRNA. New translation was measured by luminescence. (D) Comparison of translation loss and rRNA cleavage over the duration of dsRNA response. Fraction of intact rRNA observed by NanoChip was quantified in GelQuant.NET. New translation relative to the untreated condition was measured by 35S metabolic labeling and ribopuromycilation (Fig. 1C; S1A). Kinetic parameters of the time profiles are shown below the graph. See also Figure S3, Table S2.