Figure 3. MTS chemistry reveals fast- and slow-turnover miRNAs in miRNA RATE-seq experiments.

(A) Schematic of s⁴U treatments used in miRNA RATE-seq. (B) Cartoon of anticipated behavior of fast-turnover and slow-turnover miRNAs in comparison to average. Fast-turnover miRNAs are expected to be over-represented in the early time points whereas slow-turnover miRNAs are depleted, relative to steady state (ss). (C) Heatmap depicting correlation coefficients (Pearson’s r) between miRNA levels at different times after s⁴U treatment. Replicate samples are indicated by (rep). (D) Volcano plot depicting results from a comparative analysis of miRNAs that are significantly enriched or depleted in early time points (20 min, 1 hr) relative to steady state levels (6 and 22 days). Fast-turnover miRNAs (fold difference early time points from steady state > 4; p-value < 2x10⁻⁵; Bonferroni family-wise error rate < 0.005) are colored red; slow-turnover miRNAs (fold difference early time points from steady state < 0.25; p-value < 2x10⁻⁵; Bonferroni family-wise error rate < 0.005) are shown in blue. Stars indicate miRNAs defined as miRNA-stars (see Experimental Procedures); the others are indicated with circles. (E) Heatmap indicating normalized miRNA enrichment relative to steady state level at each time point in RATE-seq for the fast- and slow-turnover miRNAs in (C). For clarity of presentation, the most significant fast-turnover miRNA in this analysis (miR-4521, log₂(fold-change) = 10.8; p-value = 2.9x10⁻⁴⁰) has been omitted from (C) and (D) due to values exceeding the indicated scales. See also Figure S5 and Table S2.