Figure 1.

A genome-wide screen identified mRNAs with rhythms in poly(A) tail length in mouse livers. (A) Oligo(dT) chromatography was used to separate total RNA into fractions with either short or long poly(A) tails by varying salt concentrations in the elution. Total polyA⁺ (nonfractionated) RNAs were also isolated as a reference. The poly(A) tail length (nucleotides) of each fraction was tested by 3′ end label assay. Representative gel images are shown on the right. (B) Shown is the tissue sampling scheme in which mouse livers (n = 3 per time point) were harvested every 4 h under 12L:12D conditions, and total RNA was extracted from each sample followed by RNA fractionation as depicted in A. Following the fractionation, each RNA sample was subjected to microarray analysis. (C–F) Comparison of the signal intensity at ZT0 of each mRNA in the polyA⁺ samples versus the short-tailed samples (C), in the polyA⁺ samples versus the long-tailed samples (D), in the short-tailed samples versus the long-tailed samples (E), and in the polyA⁺ samples versus the log₁₀(long/short ratio) (F). The degree of correlation is shown in the top left corner of each graph. The data for other time points are shown in Supplemental Figure S1. (G) Phase-sorted heat map of the long/short ratio of PAR mRNAs. Red or green indicates higher or lower values in the long/short ratio (longer vs. shorter tails), respectively. Each line represents one mRNA. The heat map is double-plotted for easier visualization of rhythmicity. (H) Peak time distribution of PAR mRNAs. Peak time of poly(A) tail rhythms was calculated based on the long/short ratio of PAR mRNAs. Each wedge represents a 2-h bin. (I) Representative gel images of LM-PAT assay for PAR mRNAs. Each lane consists of pooled samples (n = 3 for each time point). Arrowheads on the right indicate the fragment derived from restriction enzyme treatment for verification of PCR specificity (see the Materials and Methods for details). Graphs are the comparison between the long/short ratio from the microarray (left Y-axis, dotted line) and the poly(A) tail length calculated from the LM-PAT assay (right X-axis, solid line).