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Figure 2. Poly(A)-tail lengths in yeast, plant, fly and vertebrate cellsa, Global tail-length distributions. For each sample,
histograms tally tail-length measurements for all poly(A) tags mapping to
annotated 3â² UTRs (bin size = 5 nt). Leftmost bin includes all
measurements <0 nt. Median tail lengths are in parentheses.
b, Intergenic tail-length distributions. For each sample,
histograms tally average tail lengths for protein-coding genes with â¥50
tags (yeasts, zebrafish and Xenopus) or â¥100 tags
(other samples). Median average tail lengths are in parentheses. c,
Intragenic tail-length distributions for 10 genes sampling the spectrum of
average tail lengths in 3T3 cells. d, Intragenic tail-length
distributions. Heatmaps show the frequency distribution of tail lengths for each
gene tallied in b. The color intensity indicates the fraction of
the total for the gene. Genes are ordered by average tail length (dashed line).
Results from the S. cerevisiae total-RNA sample are reported in this figure.
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Figure 3. Transient coupling between poly(A)-tail length and TEa, Relationship between mean tail length and TE for genes
with â¥50 poly(A) tags from embryonic samples at the indicated
developmental stages. For each stage, tail lengths and TEs were obtained from
the same sample. MGC116473 and DDX24 fell outside the plot for
X. laevis, stages 3â4, and
LOC100049092 fell outside the plot for X.
laevis, stages 12â12.5. b, Relationship
between mean tail length and TE in the indicated cells, for genes with
â¥50 (yeasts) or â¥100 (others) tags. With the exception of
HeLa35, tail lengths
and TEs were from the same samples. Budding yeast YBR196C,
YLR355C and YDL080C, fission yeast
SPCC63.04.1, mouse-liver NM_007881 and
NM_145470, HEK293T NM_001007026,
NM_021058, and NM_003537 and HeLa
NM_001007026 fell outside their respective plots.
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Figure 4. No detectable intragenic coupling between poly(A)-tail length and TEa, Global analysis of tail lengths across the polysome
profile for 3T3 cells. UV absorbance indicates mean number of ribosomes bound
per mRNA for each fraction from the sucrose gradient (top, fractions demarcated
with vertical dashed lines). Boxplots show distributions of tail lengths in each
fraction for all tags mapping to annotated 3â² UTRs (bottom). Boxplot
percentiles are line, median; box, 25th and 75th
percentiles; whiskers, 10th and 90th percentiles. The
horizontal line indicates the overall median of the median tail lengths.
b, Relationship between tail lengths and ribosomes bound per
mRNA for mRNAs from the same gene. For each gene, the data from a
were used to plot the mean tail length as a function of bound ribosomes. Log-log
plots for 8 randomly selected genes with â¥50 poly(A) tags in â¥6
fractions are shown (left), with lines indicating linear least-squared fits to
the data (adding a pseudocount of 0.5 ribosomes to the fraction with 0
ribosomes). The boxplot shows the distribution of slopes for all genes with
â¥50 poly(A) tags in â¥4 fractions (right; n = 4,079; one-sided,
one-sample Wilcoxon test; boxplot percentiles as in a).
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Figure 5. The influence of miR-155 on ribosomes, mRNA and tails in the early zebrafish
embryoa, Relationship between changes in ribosome protected
fragments (RPFs) and changes in mRNA levels after injecting miR-155. Changes
observed between miRNA- and mock-injected embryos are plotted at the indicated
stages for predicted miR-155 target genes (red, genes with â¥1 miR-155
site in their 3â² UTR) and control genes (gray, genes that have no
miR-155 site yet resemble the predicted targets with respect to 3â² UTR
length). To ensure that differences observed between 4 and 6 hpf were not the
result of examining different genes, only site-containing genes and no-site
control genes detected at both 4 and 6 hpf are shown for these stages. Lines
indicate mean changes for the respective gene sets, with statistically
significant differences between the sets indicated (*, P
â¤0.05; **, P <10â4,
one-tailed KolmogorovâSmirnov test). Because injected miRNAs partially
inhibited miR-430âmediated repression, genes with miR-430 sites were not
considered. Data were normalized to the median changes observed for the
controls. b, Relationship between RPF changes and mean tail-length
changes after injecting miR-155. Tail-lengths were determined using PAL-seq,
otherwise as in a. c, A developmental switch in the
dominant mode of miRNAâmediated repression. The schematic (left) depicts
the components of the bar graphs, showing how the RPF changes comprise both mRNA
and TE changes. The compound bar graphs show the fraction of repression
attributed to mRNA degradation (blue) and TE (green) for the indicated stage,
depicting the overall impact of miR-155 (center; plotting results from
a and b for genes with sites) and miR-132 (right,
plotting results from Extended
Data Fig. 8b for genes with sites). Slight,
statistically insignificant, increases in mRNA for predicted targets resulted in
blue bars extending above the axis. For samples from stages in which tail length
and TE are coupled, a bracket adjacent to the compound bar indicates the
fraction of repression attributable to shortened tails. Significant changes for
each component are indicated with asterisks of the corresponding color (*,
P â¤0.05; **, P
<10â4, one-tailed KolmogorovâSmirnov
test).
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