1. Volume 64, Issue 6, Pages 1088-1101 (December 2016)
Survival in Quiescence Requires the Euchromatic Deployment of Clr4/SUV39H by Argonaute-Associated Small RNAs 
Richard I. Joh, Jasbeer S. Khanduja, Isabel A. Calvo, Meeta Mistry, Christina M. Palmieri, Andrej J. Savol, Shannan J. Ho Sui, Ruslan I. Sadreyev, Martin J. Aryee, Mo Motamedi 
Molecular Cell 
Volume 64, Issue 6, Pages (December 2016)
DOI: /j.molcel
Copyright © 2016 Elsevier Inc. Terms and Conditions

2. Molecular Cell 2016 64, 1088-1101DOI: (10.1016/j.molcel.2016.11.020)
Copyright © 2016 Elsevier Inc. Terms and Conditions

3. Figure 1
Survival in G0 Requires RNAi and Clr4 Proteins and Is Concomitant with Clr4-Dependent Transcriptional Reprogramming
(A and B) Viability of indicated strains at various time points after N starvation (A) and glucose deprivation (0.02%) (B). The error bars represent SD of three biological replicates.
(C) Heatmap (RNA log2 fold change [FC]) depicting the hierarchical clustering of the transcriptome of WT cells at various time points after N starvation relative to proliferative cells (t = 0). Consistent with previous reports (Marguerat et al., 2012; Shimanuki et al., 2007), the early (1 hr and 2 hr) (N starvation and mating) and late (10 hr and 24 hr) (G0) transcriptomes are significantly different from one another.
(D) Difference of RNA log2 FC in transcriptomes of clr4Δ relative to WT cells at indicated times following N starvation. The red dots denote transcripts which show a statistically significant change in clr4Δ relative to WT cells (FDR < 0.05; two biological replicates).
Molecular Cell  , DOI: ( /j.molcel )
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4. Figure 2
G0 Entry Results in RNAi-Dependent H3K9 Methylation of Several Euchromatic Loci
(A) ChIP experiments showing that H3K9me2 accumulates at act1 following N starvation. The fold enrichment of H3K9me2 at act1 relative to cen1 is depicted. The mean and SE of five biological replicates are shown.
(B and C) H3K9me2 accumulation at several euchromatic regions. The average H3K9me2 enrichment (IP over input) of all “H3K9me-enriched” peaks identified by ChIP-seq in WT (B) and clr4Δ (C) at 0, 1, and 24 hr after N starvation is shown. The error bars represent 95% bootstrap confidence interval from 1,000 resamplings.
(D) The genomic distribution of G0-specific H3K9me2 peaks in WT cells is shown as blue bars. The black boxes depict centromeres (chromosome, Ch).
(E) qPCR analyses of H3K9me2 and H3 enrichment at act1 relative to cen1 in G0 (24 hr) of the indicated strains relative to proliferative (0 hr) WT cells. The mean and SE of three biological replicates are graphed.
(F) Average H3K9me2 enrichment of H3K9me-enriched peaks in WT and ago1Δ at 0 and 24 hr after N starvation. The error bars represent 95% bootstrap confidence interval from 1,000 resamplings.
Molecular Cell  , DOI: ( /j.molcel )
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5. Figure 3
G0 Entry Results in the Emergence of Ago1-Associated sRNAs from Several Euchromatic Loci
(A and B) Pie charts depicting the distribution of Ago1-associated sRNAs in proliferative (0 hr) (A) and G0 (24 hr) (B) cells. These graphs reveal that G0 cells have a distinct sRNA profile and show a 5-fold increase in relative abundance of Ago1-associated sRNAs. Here, “heterochromatin” refers to centromeres, telomeres, and the mating type locus; rDNA is shown separately.
(C) The fraction (%) of uniquely mapped Ago1-associated sRNA reads (mapping quality ≥ 10) in proliferative (0 hr) and G0 (24 hr) WT cells. The mean and SE of three biological replicates are graphed.
(D) The genomic distribution of Ago1-associated sRNA-enriched regions in G0 is shown as red bars. The black boxes represent centromeres (chromosome, Ch).
(E and F) Schematic representation of rDNA (E) and the left telomere and chromosome 1 (F) and their sRNA and H3K9me2 dynamics in G0. The strand-specific log2 FC of Ago1-associated sRNAs (magenta) and log2 FC of H3K9me2 (orange) in WT G0 (24 hr) relative to WT proliferative (0 hr) cells are shown. Note that because the sRNA graphs are strand specific, decrease or no change in sRNA levels will appear as a flat part of these graphs.
Molecular Cell  , DOI: ( /j.molcel )
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6. Figure 4
Clr4 and RNAi Proteins Regulate the Expression of Several Euchromatic Gene Clusters in G0
(A) Fold enrichment (FE) of overlap among Clr4-repressed, G0-repressed, sRNA-enriched, and H3K9me-enriched genes. The dotted line represents the expected overlap from random sampling of the same number of genes for each gene set (∗p < 0.05 and ∗∗p < 10−4; two-tailed binomial test).
(B) Ranked distances between Clr4-repressed, sRNA-, and H3K9me-enriched gene sets (red line) compared to the distances based on 10,000 random gene samplings (black line). The median distance (black line) and the 95% confidence interval (gray shading) of the 10,000 samplings are depicted.
(C) Heatmap depicting the spatial distribution of Clr4-repressed, sRNA-, and H3K9me-enriched gene sets. The colors indicate the number of aforementioned genes (0–12) found within a 100 kb region.
Molecular Cell  , DOI: ( /j.molcel )
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7. Figure 5
RNAi- and Clr4-Dependent H3K9me Are Required for the Transcriptional Repression of ght3 and cta3 Regions in Quiescence
(A and B) Schematic representation of ght3 and cta3 regions and fold change (FC) (log2) in RNA, Ago1-associated sRNA, and H3K9me2 levels. The blue, cyan, magenta, and orange graphs represent FC in transcript level in G0 (24 hr) relative to proliferative (0 hr) WT cells, FC in transcript level in G0 (24 hr) clr4Δ relative to G0 (24 hr) WT cells, FC in Ago1-associated sRNA levels in G0 (24 hr) relative to proliferative (0 hr) WT cells, and FC in H3K9me2 in G0 (24 hr) relative to proliferative (0 hr) WT cells, respectively. These data are derived from RNA-, sRNA-, and H3K9me ChIP-seq experiments and all RNA graphs are strand specific.
(C–F) qRT-PCR analyses of gene expression (C and E) and qPCR analyses of H3K9me2 ChIP (D and F) at ght3 and cta3 regions in G0 (24 hr) of the indicated strains relative to proliferative WT cells, respectively. The mean and SE of three to five biological replicates are shown.
Molecular Cell  , DOI: ( /j.molcel )
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8. Figure 6
The Emergence of G0-Specific Ago1-Dependent sRNAs Is Concomitant with the Downregulation of Exosome and TRAMP and Upregulation of RNAi Factors
(A) Venn diagram depicting the overlap among the different total sRNA-enriched genes in ago1Δ, clr4Δ, and dcr1Δ strains.
(B) Graph depicting the total sRNA dynamics of Ago1-associated sRNA-enriched gene set relative to proliferative (0 hr) cells. The majority of these changes occur 1 hr after N starvation.
(C) Graph depicting total sRNA FC (log2) at 1 hr versus 24 hr relative to proliferative (0 hr) WT cells.
(D) Graph depicting total sRNA FC (log2) in G0 in WT and ago1Δ cells. Only Ago1-associated sRNA-enriched genes are shown. r is the co-efficient of correlation.
(E) Relative FC in average expression levels of all exosome, TRAMP, and RNAi components in WT cells at the indicated time points after N starvation relative to proliferative WT cells (mean and SD of two biological replicates).
(F) Heatmap depicting the relative RNA FC of individual RNAi, exosome, and TRAMP components at the indicated time points after N starvation relative to proliferative WT cells. The separate column on the right shows the FC of the same set of genes calculated from a previously published data set (Marguerat et al., 2012). These data show striking similarities to our transcriptome results.
Molecular Cell  , DOI: ( /j.molcel )
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9. Figure 7
RNAi- and H3K9me-Dependent Regulation of Transcription in Quiescence
A decrease in nuclear exosome and co-factors, and an increase in RNAi proteins allow for the generation of Ago1-dependent sRNAs 1 hr post N starvation. These sRNAs act as specificity factors for the recruitment of CLRC and subsequent H3K9 methylation of complementary euchromatic regions in G0 cells.
Molecular Cell  , DOI: ( /j.molcel )
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