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Silencing is noisy: population and cell level noise in telomere-adjacent genes is dependent on telomere position and sir2.

Anderson MZ, Gerstein AC, Wigen L, Baller JA, Berman J - PLoS Genet. (2014)

Bottom Line: Finally, we found that telomere silencing and TAGEN are tightly linked and regulated in cis: selection for either silencing or activation of a TLO-adjacent URA3 gene resulted in reduced noise at the neighboring TLO but not at other TLO genes.This provides experimental support to computational predictions that the ability to shift between silent and active chromatin states has a major effect on cell-to-cell noise.Furthermore, it demonstrates that these shifts affect the degree of expression variation at each telomere individually.

View Article: PubMed Central - PubMed

Affiliation: Department of Genetics, Cell Biology and Development, University of Minnesota - Twin Cities, Minneapolis, Minnesota, United States of America.

ABSTRACT
Cell-to-cell gene expression noise is thought to be an important mechanism for generating phenotypic diversity. Furthermore, telomeric regions are major sites for gene amplification, which is thought to drive genetic diversity. Here we found that individual subtelomeric TLO genes exhibit increased variation in transcript and protein levels at both the cell-to-cell level as well as at the population-level. The cell-to-cell variation, termed Telomere-Adjacent Gene Expression Noise (TAGEN) was largely intrinsic noise and was dependent upon genome position: noise was reduced when a TLO gene was expressed at an ectopic internal locus and noise was elevated when a non-telomeric gene was expressed at a telomere-adjacent locus. This position-dependent TAGEN also was dependent on Sir2p, an NAD+-dependent histone deacetylase. Finally, we found that telomere silencing and TAGEN are tightly linked and regulated in cis: selection for either silencing or activation of a TLO-adjacent URA3 gene resulted in reduced noise at the neighboring TLO but not at other TLO genes. This provides experimental support to computational predictions that the ability to shift between silent and active chromatin states has a major effect on cell-to-cell noise. Furthermore, it demonstrates that these shifts affect the degree of expression variation at each telomere individually.

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Sir2 contributes to TLO TAGEN.(A) Transcript abundance measurements of six TLOs and two control genes were collected from either SIR2 or sir2Δ/Δ cells and in the presence or absence of the Sir-type HDAC inhibitor nicotinamide (NAM). Subtelomeric TLO expression plasticity specifically decreased when either treated with NAM or in the sir2Δ/Δ background but mean expression was not affected. Treatment of sir2Δ/Δ cells with NAM does not further decrease expression variability. (B) Fluorescence microscopy of GFP-tagged Tlos in either a SIR2 or sir2Δ/Δ background showed reduced cell-to-cell variation in a sir2Δ/Δ background. (C) Flow cytometry of GFP tagged Nup49, Tloα10, and Tloα12 also identified significantly reduced noise for both Tlos in the sir2Δ/Δ background. Fluorescence signal of Tloα10 was also increased in a SIR2 deletion strain. (D) Flow cytometry measured fluorescence signal of Nup49-GFP expressed at either the subtelomeric TLOα9 or internal NUP49 locus in both a SIR2 and sir2Δ/Δ background. Gene noise of subtelomeric Nup49-GFP decreased significantly in the sir2Δ/Δ background. * denotes p<0.05. ** denotes p<0.01.
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pgen-1004436-g007: Sir2 contributes to TLO TAGEN.(A) Transcript abundance measurements of six TLOs and two control genes were collected from either SIR2 or sir2Δ/Δ cells and in the presence or absence of the Sir-type HDAC inhibitor nicotinamide (NAM). Subtelomeric TLO expression plasticity specifically decreased when either treated with NAM or in the sir2Δ/Δ background but mean expression was not affected. Treatment of sir2Δ/Δ cells with NAM does not further decrease expression variability. (B) Fluorescence microscopy of GFP-tagged Tlos in either a SIR2 or sir2Δ/Δ background showed reduced cell-to-cell variation in a sir2Δ/Δ background. (C) Flow cytometry of GFP tagged Nup49, Tloα10, and Tloα12 also identified significantly reduced noise for both Tlos in the sir2Δ/Δ background. Fluorescence signal of Tloα10 was also increased in a SIR2 deletion strain. (D) Flow cytometry measured fluorescence signal of Nup49-GFP expressed at either the subtelomeric TLOα9 or internal NUP49 locus in both a SIR2 and sir2Δ/Δ background. Gene noise of subtelomeric Nup49-GFP decreased significantly in the sir2Δ/Δ background. * denotes p<0.05. ** denotes p<0.01.

Mentions: The Sir2p HDAC was required for TLO expression variability between colonies. Therefore, we hypothesized Sir2 may also influence TLO noise among cells in a single population. We first asked whether addition of nicotinamide (NAM), an inhibitor of NAD+-dependent HDACs, or deletion of SIR2 had an effect on TAGEN at TLO genes using qRT-PCR. Addition of NAM or the lack of Sir2p significantly reduced expression plasticity (measured with qPCR, Fig. 7A, S7; background: F1 = 6.44, p = 0.020; NAM: F1 = 7.79, p = 0.011; interaction: F1 = 3.25, p = 0.086), while neither NAM nor the absence of Sir2p significantly influenced mean TLO gene expression (Fig. S7; Sir2 background: F1 = 0.03, p = 0.86; NAM: F1 = 1.42, p = 0.25). Furthermore, the effect of deleting SIR2 together with NAM exposure affected expression and plasticity to a similar degree as either NAM or deletion of SIR2 alone: reduced variability with little effect on expression levels (interaction; CV: F1 = 3.25, p = 0.086). Similar results for wild-type vs sir2Δ/Δ mutants were obtained by microscopy (Fig. 7B, S8; p<0.05) as well as by flow cytometry of Tloα10-GFP or Tloα12-GFP (Fig. 7C; Robust CV; gene: F1 = 1.21, p = 0.29; Sir2 background: F1 = 5.44, p = 0.03; interaction: F1 = 0.165, p = 0.69). Thus, Sir2p makes a significant contribution to expression plasticity of Tloα10-GFP and Tloα12-GFP.


Silencing is noisy: population and cell level noise in telomere-adjacent genes is dependent on telomere position and sir2.

Anderson MZ, Gerstein AC, Wigen L, Baller JA, Berman J - PLoS Genet. (2014)

Sir2 contributes to TLO TAGEN.(A) Transcript abundance measurements of six TLOs and two control genes were collected from either SIR2 or sir2Δ/Δ cells and in the presence or absence of the Sir-type HDAC inhibitor nicotinamide (NAM). Subtelomeric TLO expression plasticity specifically decreased when either treated with NAM or in the sir2Δ/Δ background but mean expression was not affected. Treatment of sir2Δ/Δ cells with NAM does not further decrease expression variability. (B) Fluorescence microscopy of GFP-tagged Tlos in either a SIR2 or sir2Δ/Δ background showed reduced cell-to-cell variation in a sir2Δ/Δ background. (C) Flow cytometry of GFP tagged Nup49, Tloα10, and Tloα12 also identified significantly reduced noise for both Tlos in the sir2Δ/Δ background. Fluorescence signal of Tloα10 was also increased in a SIR2 deletion strain. (D) Flow cytometry measured fluorescence signal of Nup49-GFP expressed at either the subtelomeric TLOα9 or internal NUP49 locus in both a SIR2 and sir2Δ/Δ background. Gene noise of subtelomeric Nup49-GFP decreased significantly in the sir2Δ/Δ background. * denotes p<0.05. ** denotes p<0.01.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4109849&req=5

pgen-1004436-g007: Sir2 contributes to TLO TAGEN.(A) Transcript abundance measurements of six TLOs and two control genes were collected from either SIR2 or sir2Δ/Δ cells and in the presence or absence of the Sir-type HDAC inhibitor nicotinamide (NAM). Subtelomeric TLO expression plasticity specifically decreased when either treated with NAM or in the sir2Δ/Δ background but mean expression was not affected. Treatment of sir2Δ/Δ cells with NAM does not further decrease expression variability. (B) Fluorescence microscopy of GFP-tagged Tlos in either a SIR2 or sir2Δ/Δ background showed reduced cell-to-cell variation in a sir2Δ/Δ background. (C) Flow cytometry of GFP tagged Nup49, Tloα10, and Tloα12 also identified significantly reduced noise for both Tlos in the sir2Δ/Δ background. Fluorescence signal of Tloα10 was also increased in a SIR2 deletion strain. (D) Flow cytometry measured fluorescence signal of Nup49-GFP expressed at either the subtelomeric TLOα9 or internal NUP49 locus in both a SIR2 and sir2Δ/Δ background. Gene noise of subtelomeric Nup49-GFP decreased significantly in the sir2Δ/Δ background. * denotes p<0.05. ** denotes p<0.01.
Mentions: The Sir2p HDAC was required for TLO expression variability between colonies. Therefore, we hypothesized Sir2 may also influence TLO noise among cells in a single population. We first asked whether addition of nicotinamide (NAM), an inhibitor of NAD+-dependent HDACs, or deletion of SIR2 had an effect on TAGEN at TLO genes using qRT-PCR. Addition of NAM or the lack of Sir2p significantly reduced expression plasticity (measured with qPCR, Fig. 7A, S7; background: F1 = 6.44, p = 0.020; NAM: F1 = 7.79, p = 0.011; interaction: F1 = 3.25, p = 0.086), while neither NAM nor the absence of Sir2p significantly influenced mean TLO gene expression (Fig. S7; Sir2 background: F1 = 0.03, p = 0.86; NAM: F1 = 1.42, p = 0.25). Furthermore, the effect of deleting SIR2 together with NAM exposure affected expression and plasticity to a similar degree as either NAM or deletion of SIR2 alone: reduced variability with little effect on expression levels (interaction; CV: F1 = 3.25, p = 0.086). Similar results for wild-type vs sir2Δ/Δ mutants were obtained by microscopy (Fig. 7B, S8; p<0.05) as well as by flow cytometry of Tloα10-GFP or Tloα12-GFP (Fig. 7C; Robust CV; gene: F1 = 1.21, p = 0.29; Sir2 background: F1 = 5.44, p = 0.03; interaction: F1 = 0.165, p = 0.69). Thus, Sir2p makes a significant contribution to expression plasticity of Tloα10-GFP and Tloα12-GFP.

Bottom Line: Finally, we found that telomere silencing and TAGEN are tightly linked and regulated in cis: selection for either silencing or activation of a TLO-adjacent URA3 gene resulted in reduced noise at the neighboring TLO but not at other TLO genes.This provides experimental support to computational predictions that the ability to shift between silent and active chromatin states has a major effect on cell-to-cell noise.Furthermore, it demonstrates that these shifts affect the degree of expression variation at each telomere individually.

View Article: PubMed Central - PubMed

Affiliation: Department of Genetics, Cell Biology and Development, University of Minnesota - Twin Cities, Minneapolis, Minnesota, United States of America.

ABSTRACT
Cell-to-cell gene expression noise is thought to be an important mechanism for generating phenotypic diversity. Furthermore, telomeric regions are major sites for gene amplification, which is thought to drive genetic diversity. Here we found that individual subtelomeric TLO genes exhibit increased variation in transcript and protein levels at both the cell-to-cell level as well as at the population-level. The cell-to-cell variation, termed Telomere-Adjacent Gene Expression Noise (TAGEN) was largely intrinsic noise and was dependent upon genome position: noise was reduced when a TLO gene was expressed at an ectopic internal locus and noise was elevated when a non-telomeric gene was expressed at a telomere-adjacent locus. This position-dependent TAGEN also was dependent on Sir2p, an NAD+-dependent histone deacetylase. Finally, we found that telomere silencing and TAGEN are tightly linked and regulated in cis: selection for either silencing or activation of a TLO-adjacent URA3 gene resulted in reduced noise at the neighboring TLO but not at other TLO genes. This provides experimental support to computational predictions that the ability to shift between silent and active chromatin states has a major effect on cell-to-cell noise. Furthermore, it demonstrates that these shifts affect the degree of expression variation at each telomere individually.

Show MeSH
Related in: MedlinePlus