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Slow-growing cells within isogenic populations have increased RNA polymerase error rates and DNA damage.

van Dijk D, Dhar R, Missarova AM, Espinar L, Blevins WR, Lehner B, Carey LB - Nat Commun (2015)

Bottom Line: To characterize transcriptional differences associated with this variability, we have developed a method--FitFlow--that enables the sorting of subpopulations by growth rate.More generally, we find a significantly altered transcriptome in the slow-growing subpopulation that only partially resembles that of cells growing slowly due to environmental and culture conditions.Slow-growing cells upregulate transposons and express more chromosomal, viral and plasmid-borne transcripts, and thus explore a larger genotypic--and so phenotypic--space.

View Article: PubMed Central - PubMed

Affiliation: 1] Department of Biological Sciences, Columbia University, New York, New York 10027, USA [2] Department of Systems Biology, Columbia University, New York, New York 10027, USA [3] Department of Applied Mathematics, Weizmann Institute of Science, Rehovot, 7610001, Israel [4] Department of Molecular Cell Biology, Weizmann Institute of Science, Rehovot, 7610001, Israel.

ABSTRACT
Isogenic cells show a large degree of variability in growth rate, even when cultured in the same environment. Such cell-to-cell variability in growth can alter sensitivity to antibiotics, chemotherapy and environmental stress. To characterize transcriptional differences associated with this variability, we have developed a method--FitFlow--that enables the sorting of subpopulations by growth rate. The slow-growing subpopulation shows a transcriptional stress response, but, more surprisingly, these cells have reduced RNA polymerase fidelity and exhibit a DNA damage response. As DNA damage is often caused by oxidative stress, we test the addition of an antioxidant, and find that it reduces the size of the slow-growing population. More generally, we find a significantly altered transcriptome in the slow-growing subpopulation that only partially resembles that of cells growing slowly due to environmental and culture conditions. Slow-growing cells upregulate transposons and express more chromosomal, viral and plasmid-borne transcripts, and thus explore a larger genotypic--and so phenotypic--space.

No MeSH data available.


Related in: MedlinePlus

More transcriptional diversity in slow-growing subpopulations.(a) At low expression (<5 FPKM (fragments per kilobase of transcript per million mapped reads)), slow- and fast-growing cells express similar numbers of transcripts, but at medium (5–30 FPKM), slow-growing cells express both more genes and more unique gene functions (paired t-test P<1e−35 for transcripts and GO terms). (b) The slow-growing subpopulation expresses more unannotated transcripts (paired ks-test P=5.36 × 10−10) and antisense transcripts (paired ks-test P=1.34 × 10−22) at >10 FPKM. (c) Highly expressed genes (higher than one s.d., red) are upregulated (paired ks-test P=1.1 × 10−63), while lowly expressed genes (lower than one s.d., blue) tend to be downregulated with increasing subpopulation growth rate (paired ks-test P=4.6 × 10−35). y axis shows the average expression level in all measured populations. The x axis shows expression change from slow to fast subpopulation growth, computed as the log2 ratio.
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f2: More transcriptional diversity in slow-growing subpopulations.(a) At low expression (<5 FPKM (fragments per kilobase of transcript per million mapped reads)), slow- and fast-growing cells express similar numbers of transcripts, but at medium (5–30 FPKM), slow-growing cells express both more genes and more unique gene functions (paired t-test P<1e−35 for transcripts and GO terms). (b) The slow-growing subpopulation expresses more unannotated transcripts (paired ks-test P=5.36 × 10−10) and antisense transcripts (paired ks-test P=1.34 × 10−22) at >10 FPKM. (c) Highly expressed genes (higher than one s.d., red) are upregulated (paired ks-test P=1.1 × 10−63), while lowly expressed genes (lower than one s.d., blue) tend to be downregulated with increasing subpopulation growth rate (paired ks-test P=4.6 × 10−35). y axis shows the average expression level in all measured populations. The x axis shows expression change from slow to fast subpopulation growth, computed as the log2 ratio.

Mentions: The faster a population grows, the greater the proportion of the transcriptome is dedicated to ribosome production9. In rapidly growing cells, 50% of mRNA synthesis is dedicated to ∼10% of genes19. Consistent with these population-level results, we found that, in the fast subpopulation, the most highly expressed genes account for a large fraction of the total transcriptome. In contrast, in the slow subpopulation, the rest of the genome is more highly expressed (Fig. 2). This is not an artefact of detection bias due to fast cells having higher expression of highly expressed genes (Supplementary Figs 6 and 7). In addition to expressing more genes, slow-growing cells also express more unique gene functions (Fig. 2a). An increase in transcription of highly expressed genes results in a far greater commitment of cellular resources than a similar fold change in the transcription of genes with low expression. This suggests that the slow subpopulation shifts resources from high expression of only a few genes to the more moderate expression of a large number of genes. While deletion of most genes does not cause a growth defect in rich media, there exists some condition in which each gene is useful20. We find that these slow cells have higher expression of genes that, in fast-growing cells, have little or no expression (Fig. 2). Furthermore, novel (unannotated) and antisense transcripts are expressed at a higher level in the slow-growing subpopulation (Fig. 2b). These results show that the slow-growing subpopulation expresses more genes and unique enzymatic and cellular functions, which may, in turn, allow them to explore a larger phenotypic space.


Slow-growing cells within isogenic populations have increased RNA polymerase error rates and DNA damage.

van Dijk D, Dhar R, Missarova AM, Espinar L, Blevins WR, Lehner B, Carey LB - Nat Commun (2015)

More transcriptional diversity in slow-growing subpopulations.(a) At low expression (<5 FPKM (fragments per kilobase of transcript per million mapped reads)), slow- and fast-growing cells express similar numbers of transcripts, but at medium (5–30 FPKM), slow-growing cells express both more genes and more unique gene functions (paired t-test P<1e−35 for transcripts and GO terms). (b) The slow-growing subpopulation expresses more unannotated transcripts (paired ks-test P=5.36 × 10−10) and antisense transcripts (paired ks-test P=1.34 × 10−22) at >10 FPKM. (c) Highly expressed genes (higher than one s.d., red) are upregulated (paired ks-test P=1.1 × 10−63), while lowly expressed genes (lower than one s.d., blue) tend to be downregulated with increasing subpopulation growth rate (paired ks-test P=4.6 × 10−35). y axis shows the average expression level in all measured populations. The x axis shows expression change from slow to fast subpopulation growth, computed as the log2 ratio.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4557116&req=5

f2: More transcriptional diversity in slow-growing subpopulations.(a) At low expression (<5 FPKM (fragments per kilobase of transcript per million mapped reads)), slow- and fast-growing cells express similar numbers of transcripts, but at medium (5–30 FPKM), slow-growing cells express both more genes and more unique gene functions (paired t-test P<1e−35 for transcripts and GO terms). (b) The slow-growing subpopulation expresses more unannotated transcripts (paired ks-test P=5.36 × 10−10) and antisense transcripts (paired ks-test P=1.34 × 10−22) at >10 FPKM. (c) Highly expressed genes (higher than one s.d., red) are upregulated (paired ks-test P=1.1 × 10−63), while lowly expressed genes (lower than one s.d., blue) tend to be downregulated with increasing subpopulation growth rate (paired ks-test P=4.6 × 10−35). y axis shows the average expression level in all measured populations. The x axis shows expression change from slow to fast subpopulation growth, computed as the log2 ratio.
Mentions: The faster a population grows, the greater the proportion of the transcriptome is dedicated to ribosome production9. In rapidly growing cells, 50% of mRNA synthesis is dedicated to ∼10% of genes19. Consistent with these population-level results, we found that, in the fast subpopulation, the most highly expressed genes account for a large fraction of the total transcriptome. In contrast, in the slow subpopulation, the rest of the genome is more highly expressed (Fig. 2). This is not an artefact of detection bias due to fast cells having higher expression of highly expressed genes (Supplementary Figs 6 and 7). In addition to expressing more genes, slow-growing cells also express more unique gene functions (Fig. 2a). An increase in transcription of highly expressed genes results in a far greater commitment of cellular resources than a similar fold change in the transcription of genes with low expression. This suggests that the slow subpopulation shifts resources from high expression of only a few genes to the more moderate expression of a large number of genes. While deletion of most genes does not cause a growth defect in rich media, there exists some condition in which each gene is useful20. We find that these slow cells have higher expression of genes that, in fast-growing cells, have little or no expression (Fig. 2). Furthermore, novel (unannotated) and antisense transcripts are expressed at a higher level in the slow-growing subpopulation (Fig. 2b). These results show that the slow-growing subpopulation expresses more genes and unique enzymatic and cellular functions, which may, in turn, allow them to explore a larger phenotypic space.

Bottom Line: To characterize transcriptional differences associated with this variability, we have developed a method--FitFlow--that enables the sorting of subpopulations by growth rate.More generally, we find a significantly altered transcriptome in the slow-growing subpopulation that only partially resembles that of cells growing slowly due to environmental and culture conditions.Slow-growing cells upregulate transposons and express more chromosomal, viral and plasmid-borne transcripts, and thus explore a larger genotypic--and so phenotypic--space.

View Article: PubMed Central - PubMed

Affiliation: 1] Department of Biological Sciences, Columbia University, New York, New York 10027, USA [2] Department of Systems Biology, Columbia University, New York, New York 10027, USA [3] Department of Applied Mathematics, Weizmann Institute of Science, Rehovot, 7610001, Israel [4] Department of Molecular Cell Biology, Weizmann Institute of Science, Rehovot, 7610001, Israel.

ABSTRACT
Isogenic cells show a large degree of variability in growth rate, even when cultured in the same environment. Such cell-to-cell variability in growth can alter sensitivity to antibiotics, chemotherapy and environmental stress. To characterize transcriptional differences associated with this variability, we have developed a method--FitFlow--that enables the sorting of subpopulations by growth rate. The slow-growing subpopulation shows a transcriptional stress response, but, more surprisingly, these cells have reduced RNA polymerase fidelity and exhibit a DNA damage response. As DNA damage is often caused by oxidative stress, we test the addition of an antioxidant, and find that it reduces the size of the slow-growing population. More generally, we find a significantly altered transcriptome in the slow-growing subpopulation that only partially resembles that of cells growing slowly due to environmental and culture conditions. Slow-growing cells upregulate transposons and express more chromosomal, viral and plasmid-borne transcripts, and thus explore a larger genotypic--and so phenotypic--space.

No MeSH data available.


Related in: MedlinePlus