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Evolution acts on enhancer organization to fine-tune gradient threshold readouts.

Crocker J, Tamori Y, Erives A - PLoS Biol. (2008)

Bottom Line: Furthermore, by precisely altering the organization of NEEs with different morphogen gradient threshold readouts, we show that CRM organizational evolution is sufficient for explaining changes in enhancer activity.Thus, evolution can act on CRM organization to fine-tune morphogen gradient threshold readouts over a wide dynamic range.Our study demonstrates that equivalence classes of CRMs are powerful tools for detecting lineage-specific adaptations by gene regulatory sequences.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences, Dartmouth College, Hanover, NH, USA.

ABSTRACT
The elucidation of principles governing evolution of gene regulatory sequence is critical to the study of metazoan diversification. We are therefore exploring the structure and organizational constraints of regulatory sequences by studying functionally equivalent cis-regulatory modules (CRMs) that have been evolving in parallel across several loci. Such an independent dataset allows a multi-locus study that is not hampered by nonfunctional or constrained homology. The neurogenic ectoderm enhancers (NEEs) of Drosophila melanogaster are one such class of coordinately regulated CRMs. The NEEs share a common organization of binding sites and as a set would be useful to study the relationship between CRM organization and CRM activity across evolving lineages. We used the D. melanogaster transgenic system to screen for functional adaptations in the NEEs from divergent drosophilid species. We show that the individual NEE modules across a genome in any one lineage have independently evolved adaptations to compensate for lineage-specific developmental and/or genomic changes. Specifically, we show that both the site composition and the site organization of NEEs have been finely tuned by distinct, lineage-specific selection pressures in each of the three divergent species that we have examined: D. melanogaster, D. pseudoobscura, and D. virilis. Furthermore, by precisely altering the organization of NEEs with different morphogen gradient threshold readouts, we show that CRM organizational evolution is sufficient for explaining changes in enhancer activity. Thus, evolution can act on CRM organization to fine-tune morphogen gradient threshold readouts over a wide dynamic range. Our study demonstrates that equivalence classes of CRMs are powerful tools for detecting lineage-specific adaptations by gene regulatory sequences.

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Fluorescent Double-Labeling of rho NEE-Driven Transgenes(A–C) Double-staining (lacZ and snail expression in green and purple, respectively) for rho NEE driven transgenes from D. pseudoobscura (A), D. melanogaster (B), and D. virilis (C) and snail (sna) expression in stage 5 embryos. The sharp snail border of expression provides a landmark to align expression patterns across embryos.(D) All three enhancers drive expression patterns of similar intensity although the D/V axis although the width of the stripe is narrower for D. pseudoobscura, and wider for D. virilis than the D. melanogaster NEE driven transgene.
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pbio-0060263-g005: Fluorescent Double-Labeling of rho NEE-Driven Transgenes(A–C) Double-staining (lacZ and snail expression in green and purple, respectively) for rho NEE driven transgenes from D. pseudoobscura (A), D. melanogaster (B), and D. virilis (C) and snail (sna) expression in stage 5 embryos. The sharp snail border of expression provides a landmark to align expression patterns across embryos.(D) All three enhancers drive expression patterns of similar intensity although the D/V axis although the width of the stripe is narrower for D. pseudoobscura, and wider for D. virilis than the D. melanogaster NEE driven transgene.

Mentions: Interestingly, despite similar functional outputs of orthologous NEEs in the context of their native genomes (Figure 1F), the NEEs from each species have unidirectionally modified activities in transgenic D. melanogaster embryos when compared with all the NEEs as a group from other species (Figure 3; in situ detection experiments were conducted in parallel). Specifically, the D. virilis enhancers consistently drive expression of reporters in a significantly more robust and expansive lateral stripe than the D. melanogaster enhancers in transgenic D. melanogaster stage 5(2) embryos, whereas D. pseudoobscura enhancers drive expression in a narrower stripes than the D. melanogaster enhancers (Figure 3). We have also verified this by conducting fluorescent double-label in situ hybridization of NEE-driven lacZ reporter lines using anti-lacZ and anti-snail RNA probes; snail labels the mesoderm (Figures 4 and 5). These experiments reveal that NEE-driven lacZ expression immediately abuts the mesodermal border without overlapping with it, which is consistent with ventral repression of NEEs by Snail.


Evolution acts on enhancer organization to fine-tune gradient threshold readouts.

Crocker J, Tamori Y, Erives A - PLoS Biol. (2008)

Fluorescent Double-Labeling of rho NEE-Driven Transgenes(A–C) Double-staining (lacZ and snail expression in green and purple, respectively) for rho NEE driven transgenes from D. pseudoobscura (A), D. melanogaster (B), and D. virilis (C) and snail (sna) expression in stage 5 embryos. The sharp snail border of expression provides a landmark to align expression patterns across embryos.(D) All three enhancers drive expression patterns of similar intensity although the D/V axis although the width of the stripe is narrower for D. pseudoobscura, and wider for D. virilis than the D. melanogaster NEE driven transgene.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2577699&req=5

pbio-0060263-g005: Fluorescent Double-Labeling of rho NEE-Driven Transgenes(A–C) Double-staining (lacZ and snail expression in green and purple, respectively) for rho NEE driven transgenes from D. pseudoobscura (A), D. melanogaster (B), and D. virilis (C) and snail (sna) expression in stage 5 embryos. The sharp snail border of expression provides a landmark to align expression patterns across embryos.(D) All three enhancers drive expression patterns of similar intensity although the D/V axis although the width of the stripe is narrower for D. pseudoobscura, and wider for D. virilis than the D. melanogaster NEE driven transgene.
Mentions: Interestingly, despite similar functional outputs of orthologous NEEs in the context of their native genomes (Figure 1F), the NEEs from each species have unidirectionally modified activities in transgenic D. melanogaster embryos when compared with all the NEEs as a group from other species (Figure 3; in situ detection experiments were conducted in parallel). Specifically, the D. virilis enhancers consistently drive expression of reporters in a significantly more robust and expansive lateral stripe than the D. melanogaster enhancers in transgenic D. melanogaster stage 5(2) embryos, whereas D. pseudoobscura enhancers drive expression in a narrower stripes than the D. melanogaster enhancers (Figure 3). We have also verified this by conducting fluorescent double-label in situ hybridization of NEE-driven lacZ reporter lines using anti-lacZ and anti-snail RNA probes; snail labels the mesoderm (Figures 4 and 5). These experiments reveal that NEE-driven lacZ expression immediately abuts the mesodermal border without overlapping with it, which is consistent with ventral repression of NEEs by Snail.

Bottom Line: Furthermore, by precisely altering the organization of NEEs with different morphogen gradient threshold readouts, we show that CRM organizational evolution is sufficient for explaining changes in enhancer activity.Thus, evolution can act on CRM organization to fine-tune morphogen gradient threshold readouts over a wide dynamic range.Our study demonstrates that equivalence classes of CRMs are powerful tools for detecting lineage-specific adaptations by gene regulatory sequences.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences, Dartmouth College, Hanover, NH, USA.

ABSTRACT
The elucidation of principles governing evolution of gene regulatory sequence is critical to the study of metazoan diversification. We are therefore exploring the structure and organizational constraints of regulatory sequences by studying functionally equivalent cis-regulatory modules (CRMs) that have been evolving in parallel across several loci. Such an independent dataset allows a multi-locus study that is not hampered by nonfunctional or constrained homology. The neurogenic ectoderm enhancers (NEEs) of Drosophila melanogaster are one such class of coordinately regulated CRMs. The NEEs share a common organization of binding sites and as a set would be useful to study the relationship between CRM organization and CRM activity across evolving lineages. We used the D. melanogaster transgenic system to screen for functional adaptations in the NEEs from divergent drosophilid species. We show that the individual NEE modules across a genome in any one lineage have independently evolved adaptations to compensate for lineage-specific developmental and/or genomic changes. Specifically, we show that both the site composition and the site organization of NEEs have been finely tuned by distinct, lineage-specific selection pressures in each of the three divergent species that we have examined: D. melanogaster, D. pseudoobscura, and D. virilis. Furthermore, by precisely altering the organization of NEEs with different morphogen gradient threshold readouts, we show that CRM organizational evolution is sufficient for explaining changes in enhancer activity. Thus, evolution can act on CRM organization to fine-tune morphogen gradient threshold readouts over a wide dynamic range. Our study demonstrates that equivalence classes of CRMs are powerful tools for detecting lineage-specific adaptations by gene regulatory sequences.

Show MeSH
Related in: MedlinePlus