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Dynamic evolution of precise regulatory encodings creates the clustered site signature of enhancers.

Crocker J, Potter N, Erives A - Nat Commun (2010)

Bottom Line: However, NEEs also possess dense clusters of variant Dl sites.Here, we show that these increasingly variant sites are eclipsed relic elements, which were superseded by more recently evolved threshold encodings.Given the divergence in egg size during Drosophila lineage evolution, the observed characteristic clusters of divergent sites indicate a history of frequent selection for changes in threshold responses to the Dl morphogen gradient and confirm the NEE structure/function model.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences, Dartmouth College, Hanover, New Hampshire 03755, USA.

ABSTRACT
Concentration gradients of morphogenic proteins pattern the embryonic axes of Drosophila by activating different genes at different concentrations. The neurogenic ectoderm enhancers (NEEs) activate different genes at different threshold levels of the Dorsal (Dl) morphogen, which patterns the dorsal/ventral axis. NEEs share a unique arrangement of highly constrained DNA-binding sites for Dl, Twist (Twi), Snail (Sna) and Suppressor of Hairless (Su(H)), and encode the threshold variable in the precise length of DNA that separates one well-defined Dl element from a Twi element. However, NEEs also possess dense clusters of variant Dl sites. Here, we show that these increasingly variant sites are eclipsed relic elements, which were superseded by more recently evolved threshold encodings. Given the divergence in egg size during Drosophila lineage evolution, the observed characteristic clusters of divergent sites indicate a history of frequent selection for changes in threshold responses to the Dl morphogen gradient and confirm the NEE structure/function model.

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Relic sites are non-functional and accumulate as the enhancer ages.(a) Graph showing the percentage of CA-dinucleotide and CAC-trinucleotide content of several orthologous enhancer sequences from D. melanogaster, D. pseudoobscura, D. willistoni and D. virilis. Each window of NEE sequence is taken ±480 bp from Dβ for each species. Each window of an A/P enhancer is a 960 bp sequence centred around the Bicoid-binding site cluster. Each orthologous set of NEEs is boxed separately to visualize enrichment relative to other groups. The red boxes show the regions occupied by all data points corresponding to a single orthologous set of NEEs located at the indicated locus across many species. The blue box shows the region occupied by all data points corresponding to all A/P enhancers for all species. (b) Identical graph as in panel a, except the data points are boxed by species to visualize genome-specific effects in satellite enrichment or depletion. Red boxes show the region occupied by all data points corresponding to all NEEs within a single species. Canonical A/P enhancers at the eve, gt, Kr and hb loci for all four species are boxed in both panels (blue rectangular area). (c) Graph showing the number of cells spanned by the lacZ expression pattern (vertical axis), as driven by NEEs containing different numbers of Dl half-sites, 5′-SGGAAW-3′ (horizontal axis). (d) Graph showing the number of cells spanned by the lacZ expression pattern (vertical axis), as driven by NEEs characterized by different E-to-D spacer lengths (horizontal axis). Error bars in c and d represent ±1 s.d., as derived from a replicate pool of 20–120 embryos for each construct.
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f5: Relic sites are non-functional and accumulate as the enhancer ages.(a) Graph showing the percentage of CA-dinucleotide and CAC-trinucleotide content of several orthologous enhancer sequences from D. melanogaster, D. pseudoobscura, D. willistoni and D. virilis. Each window of NEE sequence is taken ±480 bp from Dβ for each species. Each window of an A/P enhancer is a 960 bp sequence centred around the Bicoid-binding site cluster. Each orthologous set of NEEs is boxed separately to visualize enrichment relative to other groups. The red boxes show the regions occupied by all data points corresponding to a single orthologous set of NEEs located at the indicated locus across many species. The blue box shows the region occupied by all data points corresponding to all A/P enhancers for all species. (b) Identical graph as in panel a, except the data points are boxed by species to visualize genome-specific effects in satellite enrichment or depletion. Red boxes show the region occupied by all data points corresponding to all NEEs within a single species. Canonical A/P enhancers at the eve, gt, Kr and hb loci for all four species are boxed in both panels (blue rectangular area). (c) Graph showing the number of cells spanned by the lacZ expression pattern (vertical axis), as driven by NEEs containing different numbers of Dl half-sites, 5′-SGGAAW-3′ (horizontal axis). (d) Graph showing the number of cells spanned by the lacZ expression pattern (vertical axis), as driven by NEEs characterized by different E-to-D spacer lengths (horizontal axis). Error bars in c and d represent ±1 s.d., as derived from a replicate pool of 20–120 embryos for each construct.

Mentions: To address the possibility that elevated CA-satellite composition is a feature common to developmental enhancers, we looked at several embryonic enhancers that respond to the Bicoid morphogen gradient, which patterns the A/P axis. We identified complete orthologous sequence sets for the hb embryonic enhancer42, the gt posterior stripe enhancer43, the Kr central domain enhancer4445 and the eve stripe 2 enhancer46 from each of four genomes, namely, D. melanogaster, D. pseudoobscura, D. willistoni and D. virilis. All of these enhancers are active in the same embryonic nuclei as the NEEs and thus constitute a well-matched control group. We find that while the NEE set from any genome is enriched in CA-satellite dinucleotide and trinucleotide fragments, none of the 16 A/P enhancer sets possess the elevated CA-satellite levels that characterize canonical NEEs from these same species, even in genomes with elevated CA-satellite content (Fig. 5a–b).


Dynamic evolution of precise regulatory encodings creates the clustered site signature of enhancers.

Crocker J, Potter N, Erives A - Nat Commun (2010)

Relic sites are non-functional and accumulate as the enhancer ages.(a) Graph showing the percentage of CA-dinucleotide and CAC-trinucleotide content of several orthologous enhancer sequences from D. melanogaster, D. pseudoobscura, D. willistoni and D. virilis. Each window of NEE sequence is taken ±480 bp from Dβ for each species. Each window of an A/P enhancer is a 960 bp sequence centred around the Bicoid-binding site cluster. Each orthologous set of NEEs is boxed separately to visualize enrichment relative to other groups. The red boxes show the regions occupied by all data points corresponding to a single orthologous set of NEEs located at the indicated locus across many species. The blue box shows the region occupied by all data points corresponding to all A/P enhancers for all species. (b) Identical graph as in panel a, except the data points are boxed by species to visualize genome-specific effects in satellite enrichment or depletion. Red boxes show the region occupied by all data points corresponding to all NEEs within a single species. Canonical A/P enhancers at the eve, gt, Kr and hb loci for all four species are boxed in both panels (blue rectangular area). (c) Graph showing the number of cells spanned by the lacZ expression pattern (vertical axis), as driven by NEEs containing different numbers of Dl half-sites, 5′-SGGAAW-3′ (horizontal axis). (d) Graph showing the number of cells spanned by the lacZ expression pattern (vertical axis), as driven by NEEs characterized by different E-to-D spacer lengths (horizontal axis). Error bars in c and d represent ±1 s.d., as derived from a replicate pool of 20–120 embryos for each construct.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2963808&req=5

f5: Relic sites are non-functional and accumulate as the enhancer ages.(a) Graph showing the percentage of CA-dinucleotide and CAC-trinucleotide content of several orthologous enhancer sequences from D. melanogaster, D. pseudoobscura, D. willistoni and D. virilis. Each window of NEE sequence is taken ±480 bp from Dβ for each species. Each window of an A/P enhancer is a 960 bp sequence centred around the Bicoid-binding site cluster. Each orthologous set of NEEs is boxed separately to visualize enrichment relative to other groups. The red boxes show the regions occupied by all data points corresponding to a single orthologous set of NEEs located at the indicated locus across many species. The blue box shows the region occupied by all data points corresponding to all A/P enhancers for all species. (b) Identical graph as in panel a, except the data points are boxed by species to visualize genome-specific effects in satellite enrichment or depletion. Red boxes show the region occupied by all data points corresponding to all NEEs within a single species. Canonical A/P enhancers at the eve, gt, Kr and hb loci for all four species are boxed in both panels (blue rectangular area). (c) Graph showing the number of cells spanned by the lacZ expression pattern (vertical axis), as driven by NEEs containing different numbers of Dl half-sites, 5′-SGGAAW-3′ (horizontal axis). (d) Graph showing the number of cells spanned by the lacZ expression pattern (vertical axis), as driven by NEEs characterized by different E-to-D spacer lengths (horizontal axis). Error bars in c and d represent ±1 s.d., as derived from a replicate pool of 20–120 embryos for each construct.
Mentions: To address the possibility that elevated CA-satellite composition is a feature common to developmental enhancers, we looked at several embryonic enhancers that respond to the Bicoid morphogen gradient, which patterns the A/P axis. We identified complete orthologous sequence sets for the hb embryonic enhancer42, the gt posterior stripe enhancer43, the Kr central domain enhancer4445 and the eve stripe 2 enhancer46 from each of four genomes, namely, D. melanogaster, D. pseudoobscura, D. willistoni and D. virilis. All of these enhancers are active in the same embryonic nuclei as the NEEs and thus constitute a well-matched control group. We find that while the NEE set from any genome is enriched in CA-satellite dinucleotide and trinucleotide fragments, none of the 16 A/P enhancer sets possess the elevated CA-satellite levels that characterize canonical NEEs from these same species, even in genomes with elevated CA-satellite content (Fig. 5a–b).

Bottom Line: However, NEEs also possess dense clusters of variant Dl sites.Here, we show that these increasingly variant sites are eclipsed relic elements, which were superseded by more recently evolved threshold encodings.Given the divergence in egg size during Drosophila lineage evolution, the observed characteristic clusters of divergent sites indicate a history of frequent selection for changes in threshold responses to the Dl morphogen gradient and confirm the NEE structure/function model.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences, Dartmouth College, Hanover, New Hampshire 03755, USA.

ABSTRACT
Concentration gradients of morphogenic proteins pattern the embryonic axes of Drosophila by activating different genes at different concentrations. The neurogenic ectoderm enhancers (NEEs) activate different genes at different threshold levels of the Dorsal (Dl) morphogen, which patterns the dorsal/ventral axis. NEEs share a unique arrangement of highly constrained DNA-binding sites for Dl, Twist (Twi), Snail (Sna) and Suppressor of Hairless (Su(H)), and encode the threshold variable in the precise length of DNA that separates one well-defined Dl element from a Twi element. However, NEEs also possess dense clusters of variant Dl sites. Here, we show that these increasingly variant sites are eclipsed relic elements, which were superseded by more recently evolved threshold encodings. Given the divergence in egg size during Drosophila lineage evolution, the observed characteristic clusters of divergent sites indicate a history of frequent selection for changes in threshold responses to the Dl morphogen gradient and confirm the NEE structure/function model.

Show MeSH