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An arthropod cis-regulatory element functioning in sensory organ precursor development dates back to the Cambrian.

Ayyar S, Negre B, Simpson P, Stollewerk A - BMC Biol. (2010)

Bottom Line: The SOPEs of the spider Cupiennius salei and the insect Tribolium castaneum are shown to be functional in transgenic Drosophila.This would place the origin of this regulatory sequence as far back as the last common ancestor of the Arthropoda, that is, in the Cambrian, 550 million years ago.The SOPE is not detectable by inter-specific sequence comparison, raising the possibility that other ancient regulatory modules in invertebrates might have escaped detection.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Zoology, University of Cambridge, Downing Street, Cambridge CB2 3EJ, UK.

ABSTRACT

Background: An increasing number of publications demonstrate conservation of function of cis-regulatory elements without sequence similarity. In invertebrates such functional conservation has only been shown for closely related species. Here we demonstrate the existence of an ancient arthropod regulatory element that functions during the selection of neural precursors. The activity of genes of the achaete-scute (ac-sc) family endows cells with neural potential. An essential, conserved characteristic of proneural genes is their ability to restrict their own activity to single or a small number of progenitor cells from their initially broad domains of expression. This is achieved through a process called lateral inhibition. A regulatory element, the sensory organ precursor enhancer (SOPE), is required for this process. First identified in Drosophila, the SOPE contains discrete binding sites for four regulatory factors. The SOPE of the Drosophila asense gene is situated in the 5' UTR.

Results: Through a manual comparison of consensus binding site sequences we have been able to identify a SOPE in UTR sequences of asense-like genes in species belonging to all four arthropod groups (Crustacea, Myriapoda, Chelicerata and Insecta). The SOPEs of the spider Cupiennius salei and the insect Tribolium castaneum are shown to be functional in transgenic Drosophila. This would place the origin of this regulatory sequence as far back as the last common ancestor of the Arthropoda, that is, in the Cambrian, 550 million years ago.

Conclusions: The SOPE is not detectable by inter-specific sequence comparison, raising the possibility that other ancient regulatory modules in invertebrates might have escaped detection.

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Comparison of the number of ectopic bristles (macrochaetes) displayed by transgenic flies after ectopic expression. UAS constructs containing D. melanogaster ase, T. castenum ase and C. salei CsASH2 (ORF alone or the entire transcribed region (ORF+SOPE) of ase or CsASH2) were each crossed to four different Gal4 drivers allowing expression in different parts of the thorax (see Materials and methods). (A) The number of ectopic bristles was counted in the respective Gal4 expression domains of each of the driver lines. Columns give the number of ectopic bristles and data from all four crosses have been pooled for each UAS construct (UAS-ORF+SOPE or UAS-ORF). The number of ectopic bristles is significantly reduced in flies carrying the UAS-ORF+SOPE constructs (see Additional file 4 for details). The error bars give the standard error of the mean. (B-G) Thoraces illustrating the phenotypes obtained: (B, C) ptc-Gal4 > UAS Dm-ase; (D, E) sca-Gal4 > UAS Tc-ase; (F, G) MD806-Gal4 > UAS CsASH2.
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Figure 5: Comparison of the number of ectopic bristles (macrochaetes) displayed by transgenic flies after ectopic expression. UAS constructs containing D. melanogaster ase, T. castenum ase and C. salei CsASH2 (ORF alone or the entire transcribed region (ORF+SOPE) of ase or CsASH2) were each crossed to four different Gal4 drivers allowing expression in different parts of the thorax (see Materials and methods). (A) The number of ectopic bristles was counted in the respective Gal4 expression domains of each of the driver lines. Columns give the number of ectopic bristles and data from all four crosses have been pooled for each UAS construct (UAS-ORF+SOPE or UAS-ORF). The number of ectopic bristles is significantly reduced in flies carrying the UAS-ORF+SOPE constructs (see Additional file 4 for details). The error bars give the standard error of the mean. (B-G) Thoraces illustrating the phenotypes obtained: (B, C) ptc-Gal4 > UAS Dm-ase; (D, E) sca-Gal4 > UAS Tc-ase; (F, G) MD806-Gal4 > UAS CsASH2.

Mentions: The Dm-ase SOPE had been shown to display enhancer activity when placed upstream of an hsp70 promoter and a reporter gene [13]. To test its effects when positioned in the UTR, we generated transgenic lines carrying UAS constructs containing either the entire transcribed region (including the UTR sequences) or merely the ORF. Since reporter gene fusion constructs that cover different regions upstream of the ORF only restrict expression to single SOPs if the 560-bp UTR containing the SOPE is present [13], the ORF+SOPE constructs should reduce the number of bristles. Three independent lines of each construct were crossed to four different Gal4 lines, each of which drives expression in all or part of the D. melanogaster notum. As expected, both transgenes caused the development of ectopic bristles but their number was significantly reduced in the construct containing the entire UTR. Flies expressing the UAS-Dm-ase ORF displayed, in total, an average of 10.9 ectopic bristles, compared with 7.3 in flies expressing the UAS-Dm-ase ORF+SOPE (Figure 5A-C; Additional file 4). We therefore conclude that the SOPE regulates gene activity from its position in the UTR and that, when transcription is initiated from exogenous UAS sequences, it functions to dampen transcription. This is consistent with its proposed function to restrict proneural gene activity from broad expression domains to single neural progenitors.


An arthropod cis-regulatory element functioning in sensory organ precursor development dates back to the Cambrian.

Ayyar S, Negre B, Simpson P, Stollewerk A - BMC Biol. (2010)

Comparison of the number of ectopic bristles (macrochaetes) displayed by transgenic flies after ectopic expression. UAS constructs containing D. melanogaster ase, T. castenum ase and C. salei CsASH2 (ORF alone or the entire transcribed region (ORF+SOPE) of ase or CsASH2) were each crossed to four different Gal4 drivers allowing expression in different parts of the thorax (see Materials and methods). (A) The number of ectopic bristles was counted in the respective Gal4 expression domains of each of the driver lines. Columns give the number of ectopic bristles and data from all four crosses have been pooled for each UAS construct (UAS-ORF+SOPE or UAS-ORF). The number of ectopic bristles is significantly reduced in flies carrying the UAS-ORF+SOPE constructs (see Additional file 4 for details). The error bars give the standard error of the mean. (B-G) Thoraces illustrating the phenotypes obtained: (B, C) ptc-Gal4 > UAS Dm-ase; (D, E) sca-Gal4 > UAS Tc-ase; (F, G) MD806-Gal4 > UAS CsASH2.
© Copyright Policy - open-access
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2958161&req=5

Figure 5: Comparison of the number of ectopic bristles (macrochaetes) displayed by transgenic flies after ectopic expression. UAS constructs containing D. melanogaster ase, T. castenum ase and C. salei CsASH2 (ORF alone or the entire transcribed region (ORF+SOPE) of ase or CsASH2) were each crossed to four different Gal4 drivers allowing expression in different parts of the thorax (see Materials and methods). (A) The number of ectopic bristles was counted in the respective Gal4 expression domains of each of the driver lines. Columns give the number of ectopic bristles and data from all four crosses have been pooled for each UAS construct (UAS-ORF+SOPE or UAS-ORF). The number of ectopic bristles is significantly reduced in flies carrying the UAS-ORF+SOPE constructs (see Additional file 4 for details). The error bars give the standard error of the mean. (B-G) Thoraces illustrating the phenotypes obtained: (B, C) ptc-Gal4 > UAS Dm-ase; (D, E) sca-Gal4 > UAS Tc-ase; (F, G) MD806-Gal4 > UAS CsASH2.
Mentions: The Dm-ase SOPE had been shown to display enhancer activity when placed upstream of an hsp70 promoter and a reporter gene [13]. To test its effects when positioned in the UTR, we generated transgenic lines carrying UAS constructs containing either the entire transcribed region (including the UTR sequences) or merely the ORF. Since reporter gene fusion constructs that cover different regions upstream of the ORF only restrict expression to single SOPs if the 560-bp UTR containing the SOPE is present [13], the ORF+SOPE constructs should reduce the number of bristles. Three independent lines of each construct were crossed to four different Gal4 lines, each of which drives expression in all or part of the D. melanogaster notum. As expected, both transgenes caused the development of ectopic bristles but their number was significantly reduced in the construct containing the entire UTR. Flies expressing the UAS-Dm-ase ORF displayed, in total, an average of 10.9 ectopic bristles, compared with 7.3 in flies expressing the UAS-Dm-ase ORF+SOPE (Figure 5A-C; Additional file 4). We therefore conclude that the SOPE regulates gene activity from its position in the UTR and that, when transcription is initiated from exogenous UAS sequences, it functions to dampen transcription. This is consistent with its proposed function to restrict proneural gene activity from broad expression domains to single neural progenitors.

Bottom Line: The SOPEs of the spider Cupiennius salei and the insect Tribolium castaneum are shown to be functional in transgenic Drosophila.This would place the origin of this regulatory sequence as far back as the last common ancestor of the Arthropoda, that is, in the Cambrian, 550 million years ago.The SOPE is not detectable by inter-specific sequence comparison, raising the possibility that other ancient regulatory modules in invertebrates might have escaped detection.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Zoology, University of Cambridge, Downing Street, Cambridge CB2 3EJ, UK.

ABSTRACT

Background: An increasing number of publications demonstrate conservation of function of cis-regulatory elements without sequence similarity. In invertebrates such functional conservation has only been shown for closely related species. Here we demonstrate the existence of an ancient arthropod regulatory element that functions during the selection of neural precursors. The activity of genes of the achaete-scute (ac-sc) family endows cells with neural potential. An essential, conserved characteristic of proneural genes is their ability to restrict their own activity to single or a small number of progenitor cells from their initially broad domains of expression. This is achieved through a process called lateral inhibition. A regulatory element, the sensory organ precursor enhancer (SOPE), is required for this process. First identified in Drosophila, the SOPE contains discrete binding sites for four regulatory factors. The SOPE of the Drosophila asense gene is situated in the 5' UTR.

Results: Through a manual comparison of consensus binding site sequences we have been able to identify a SOPE in UTR sequences of asense-like genes in species belonging to all four arthropod groups (Crustacea, Myriapoda, Chelicerata and Insecta). The SOPEs of the spider Cupiennius salei and the insect Tribolium castaneum are shown to be functional in transgenic Drosophila. This would place the origin of this regulatory sequence as far back as the last common ancestor of the Arthropoda, that is, in the Cambrian, 550 million years ago.

Conclusions: The SOPE is not detectable by inter-specific sequence comparison, raising the possibility that other ancient regulatory modules in invertebrates might have escaped detection.

Show MeSH
Related in: MedlinePlus