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cis-regulatory analysis of the Drosophila pdm locus reveals a diversity of neural enhancers.

Ross J, Kuzin A, Brody T, Odenwald WF - BMC Genomics (2015)

Bottom Line: To facilitate data accessibility, the results of our analysis are catalogued in cisPatterns, an online database of the structure and function of these and other Drosophila enhancers.A subset of these enhancers share conserved elements including sequences that correspond to known TF DNA binding sites.Although comparative analysis of the nubbin and pdm-2 encoding sequences indicate that these two genes most likely arose from a duplication event, we found only partial evidence of sequence duplication between their enhancers, suggesting that after the putative duplication their cis-regulatory DNA diverged at a higher rate than their coding sequences.

View Article: PubMed Central - PubMed

Affiliation: The Neural Cell-Fate Determinants Section, NINDS, NIH, Bethesda, MD, USA. rossje@ninds.nih.gov.

ABSTRACT

Background: One of the major challenges in developmental biology is to understand the regulatory events that generate neuronal diversity. During Drosophila embryonic neural lineage development, cellular temporal identity is established in part by a transcription factor (TF) regulatory network that mediates a cascade of cellular identity decisions. Two of the regulators essential to this network are the POU-domain TFs Nubbin and Pdm-2, encoded by adjacent genes collectively known as pdm. The focus of this study is the discovery and characterization of cis-regulatory DNA that governs their expression.

Results: Phylogenetic footprinting analysis of a 125 kb genomic region that spans the pdm locus identified 116 conserved sequence clusters. To determine which of these regions function as cis-regulatory enhancers that regulate the dynamics of pdm gene expression, we tested each for in vivo enhancer activity during embryonic development and postembryonic neurogenesis. Our screen revealed 77 unique enhancers positioned throughout the noncoding region of the pdm locus. Many of these activated neural-specific gene expression during different developmental stages and many drove expression in overlapping patterns. Sequence comparisons of functionally related enhancers that activate overlapping expression patterns revealed that they share conserved elements that can be predictive of enhancer behavior. To facilitate data accessibility, the results of our analysis are catalogued in cisPatterns, an online database of the structure and function of these and other Drosophila enhancers.

Conclusions: These studies reveal a diversity of modular enhancers that most likely regulate pdm gene expression during embryonic and adult development, highlighting a high level of temporal and spatial expression specificity. In addition, we discovered clusters of functionally related enhancers throughout the pdm locus. A subset of these enhancers share conserved elements including sequences that correspond to known TF DNA binding sites. Although comparative analysis of the nubbin and pdm-2 encoding sequences indicate that these two genes most likely arose from a duplication event, we found only partial evidence of sequence duplication between their enhancers, suggesting that after the putative duplication their cis-regulatory DNA diverged at a higher rate than their coding sequences.

No MeSH data available.


pdm locus enhancers drives expression in specific classes of larval NBs. a–dpdm enhancers activate reporter expression during larval NB lineage development. Shown is membrane-bound GFP (mCD8-GFP) expression (green) driven by each enhancer. a The nub-46 enhancer regulates central brain and VNC expression. a’ The nub-46 enhancer regulates expression in type I and type II NBs. Shown is a single confocal plane view of a larval brain lobe stained with anti-GFP (green), anti-Ase (red), and anti-Dpn (blue). a” The inset is a magnified view of the yellow dashed square and highlights type I NBs (Ase+ Dpn+, asterisk), type II NBs (Ase− Dpn+, arrowheads), GMCs (Ase+ Dpn−, arrows) and individual NB lineages (yellow dashed outlines). b The nub-49b enhancer regulates a subset of optic lobe, central brain, and VNC NB lineages. b’ and b” show that nub-49b directs expression in type I and type II NBs. c The pdm2-34 enhancer directs central brain and VNC expression. c’ and c” reveals that pdm2-34 drives expression in type I NBs. d–d” The pdm-37a enhancer activates expression in type I and type II larval NBs
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Fig8: pdm locus enhancers drives expression in specific classes of larval NBs. a–dpdm enhancers activate reporter expression during larval NB lineage development. Shown is membrane-bound GFP (mCD8-GFP) expression (green) driven by each enhancer. a The nub-46 enhancer regulates central brain and VNC expression. a’ The nub-46 enhancer regulates expression in type I and type II NBs. Shown is a single confocal plane view of a larval brain lobe stained with anti-GFP (green), anti-Ase (red), and anti-Dpn (blue). a” The inset is a magnified view of the yellow dashed square and highlights type I NBs (Ase+ Dpn+, asterisk), type II NBs (Ase− Dpn+, arrowheads), GMCs (Ase+ Dpn−, arrows) and individual NB lineages (yellow dashed outlines). b The nub-49b enhancer regulates a subset of optic lobe, central brain, and VNC NB lineages. b’ and b” show that nub-49b directs expression in type I and type II NBs. c The pdm2-34 enhancer directs central brain and VNC expression. c’ and c” reveals that pdm2-34 drives expression in type I NBs. d–d” The pdm-37a enhancer activates expression in type I and type II larval NBs

Mentions: A subset of enhancers drove expression in putative larval NBs. For example, the two embryonic NB enhancers for nub and pdm-2, nub-46 and pdm2-34, are also active in larval NBs (Fig. 8a, c, respectively). In addition, nub-49b and pdm2-37a also regulated expression during larval NB lineage development (Fig. 8b, d).Fig. 8


cis-regulatory analysis of the Drosophila pdm locus reveals a diversity of neural enhancers.

Ross J, Kuzin A, Brody T, Odenwald WF - BMC Genomics (2015)

pdm locus enhancers drives expression in specific classes of larval NBs. a–dpdm enhancers activate reporter expression during larval NB lineage development. Shown is membrane-bound GFP (mCD8-GFP) expression (green) driven by each enhancer. a The nub-46 enhancer regulates central brain and VNC expression. a’ The nub-46 enhancer regulates expression in type I and type II NBs. Shown is a single confocal plane view of a larval brain lobe stained with anti-GFP (green), anti-Ase (red), and anti-Dpn (blue). a” The inset is a magnified view of the yellow dashed square and highlights type I NBs (Ase+ Dpn+, asterisk), type II NBs (Ase− Dpn+, arrowheads), GMCs (Ase+ Dpn−, arrows) and individual NB lineages (yellow dashed outlines). b The nub-49b enhancer regulates a subset of optic lobe, central brain, and VNC NB lineages. b’ and b” show that nub-49b directs expression in type I and type II NBs. c The pdm2-34 enhancer directs central brain and VNC expression. c’ and c” reveals that pdm2-34 drives expression in type I NBs. d–d” The pdm-37a enhancer activates expression in type I and type II larval NBs
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

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Fig8: pdm locus enhancers drives expression in specific classes of larval NBs. a–dpdm enhancers activate reporter expression during larval NB lineage development. Shown is membrane-bound GFP (mCD8-GFP) expression (green) driven by each enhancer. a The nub-46 enhancer regulates central brain and VNC expression. a’ The nub-46 enhancer regulates expression in type I and type II NBs. Shown is a single confocal plane view of a larval brain lobe stained with anti-GFP (green), anti-Ase (red), and anti-Dpn (blue). a” The inset is a magnified view of the yellow dashed square and highlights type I NBs (Ase+ Dpn+, asterisk), type II NBs (Ase− Dpn+, arrowheads), GMCs (Ase+ Dpn−, arrows) and individual NB lineages (yellow dashed outlines). b The nub-49b enhancer regulates a subset of optic lobe, central brain, and VNC NB lineages. b’ and b” show that nub-49b directs expression in type I and type II NBs. c The pdm2-34 enhancer directs central brain and VNC expression. c’ and c” reveals that pdm2-34 drives expression in type I NBs. d–d” The pdm-37a enhancer activates expression in type I and type II larval NBs
Mentions: A subset of enhancers drove expression in putative larval NBs. For example, the two embryonic NB enhancers for nub and pdm-2, nub-46 and pdm2-34, are also active in larval NBs (Fig. 8a, c, respectively). In addition, nub-49b and pdm2-37a also regulated expression during larval NB lineage development (Fig. 8b, d).Fig. 8

Bottom Line: To facilitate data accessibility, the results of our analysis are catalogued in cisPatterns, an online database of the structure and function of these and other Drosophila enhancers.A subset of these enhancers share conserved elements including sequences that correspond to known TF DNA binding sites.Although comparative analysis of the nubbin and pdm-2 encoding sequences indicate that these two genes most likely arose from a duplication event, we found only partial evidence of sequence duplication between their enhancers, suggesting that after the putative duplication their cis-regulatory DNA diverged at a higher rate than their coding sequences.

View Article: PubMed Central - PubMed

Affiliation: The Neural Cell-Fate Determinants Section, NINDS, NIH, Bethesda, MD, USA. rossje@ninds.nih.gov.

ABSTRACT

Background: One of the major challenges in developmental biology is to understand the regulatory events that generate neuronal diversity. During Drosophila embryonic neural lineage development, cellular temporal identity is established in part by a transcription factor (TF) regulatory network that mediates a cascade of cellular identity decisions. Two of the regulators essential to this network are the POU-domain TFs Nubbin and Pdm-2, encoded by adjacent genes collectively known as pdm. The focus of this study is the discovery and characterization of cis-regulatory DNA that governs their expression.

Results: Phylogenetic footprinting analysis of a 125 kb genomic region that spans the pdm locus identified 116 conserved sequence clusters. To determine which of these regions function as cis-regulatory enhancers that regulate the dynamics of pdm gene expression, we tested each for in vivo enhancer activity during embryonic development and postembryonic neurogenesis. Our screen revealed 77 unique enhancers positioned throughout the noncoding region of the pdm locus. Many of these activated neural-specific gene expression during different developmental stages and many drove expression in overlapping patterns. Sequence comparisons of functionally related enhancers that activate overlapping expression patterns revealed that they share conserved elements that can be predictive of enhancer behavior. To facilitate data accessibility, the results of our analysis are catalogued in cisPatterns, an online database of the structure and function of these and other Drosophila enhancers.

Conclusions: These studies reveal a diversity of modular enhancers that most likely regulate pdm gene expression during embryonic and adult development, highlighting a high level of temporal and spatial expression specificity. In addition, we discovered clusters of functionally related enhancers throughout the pdm locus. A subset of these enhancers share conserved elements including sequences that correspond to known TF DNA binding sites. Although comparative analysis of the nubbin and pdm-2 encoding sequences indicate that these two genes most likely arose from a duplication event, we found only partial evidence of sequence duplication between their enhancers, suggesting that after the putative duplication their cis-regulatory DNA diverged at a higher rate than their coding sequences.

No MeSH data available.