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Positional cues in the Drosophila nerve cord: semaphorins pattern the dorso-ventral axis.

Zlatic M, Li F, Strigini M, Grueber W, Bate M - PLoS Biol. (2009)

Bottom Line: The range of alternative targets is reduced by well organised patterns of growth, termination, and branching that deliver the terminals of appropriate pre- and postsynaptic partners to restricted volumes of the developing nervous system.We show that a combination of Plexin A (Plex A) and Plexin B (Plex B) receptors specifies the ventral projection of sensory neurons by responding to high concentrations of Semaphorin 1a (Sema 1a) and Semaphorin 2a (Sema 2a).Together our findings support the idea that axons are delivered to particular regions of the neuropile by their responses to systems of positional cues in each dimension.

View Article: PubMed Central - PubMed

Affiliation: Department of Zoology, University of Cambridge, Cambridge, United Kingdom. zlaticm@janelia.hhmi.org

ABSTRACT
During the development of neural circuitry, neurons of different kinds establish specific synaptic connections by selecting appropriate targets from large numbers of alternatives. The range of alternative targets is reduced by well organised patterns of growth, termination, and branching that deliver the terminals of appropriate pre- and postsynaptic partners to restricted volumes of the developing nervous system. We use the axons of embryonic Drosophila sensory neurons as a model system in which to study the way in which growing neurons are guided to terminate in specific volumes of the developing nervous system. The mediolateral positions of sensory arbors are controlled by the response of Robo receptors to a Slit gradient. Here we make a genetic analysis of factors regulating position in the dorso-ventral axis. We find that dorso-ventral layers of neuropile contain different levels and combinations of Semaphorins. We demonstrate the existence of a central to dorsal and central to ventral gradient of Sema 2a, perpendicular to the Slit gradient. We show that a combination of Plexin A (Plex A) and Plexin B (Plex B) receptors specifies the ventral projection of sensory neurons by responding to high concentrations of Semaphorin 1a (Sema 1a) and Semaphorin 2a (Sema 2a). Together our findings support the idea that axons are delivered to particular regions of the neuropile by their responses to systems of positional cues in each dimension.

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sema 2a and sema 1a mutations suppress the phenotypes of Plex B and Plex A overexpression in sensory neurons.(A–D) Representative images of sensory terminals labelled with PO163GAL4, UAS-n-syb-GFP (green) with respect to Fas II tracts (red) in 21-h embryos (left) and diagrams showing patterns of sensory terminals in different genotypes superimposed (right). In all cases images show projections of a confocal z series of transverse sections through A7. Dorsal is up. Arrowheads show midline. White lines, layer boundaries. Numbers indicate layers: M, medial; I, intermediate; L, lateral domains. Scale bar: 10 µm. (A) In sema 2a03021 mutant embryos, sensory terminals aberrantly invade neuropile layer 2 and to a lesser extent layer 1. Right: Diagram showing the pattern of sensory terminals in sema 2a03021 mutant (green) and (yellow) embryos, superimposed. Quantification of SA in layer 2 in sema 2a03021 mutant embryos reveals a significant increase (***, p = 5×10−5; Student's t-test; average SA2/h = 0.1; SD = 0.05; n = 24 hemisegments) with respect to wild type (average SA2/h = 0.06; SD = 0.02; n = 32 hemisegments). (B) Expressing Plex B in sensory neurons in a sema 2a03021 mutant background fails to exclude sensory terminals from neuropile layer 2. Right: Diagram showing the patterns of Plex B expressing sensory terminals in sema 2a03021 (green) mutant and wild-type (yellow) backgrounds, superimposed. Quantification of SA in layer 2 reveals a significant increase (***, p = 10×10−12; Student's t-test; average SA2/h = 0.06 and SD = 0.03, n = 31 hemisegments), with respect to Plex B expression in wild-type background (average SA2/h = 0.003; SD = 0.006, n = 24 hemisegments). (C) In sema 1aP1 mutant embryos, sensory terminals aberrantly invade neuropile layer 1 and to a lesser extent layers 2 and 3. This results in an large increase in the surface area occupied by sensory neuron terminals in layers 1 and 3. Right: Diagram showing the patterns of sensory terminals in sema 1aP1 mutant (green) and (yellow) embryos, superimposed. Quantification of SA1+3+4/h reveals a significant increase (***, p = 5×10−15; Student's t-test) in sema 1aP1 mutants (average SA1+3+4/h = 0.6 and SD = 0.09, n = 23 hemisegments), compared to wild type (average SA1+3+4/h = 0.3; SD = 0.05, n = 32 hemisegments). (D) Expressing Plex A in sensory neurons in a sema 1aP1 background fails to exclude sensory terminals from neuropile layers 1 and 3. Right: Diagram showing the patterns of sensory terminals that express Plex A in sema 1aP1 mutant (green) and wild-type (yellow) backgrounds, superimposed. Quantification of SA1+3+4/h reveals a significant increase (***, p = 2×10−25; Student's t-test) in sema 1aP1 mutant (average SA1+3+4/h = 0.5 and SD = 0.09, n = 30 hemisegments), compared to wild-type (average SA1+3+4/h = 0.1; SD = 0.04; n = 38) backgrounds.
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pbio-1000135-g003: sema 2a and sema 1a mutations suppress the phenotypes of Plex B and Plex A overexpression in sensory neurons.(A–D) Representative images of sensory terminals labelled with PO163GAL4, UAS-n-syb-GFP (green) with respect to Fas II tracts (red) in 21-h embryos (left) and diagrams showing patterns of sensory terminals in different genotypes superimposed (right). In all cases images show projections of a confocal z series of transverse sections through A7. Dorsal is up. Arrowheads show midline. White lines, layer boundaries. Numbers indicate layers: M, medial; I, intermediate; L, lateral domains. Scale bar: 10 µm. (A) In sema 2a03021 mutant embryos, sensory terminals aberrantly invade neuropile layer 2 and to a lesser extent layer 1. Right: Diagram showing the pattern of sensory terminals in sema 2a03021 mutant (green) and (yellow) embryos, superimposed. Quantification of SA in layer 2 in sema 2a03021 mutant embryos reveals a significant increase (***, p = 5×10−5; Student's t-test; average SA2/h = 0.1; SD = 0.05; n = 24 hemisegments) with respect to wild type (average SA2/h = 0.06; SD = 0.02; n = 32 hemisegments). (B) Expressing Plex B in sensory neurons in a sema 2a03021 mutant background fails to exclude sensory terminals from neuropile layer 2. Right: Diagram showing the patterns of Plex B expressing sensory terminals in sema 2a03021 (green) mutant and wild-type (yellow) backgrounds, superimposed. Quantification of SA in layer 2 reveals a significant increase (***, p = 10×10−12; Student's t-test; average SA2/h = 0.06 and SD = 0.03, n = 31 hemisegments), with respect to Plex B expression in wild-type background (average SA2/h = 0.003; SD = 0.006, n = 24 hemisegments). (C) In sema 1aP1 mutant embryos, sensory terminals aberrantly invade neuropile layer 1 and to a lesser extent layers 2 and 3. This results in an large increase in the surface area occupied by sensory neuron terminals in layers 1 and 3. Right: Diagram showing the patterns of sensory terminals in sema 1aP1 mutant (green) and (yellow) embryos, superimposed. Quantification of SA1+3+4/h reveals a significant increase (***, p = 5×10−15; Student's t-test) in sema 1aP1 mutants (average SA1+3+4/h = 0.6 and SD = 0.09, n = 23 hemisegments), compared to wild type (average SA1+3+4/h = 0.3; SD = 0.05, n = 32 hemisegments). (D) Expressing Plex A in sensory neurons in a sema 1aP1 background fails to exclude sensory terminals from neuropile layers 1 and 3. Right: Diagram showing the patterns of sensory terminals that express Plex A in sema 1aP1 mutant (green) and wild-type (yellow) backgrounds, superimposed. Quantification of SA1+3+4/h reveals a significant increase (***, p = 2×10−25; Student's t-test) in sema 1aP1 mutant (average SA1+3+4/h = 0.5 and SD = 0.09, n = 30 hemisegments), compared to wild-type (average SA1+3+4/h = 0.1; SD = 0.04; n = 38) backgrounds.

Mentions: We analysed patterns of sensory terminals in sema 2a03021 loss of function embryos [25] and in embryos in which plex B was overexpressed in sensory neurons in a sema 2a03021 background. In sema 2a03021 embryos we find ectopic sensory terminals in layer 2 (Figure 3A). Overexpression of Plex B in sensory neurons in a sema 2a03021 background fails to exclude sensory terminals from central and dorsal neuropile (compare Figures 2C and 3B). The pattern of sensory terminals in these embryos is similar to their pattern in sema 2a03021 mutants (compare Figure 3A and 3B). We conclude that Sema 2a is the functional ligand for Plex B in this system.


Positional cues in the Drosophila nerve cord: semaphorins pattern the dorso-ventral axis.

Zlatic M, Li F, Strigini M, Grueber W, Bate M - PLoS Biol. (2009)

sema 2a and sema 1a mutations suppress the phenotypes of Plex B and Plex A overexpression in sensory neurons.(A–D) Representative images of sensory terminals labelled with PO163GAL4, UAS-n-syb-GFP (green) with respect to Fas II tracts (red) in 21-h embryos (left) and diagrams showing patterns of sensory terminals in different genotypes superimposed (right). In all cases images show projections of a confocal z series of transverse sections through A7. Dorsal is up. Arrowheads show midline. White lines, layer boundaries. Numbers indicate layers: M, medial; I, intermediate; L, lateral domains. Scale bar: 10 µm. (A) In sema 2a03021 mutant embryos, sensory terminals aberrantly invade neuropile layer 2 and to a lesser extent layer 1. Right: Diagram showing the pattern of sensory terminals in sema 2a03021 mutant (green) and (yellow) embryos, superimposed. Quantification of SA in layer 2 in sema 2a03021 mutant embryos reveals a significant increase (***, p = 5×10−5; Student's t-test; average SA2/h = 0.1; SD = 0.05; n = 24 hemisegments) with respect to wild type (average SA2/h = 0.06; SD = 0.02; n = 32 hemisegments). (B) Expressing Plex B in sensory neurons in a sema 2a03021 mutant background fails to exclude sensory terminals from neuropile layer 2. Right: Diagram showing the patterns of Plex B expressing sensory terminals in sema 2a03021 (green) mutant and wild-type (yellow) backgrounds, superimposed. Quantification of SA in layer 2 reveals a significant increase (***, p = 10×10−12; Student's t-test; average SA2/h = 0.06 and SD = 0.03, n = 31 hemisegments), with respect to Plex B expression in wild-type background (average SA2/h = 0.003; SD = 0.006, n = 24 hemisegments). (C) In sema 1aP1 mutant embryos, sensory terminals aberrantly invade neuropile layer 1 and to a lesser extent layers 2 and 3. This results in an large increase in the surface area occupied by sensory neuron terminals in layers 1 and 3. Right: Diagram showing the patterns of sensory terminals in sema 1aP1 mutant (green) and (yellow) embryos, superimposed. Quantification of SA1+3+4/h reveals a significant increase (***, p = 5×10−15; Student's t-test) in sema 1aP1 mutants (average SA1+3+4/h = 0.6 and SD = 0.09, n = 23 hemisegments), compared to wild type (average SA1+3+4/h = 0.3; SD = 0.05, n = 32 hemisegments). (D) Expressing Plex A in sensory neurons in a sema 1aP1 background fails to exclude sensory terminals from neuropile layers 1 and 3. Right: Diagram showing the patterns of sensory terminals that express Plex A in sema 1aP1 mutant (green) and wild-type (yellow) backgrounds, superimposed. Quantification of SA1+3+4/h reveals a significant increase (***, p = 2×10−25; Student's t-test) in sema 1aP1 mutant (average SA1+3+4/h = 0.5 and SD = 0.09, n = 30 hemisegments), compared to wild-type (average SA1+3+4/h = 0.1; SD = 0.04; n = 38) backgrounds.
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pbio-1000135-g003: sema 2a and sema 1a mutations suppress the phenotypes of Plex B and Plex A overexpression in sensory neurons.(A–D) Representative images of sensory terminals labelled with PO163GAL4, UAS-n-syb-GFP (green) with respect to Fas II tracts (red) in 21-h embryos (left) and diagrams showing patterns of sensory terminals in different genotypes superimposed (right). In all cases images show projections of a confocal z series of transverse sections through A7. Dorsal is up. Arrowheads show midline. White lines, layer boundaries. Numbers indicate layers: M, medial; I, intermediate; L, lateral domains. Scale bar: 10 µm. (A) In sema 2a03021 mutant embryos, sensory terminals aberrantly invade neuropile layer 2 and to a lesser extent layer 1. Right: Diagram showing the pattern of sensory terminals in sema 2a03021 mutant (green) and (yellow) embryos, superimposed. Quantification of SA in layer 2 in sema 2a03021 mutant embryos reveals a significant increase (***, p = 5×10−5; Student's t-test; average SA2/h = 0.1; SD = 0.05; n = 24 hemisegments) with respect to wild type (average SA2/h = 0.06; SD = 0.02; n = 32 hemisegments). (B) Expressing Plex B in sensory neurons in a sema 2a03021 mutant background fails to exclude sensory terminals from neuropile layer 2. Right: Diagram showing the patterns of Plex B expressing sensory terminals in sema 2a03021 (green) mutant and wild-type (yellow) backgrounds, superimposed. Quantification of SA in layer 2 reveals a significant increase (***, p = 10×10−12; Student's t-test; average SA2/h = 0.06 and SD = 0.03, n = 31 hemisegments), with respect to Plex B expression in wild-type background (average SA2/h = 0.003; SD = 0.006, n = 24 hemisegments). (C) In sema 1aP1 mutant embryos, sensory terminals aberrantly invade neuropile layer 1 and to a lesser extent layers 2 and 3. This results in an large increase in the surface area occupied by sensory neuron terminals in layers 1 and 3. Right: Diagram showing the patterns of sensory terminals in sema 1aP1 mutant (green) and (yellow) embryos, superimposed. Quantification of SA1+3+4/h reveals a significant increase (***, p = 5×10−15; Student's t-test) in sema 1aP1 mutants (average SA1+3+4/h = 0.6 and SD = 0.09, n = 23 hemisegments), compared to wild type (average SA1+3+4/h = 0.3; SD = 0.05, n = 32 hemisegments). (D) Expressing Plex A in sensory neurons in a sema 1aP1 background fails to exclude sensory terminals from neuropile layers 1 and 3. Right: Diagram showing the patterns of sensory terminals that express Plex A in sema 1aP1 mutant (green) and wild-type (yellow) backgrounds, superimposed. Quantification of SA1+3+4/h reveals a significant increase (***, p = 2×10−25; Student's t-test) in sema 1aP1 mutant (average SA1+3+4/h = 0.5 and SD = 0.09, n = 30 hemisegments), compared to wild-type (average SA1+3+4/h = 0.1; SD = 0.04; n = 38) backgrounds.
Mentions: We analysed patterns of sensory terminals in sema 2a03021 loss of function embryos [25] and in embryos in which plex B was overexpressed in sensory neurons in a sema 2a03021 background. In sema 2a03021 embryos we find ectopic sensory terminals in layer 2 (Figure 3A). Overexpression of Plex B in sensory neurons in a sema 2a03021 background fails to exclude sensory terminals from central and dorsal neuropile (compare Figures 2C and 3B). The pattern of sensory terminals in these embryos is similar to their pattern in sema 2a03021 mutants (compare Figure 3A and 3B). We conclude that Sema 2a is the functional ligand for Plex B in this system.

Bottom Line: The range of alternative targets is reduced by well organised patterns of growth, termination, and branching that deliver the terminals of appropriate pre- and postsynaptic partners to restricted volumes of the developing nervous system.We show that a combination of Plexin A (Plex A) and Plexin B (Plex B) receptors specifies the ventral projection of sensory neurons by responding to high concentrations of Semaphorin 1a (Sema 1a) and Semaphorin 2a (Sema 2a).Together our findings support the idea that axons are delivered to particular regions of the neuropile by their responses to systems of positional cues in each dimension.

View Article: PubMed Central - PubMed

Affiliation: Department of Zoology, University of Cambridge, Cambridge, United Kingdom. zlaticm@janelia.hhmi.org

ABSTRACT
During the development of neural circuitry, neurons of different kinds establish specific synaptic connections by selecting appropriate targets from large numbers of alternatives. The range of alternative targets is reduced by well organised patterns of growth, termination, and branching that deliver the terminals of appropriate pre- and postsynaptic partners to restricted volumes of the developing nervous system. We use the axons of embryonic Drosophila sensory neurons as a model system in which to study the way in which growing neurons are guided to terminate in specific volumes of the developing nervous system. The mediolateral positions of sensory arbors are controlled by the response of Robo receptors to a Slit gradient. Here we make a genetic analysis of factors regulating position in the dorso-ventral axis. We find that dorso-ventral layers of neuropile contain different levels and combinations of Semaphorins. We demonstrate the existence of a central to dorsal and central to ventral gradient of Sema 2a, perpendicular to the Slit gradient. We show that a combination of Plexin A (Plex A) and Plexin B (Plex B) receptors specifies the ventral projection of sensory neurons by responding to high concentrations of Semaphorin 1a (Sema 1a) and Semaphorin 2a (Sema 2a). Together our findings support the idea that axons are delivered to particular regions of the neuropile by their responses to systems of positional cues in each dimension.

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