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Orthodenticle Is Required for the Expression of Principal Recognition Molecules That Control Axon Targeting in the Drosophila Retina.

Mencarelli C, Pichaud F - PLoS Genet. (2015)

Bottom Line: Our data indicate that otd function in these photoreceptors is largely mediated by the recognition molecules flamingo (fmi) and golden goal (gogo).In addition, we find that otd regulates synaptic-layer targeting of R8.Our work therefore demonstrates that otd is a main component of the gene regulatory network that regulates synaptic-column and layer targeting in the fly visual system.

View Article: PubMed Central - PubMed

Affiliation: MRC Laboratory for Molecular Cell Biology, University College London, London, United Kingdom.

ABSTRACT
Parallel processing of neuronal inputs relies on assembling neural circuits into distinct synaptic-columns and layers. This is orchestrated by matching recognition molecules between afferent growth cones and target areas. Controlling the expression of these molecules during development is crucial but not well understood. The developing Drosophila visual system is a powerful genetic model for addressing this question. In this model system, the achromatic R1-6 photoreceptors project their axons in the lamina while the R7 and R8 photoreceptors, which are involved in colour detection, project their axons to two distinct synaptic-layers in the medulla. Here we show that the conserved homeodomain transcription factor Orthodenticle (Otd), which in the eye is a main regulator of rhodopsin expression, is also required for R1-6 photoreceptor synaptic-column specific innervation of the lamina. Our data indicate that otd function in these photoreceptors is largely mediated by the recognition molecules flamingo (fmi) and golden goal (gogo). In addition, we find that otd regulates synaptic-layer targeting of R8. We demonstrate that during this process, otd and the R8-specific transcription factor senseless/Gfi1 (sens) function as independent transcriptional inputs that are required for the expression of fmi, gogo and the adhesion molecule capricious (caps), which govern R8 synaptic-layer targeting. Our work therefore demonstrates that otd is a main component of the gene regulatory network that regulates synaptic-column and layer targeting in the fly visual system.

No MeSH data available.


Related in: MedlinePlus

otd and sens are required for R8 synaptic layer targeting.All panels (A-F) show photoreceptor cell projections stained with 24B10 (red). (A) Expression of UAS-sensRNAi (GFP-negative ommatidia, encircled by a dotted line) in wild-type tissue (GFP positive) 48 h after clone induction using the tub>GFP>Gal4 system. UAS-sensRNAi expressing cells show a clear reduction in Sens protein levels (blue). (B) Expression of UAS-sensRNAi transgene (GFP-negative ommatidia, encircled by a dotted line) in wild-type tissue (GFP positive) 48 h after clone induction using the tub>GFP>Gal4 system. Otd expression (red) is unaffected in UAS-sensRNAi expressing R8 cells (indicated by asterisks). (C) Rh6-lacZ-positive R8 axons in sensRNAi (R8-specific driver-Gal4109–68;UAS-sensRNAi) and in otduvi mutant combined with sensRNAi retina (D), stained with anti-β-galactosidase (green). sens knockdown in the retina leads to a few defects in R8 axon projection, however the combination of the sens knockdown and otduvi mutant leads to a complete failure of R8 layer-specific targeting, indicating that sens and otd act in parallel. (E,F) Layer-specific targeting of the R7 photoreceptors is assessed using the R7 specific transgene Rh4-lacZ. As in wild-type (E), Rh4-lacZ positive R7 terminals (green) correctly target to the M6 layer in otduvi mutants flies (F). (G) Quantification of the misprojections of Rh6-lacZ-positive R8 axons in wild-type, otduvi mutant and otduvi mutant flies where either caps, fmi or gogo expression has been restored. Since Caps is present only in R8, UAS-caps is expressed under the control of an R8-specific Gal4 driver, while UAS-gogo and UAS-fmi are expressed using the pan-photoreceptor GMR-Gal4 driver. Restoring caps expression in otd-mutant R8 photoreceptors partially rescues the otduvi mutant R8 mis-targeting phenotype (from 59% R8 misprojection in otduvi to 25% in otduvi+ UAS-caps). Similarly, expression of fmi and gogo in otduvimutant photoreceptors partially suppress the R8 misprojection phenotype (36% R8 axons misprojecting in otduvi+ UAS-fmi, n = 374 of 1039; 40% R8 axons misprojecting in otduvi+ UAS-gogo, n = 436 of 1090). Percentages indicate quantitative assessment of R8 axons as detected in M3 and M6 layers only. In wild-type, all R8 neurons target to the M3 layer, thus the mistargeting percentage is zero.
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pgen.1005303.g006: otd and sens are required for R8 synaptic layer targeting.All panels (A-F) show photoreceptor cell projections stained with 24B10 (red). (A) Expression of UAS-sensRNAi (GFP-negative ommatidia, encircled by a dotted line) in wild-type tissue (GFP positive) 48 h after clone induction using the tub>GFP>Gal4 system. UAS-sensRNAi expressing cells show a clear reduction in Sens protein levels (blue). (B) Expression of UAS-sensRNAi transgene (GFP-negative ommatidia, encircled by a dotted line) in wild-type tissue (GFP positive) 48 h after clone induction using the tub>GFP>Gal4 system. Otd expression (red) is unaffected in UAS-sensRNAi expressing R8 cells (indicated by asterisks). (C) Rh6-lacZ-positive R8 axons in sensRNAi (R8-specific driver-Gal4109–68;UAS-sensRNAi) and in otduvi mutant combined with sensRNAi retina (D), stained with anti-β-galactosidase (green). sens knockdown in the retina leads to a few defects in R8 axon projection, however the combination of the sens knockdown and otduvi mutant leads to a complete failure of R8 layer-specific targeting, indicating that sens and otd act in parallel. (E,F) Layer-specific targeting of the R7 photoreceptors is assessed using the R7 specific transgene Rh4-lacZ. As in wild-type (E), Rh4-lacZ positive R7 terminals (green) correctly target to the M6 layer in otduvi mutants flies (F). (G) Quantification of the misprojections of Rh6-lacZ-positive R8 axons in wild-type, otduvi mutant and otduvi mutant flies where either caps, fmi or gogo expression has been restored. Since Caps is present only in R8, UAS-caps is expressed under the control of an R8-specific Gal4 driver, while UAS-gogo and UAS-fmi are expressed using the pan-photoreceptor GMR-Gal4 driver. Restoring caps expression in otd-mutant R8 photoreceptors partially rescues the otduvi mutant R8 mis-targeting phenotype (from 59% R8 misprojection in otduvi to 25% in otduvi+ UAS-caps). Similarly, expression of fmi and gogo in otduvimutant photoreceptors partially suppress the R8 misprojection phenotype (36% R8 axons misprojecting in otduvi+ UAS-fmi, n = 374 of 1039; 40% R8 axons misprojecting in otduvi+ UAS-gogo, n = 436 of 1090). Percentages indicate quantitative assessment of R8 axons as detected in M3 and M6 layers only. In wild-type, all R8 neurons target to the M3 layer, thus the mistargeting percentage is zero.

Mentions: The otd mutant defects in R8 layer targeting described here are also observed in R8s that lack the gene encoding the transcription factor sens [20]. Thus, our data raise the possibility that sens might be downregulated in the absence of otd. When we tested this hypothesis, we found this not to be the case, as sens is expressed normally in the absence of otd (Fig 2B). Likewise, otd expression is unaffected in photoreceptors in which sens expression has been reduced (Fig 6B). Therefore, otd must function in parallel to sens during layer targeting of R8 photoreceptors.


Orthodenticle Is Required for the Expression of Principal Recognition Molecules That Control Axon Targeting in the Drosophila Retina.

Mencarelli C, Pichaud F - PLoS Genet. (2015)

otd and sens are required for R8 synaptic layer targeting.All panels (A-F) show photoreceptor cell projections stained with 24B10 (red). (A) Expression of UAS-sensRNAi (GFP-negative ommatidia, encircled by a dotted line) in wild-type tissue (GFP positive) 48 h after clone induction using the tub>GFP>Gal4 system. UAS-sensRNAi expressing cells show a clear reduction in Sens protein levels (blue). (B) Expression of UAS-sensRNAi transgene (GFP-negative ommatidia, encircled by a dotted line) in wild-type tissue (GFP positive) 48 h after clone induction using the tub>GFP>Gal4 system. Otd expression (red) is unaffected in UAS-sensRNAi expressing R8 cells (indicated by asterisks). (C) Rh6-lacZ-positive R8 axons in sensRNAi (R8-specific driver-Gal4109–68;UAS-sensRNAi) and in otduvi mutant combined with sensRNAi retina (D), stained with anti-β-galactosidase (green). sens knockdown in the retina leads to a few defects in R8 axon projection, however the combination of the sens knockdown and otduvi mutant leads to a complete failure of R8 layer-specific targeting, indicating that sens and otd act in parallel. (E,F) Layer-specific targeting of the R7 photoreceptors is assessed using the R7 specific transgene Rh4-lacZ. As in wild-type (E), Rh4-lacZ positive R7 terminals (green) correctly target to the M6 layer in otduvi mutants flies (F). (G) Quantification of the misprojections of Rh6-lacZ-positive R8 axons in wild-type, otduvi mutant and otduvi mutant flies where either caps, fmi or gogo expression has been restored. Since Caps is present only in R8, UAS-caps is expressed under the control of an R8-specific Gal4 driver, while UAS-gogo and UAS-fmi are expressed using the pan-photoreceptor GMR-Gal4 driver. Restoring caps expression in otd-mutant R8 photoreceptors partially rescues the otduvi mutant R8 mis-targeting phenotype (from 59% R8 misprojection in otduvi to 25% in otduvi+ UAS-caps). Similarly, expression of fmi and gogo in otduvimutant photoreceptors partially suppress the R8 misprojection phenotype (36% R8 axons misprojecting in otduvi+ UAS-fmi, n = 374 of 1039; 40% R8 axons misprojecting in otduvi+ UAS-gogo, n = 436 of 1090). Percentages indicate quantitative assessment of R8 axons as detected in M3 and M6 layers only. In wild-type, all R8 neurons target to the M3 layer, thus the mistargeting percentage is zero.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4482733&req=5

pgen.1005303.g006: otd and sens are required for R8 synaptic layer targeting.All panels (A-F) show photoreceptor cell projections stained with 24B10 (red). (A) Expression of UAS-sensRNAi (GFP-negative ommatidia, encircled by a dotted line) in wild-type tissue (GFP positive) 48 h after clone induction using the tub>GFP>Gal4 system. UAS-sensRNAi expressing cells show a clear reduction in Sens protein levels (blue). (B) Expression of UAS-sensRNAi transgene (GFP-negative ommatidia, encircled by a dotted line) in wild-type tissue (GFP positive) 48 h after clone induction using the tub>GFP>Gal4 system. Otd expression (red) is unaffected in UAS-sensRNAi expressing R8 cells (indicated by asterisks). (C) Rh6-lacZ-positive R8 axons in sensRNAi (R8-specific driver-Gal4109–68;UAS-sensRNAi) and in otduvi mutant combined with sensRNAi retina (D), stained with anti-β-galactosidase (green). sens knockdown in the retina leads to a few defects in R8 axon projection, however the combination of the sens knockdown and otduvi mutant leads to a complete failure of R8 layer-specific targeting, indicating that sens and otd act in parallel. (E,F) Layer-specific targeting of the R7 photoreceptors is assessed using the R7 specific transgene Rh4-lacZ. As in wild-type (E), Rh4-lacZ positive R7 terminals (green) correctly target to the M6 layer in otduvi mutants flies (F). (G) Quantification of the misprojections of Rh6-lacZ-positive R8 axons in wild-type, otduvi mutant and otduvi mutant flies where either caps, fmi or gogo expression has been restored. Since Caps is present only in R8, UAS-caps is expressed under the control of an R8-specific Gal4 driver, while UAS-gogo and UAS-fmi are expressed using the pan-photoreceptor GMR-Gal4 driver. Restoring caps expression in otd-mutant R8 photoreceptors partially rescues the otduvi mutant R8 mis-targeting phenotype (from 59% R8 misprojection in otduvi to 25% in otduvi+ UAS-caps). Similarly, expression of fmi and gogo in otduvimutant photoreceptors partially suppress the R8 misprojection phenotype (36% R8 axons misprojecting in otduvi+ UAS-fmi, n = 374 of 1039; 40% R8 axons misprojecting in otduvi+ UAS-gogo, n = 436 of 1090). Percentages indicate quantitative assessment of R8 axons as detected in M3 and M6 layers only. In wild-type, all R8 neurons target to the M3 layer, thus the mistargeting percentage is zero.
Mentions: The otd mutant defects in R8 layer targeting described here are also observed in R8s that lack the gene encoding the transcription factor sens [20]. Thus, our data raise the possibility that sens might be downregulated in the absence of otd. When we tested this hypothesis, we found this not to be the case, as sens is expressed normally in the absence of otd (Fig 2B). Likewise, otd expression is unaffected in photoreceptors in which sens expression has been reduced (Fig 6B). Therefore, otd must function in parallel to sens during layer targeting of R8 photoreceptors.

Bottom Line: Our data indicate that otd function in these photoreceptors is largely mediated by the recognition molecules flamingo (fmi) and golden goal (gogo).In addition, we find that otd regulates synaptic-layer targeting of R8.Our work therefore demonstrates that otd is a main component of the gene regulatory network that regulates synaptic-column and layer targeting in the fly visual system.

View Article: PubMed Central - PubMed

Affiliation: MRC Laboratory for Molecular Cell Biology, University College London, London, United Kingdom.

ABSTRACT
Parallel processing of neuronal inputs relies on assembling neural circuits into distinct synaptic-columns and layers. This is orchestrated by matching recognition molecules between afferent growth cones and target areas. Controlling the expression of these molecules during development is crucial but not well understood. The developing Drosophila visual system is a powerful genetic model for addressing this question. In this model system, the achromatic R1-6 photoreceptors project their axons in the lamina while the R7 and R8 photoreceptors, which are involved in colour detection, project their axons to two distinct synaptic-layers in the medulla. Here we show that the conserved homeodomain transcription factor Orthodenticle (Otd), which in the eye is a main regulator of rhodopsin expression, is also required for R1-6 photoreceptor synaptic-column specific innervation of the lamina. Our data indicate that otd function in these photoreceptors is largely mediated by the recognition molecules flamingo (fmi) and golden goal (gogo). In addition, we find that otd regulates synaptic-layer targeting of R8. We demonstrate that during this process, otd and the R8-specific transcription factor senseless/Gfi1 (sens) function as independent transcriptional inputs that are required for the expression of fmi, gogo and the adhesion molecule capricious (caps), which govern R8 synaptic-layer targeting. Our work therefore demonstrates that otd is a main component of the gene regulatory network that regulates synaptic-column and layer targeting in the fly visual system.

No MeSH data available.


Related in: MedlinePlus