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Cysteine proteinase-1 and cut protein isoform control dendritic innervation of two distinct sensory fields by a single neuron.

Lyons GR, Andersen RO, Abdi K, Song WS, Kuo CT - Cell Rep (2014)

Bottom Line: We found that a class of Drosophila sensory neurons, through complete pruning and regeneration, can elaborate two distinct dendritic trees, innervating independent sensory fields.Unlike known ecdysone effectors, Cp1-mutant ddaC neurons pruned larval dendrites normally but failed to regrow adult dendrites.These results identify a molecular pathway needed for dendrite regrowth after pruning, which allows the same neuron to innervate distinct sensory fields.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology, Duke University School of Medicine, Durham, NC 27710, USA; Medical Scientist Training Program, Duke University School of Medicine, Durham, NC 27710, USA.

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Cp1 Function during Dendrite Regrowth(A) Live imaging of Cp1-EGFP fluorescence in ddaC neurons during metamorphosis.(B) Live imaging of Cp1-EGFP fluorescence in ddaC neurons expressing EcR-DN receptor (ppk-Gal4; UAS-mCD8::RFP; UAS-EcR-DN). Arrows point to RFP+ ddaC neurons. Note that Cp1-EGFP expression in neighboring cells (*) is unaffected by EcR-DN expression via ppk-Gal4 driver.(C) Quantitative analyses of Cp1-EGFP fluorescence levels: average ratio of EGFP/RFP signal from ddaC neurons in WP is set to 1 (n = 6 in all groups). *p < 0.005, Wilcoxon two-sample test. Error bars represent SEM.(D) Live imaging of control and Cp1-mutant ddaC neuron clones at 95 hr APF. Three representative Cp1-mutants are shown.(E) Corresponding colorimetric representation of dendritic arbor depths in (D).(F) Quantitative analyses of Cp1-mutant dendrite regrowth defects: field area, branchpoints, depth of primary (1°) and higher-order dendrites from surface, and percentages of total dendrite length at 95 hr APF that are shallow (within 15 μm from the body wall) or deep (≥15 μm). cont., control; resc., rescue. n ≥ 6 in all groups. *p < 0.001, one-way ANOVA. Error bars represent SEM.Scale bars, 2 mm (A), 5 mm (B), and 50 μm (D). See also Figure S3.
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Figure 2: Cp1 Function during Dendrite Regrowth(A) Live imaging of Cp1-EGFP fluorescence in ddaC neurons during metamorphosis.(B) Live imaging of Cp1-EGFP fluorescence in ddaC neurons expressing EcR-DN receptor (ppk-Gal4; UAS-mCD8::RFP; UAS-EcR-DN). Arrows point to RFP+ ddaC neurons. Note that Cp1-EGFP expression in neighboring cells (*) is unaffected by EcR-DN expression via ppk-Gal4 driver.(C) Quantitative analyses of Cp1-EGFP fluorescence levels: average ratio of EGFP/RFP signal from ddaC neurons in WP is set to 1 (n = 6 in all groups). *p < 0.005, Wilcoxon two-sample test. Error bars represent SEM.(D) Live imaging of control and Cp1-mutant ddaC neuron clones at 95 hr APF. Three representative Cp1-mutants are shown.(E) Corresponding colorimetric representation of dendritic arbor depths in (D).(F) Quantitative analyses of Cp1-mutant dendrite regrowth defects: field area, branchpoints, depth of primary (1°) and higher-order dendrites from surface, and percentages of total dendrite length at 95 hr APF that are shallow (within 15 μm from the body wall) or deep (≥15 μm). cont., control; resc., rescue. n ≥ 6 in all groups. *p < 0.001, one-way ANOVA. Error bars represent SEM.Scale bars, 2 mm (A), 5 mm (B), and 50 μm (D). See also Figure S3.

Mentions: We hypothesized that if variations in molecular programs are needed to grow two different sets of dendrites in the same neuron, then the genes involved will likely change their expression levels in a context-dependent manner. We set out to identify such genes in ddaC neurons during dendrite regrowth. An expression screen of the EGFP-FlyTrap collection identified stock ZCL2854, corresponding to EGFP insertion into the Cp1 gene, that showed increased EGFP expression during ddaC neuron dendrite remodeling (Figure 2A). We quantified Cp1-EGFP fluorescence levels in ddaC neurons by normalizing EGFP intensity to internal UAS-mCD8::RFP fluorescence driven by ppk-Gal4, which remained relatively constant throughout (Figure 2C; data not shown). We showed previously that dendrite remodeling in these neurons is initiated by nuclear hormone receptor signaling (Kuo et al., 2005). To confirm that the increase in Cp1-EGFP expression during metamorphosis is controlled by the Drosophila hormone ecdysone, we blocked ecdysone signaling by expressing a dominant-negative ecdysone receptor (UAS-EcR-DN) in ddaC neurons (Kirilly et al., 2009; Kuo et al., 2005). This effectively attenuated Cp1-EGFP upregulation during metamorphosis (Figures 2B and 2C).


Cysteine proteinase-1 and cut protein isoform control dendritic innervation of two distinct sensory fields by a single neuron.

Lyons GR, Andersen RO, Abdi K, Song WS, Kuo CT - Cell Rep (2014)

Cp1 Function during Dendrite Regrowth(A) Live imaging of Cp1-EGFP fluorescence in ddaC neurons during metamorphosis.(B) Live imaging of Cp1-EGFP fluorescence in ddaC neurons expressing EcR-DN receptor (ppk-Gal4; UAS-mCD8::RFP; UAS-EcR-DN). Arrows point to RFP+ ddaC neurons. Note that Cp1-EGFP expression in neighboring cells (*) is unaffected by EcR-DN expression via ppk-Gal4 driver.(C) Quantitative analyses of Cp1-EGFP fluorescence levels: average ratio of EGFP/RFP signal from ddaC neurons in WP is set to 1 (n = 6 in all groups). *p < 0.005, Wilcoxon two-sample test. Error bars represent SEM.(D) Live imaging of control and Cp1-mutant ddaC neuron clones at 95 hr APF. Three representative Cp1-mutants are shown.(E) Corresponding colorimetric representation of dendritic arbor depths in (D).(F) Quantitative analyses of Cp1-mutant dendrite regrowth defects: field area, branchpoints, depth of primary (1°) and higher-order dendrites from surface, and percentages of total dendrite length at 95 hr APF that are shallow (within 15 μm from the body wall) or deep (≥15 μm). cont., control; resc., rescue. n ≥ 6 in all groups. *p < 0.001, one-way ANOVA. Error bars represent SEM.Scale bars, 2 mm (A), 5 mm (B), and 50 μm (D). See also Figure S3.
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Figure 2: Cp1 Function during Dendrite Regrowth(A) Live imaging of Cp1-EGFP fluorescence in ddaC neurons during metamorphosis.(B) Live imaging of Cp1-EGFP fluorescence in ddaC neurons expressing EcR-DN receptor (ppk-Gal4; UAS-mCD8::RFP; UAS-EcR-DN). Arrows point to RFP+ ddaC neurons. Note that Cp1-EGFP expression in neighboring cells (*) is unaffected by EcR-DN expression via ppk-Gal4 driver.(C) Quantitative analyses of Cp1-EGFP fluorescence levels: average ratio of EGFP/RFP signal from ddaC neurons in WP is set to 1 (n = 6 in all groups). *p < 0.005, Wilcoxon two-sample test. Error bars represent SEM.(D) Live imaging of control and Cp1-mutant ddaC neuron clones at 95 hr APF. Three representative Cp1-mutants are shown.(E) Corresponding colorimetric representation of dendritic arbor depths in (D).(F) Quantitative analyses of Cp1-mutant dendrite regrowth defects: field area, branchpoints, depth of primary (1°) and higher-order dendrites from surface, and percentages of total dendrite length at 95 hr APF that are shallow (within 15 μm from the body wall) or deep (≥15 μm). cont., control; resc., rescue. n ≥ 6 in all groups. *p < 0.001, one-way ANOVA. Error bars represent SEM.Scale bars, 2 mm (A), 5 mm (B), and 50 μm (D). See also Figure S3.
Mentions: We hypothesized that if variations in molecular programs are needed to grow two different sets of dendrites in the same neuron, then the genes involved will likely change their expression levels in a context-dependent manner. We set out to identify such genes in ddaC neurons during dendrite regrowth. An expression screen of the EGFP-FlyTrap collection identified stock ZCL2854, corresponding to EGFP insertion into the Cp1 gene, that showed increased EGFP expression during ddaC neuron dendrite remodeling (Figure 2A). We quantified Cp1-EGFP fluorescence levels in ddaC neurons by normalizing EGFP intensity to internal UAS-mCD8::RFP fluorescence driven by ppk-Gal4, which remained relatively constant throughout (Figure 2C; data not shown). We showed previously that dendrite remodeling in these neurons is initiated by nuclear hormone receptor signaling (Kuo et al., 2005). To confirm that the increase in Cp1-EGFP expression during metamorphosis is controlled by the Drosophila hormone ecdysone, we blocked ecdysone signaling by expressing a dominant-negative ecdysone receptor (UAS-EcR-DN) in ddaC neurons (Kirilly et al., 2009; Kuo et al., 2005). This effectively attenuated Cp1-EGFP upregulation during metamorphosis (Figures 2B and 2C).

Bottom Line: We found that a class of Drosophila sensory neurons, through complete pruning and regeneration, can elaborate two distinct dendritic trees, innervating independent sensory fields.Unlike known ecdysone effectors, Cp1-mutant ddaC neurons pruned larval dendrites normally but failed to regrow adult dendrites.These results identify a molecular pathway needed for dendrite regrowth after pruning, which allows the same neuron to innervate distinct sensory fields.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology, Duke University School of Medicine, Durham, NC 27710, USA; Medical Scientist Training Program, Duke University School of Medicine, Durham, NC 27710, USA.

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Related in: MedlinePlus