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The BTB/POZ zinc finger protein Broad-Z3 promotes dendritic outgrowth during metamorphic remodeling of the peripheral stretch receptor dbd.

Scott JA, Williams DW, Truman JW - Neural Dev (2011)

Bottom Line: We used the metamorphic remodeling of an identified sensory neuron, the dorsal bipolar dendrite sensory neuron (dbd), to examine the effects of BrZ3 expression on the extent and pattern of dendrite growth during metamorphosis.The adult arbor of dbd is a highly branched arbor whose branches self-fasciculate to form a compact dendritic bundle.These data suggest that the nature of their local environment can change dendrite behavior from self-adhesion to self-avoidance.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Biology, Box 351800, University of Washington, Seattle, WA 98195, USA.

ABSTRACT

Background: Various members of the family of BTB/POZ zinc-finger transcription factors influence patterns of dendritic branching. One such member, Broad, is notable because its BrZ3 isoform is widely expressed in Drosophila in immature neurons around the time of arbor outgrowth. We used the metamorphic remodeling of an identified sensory neuron, the dorsal bipolar dendrite sensory neuron (dbd), to examine the effects of BrZ3 expression on the extent and pattern of dendrite growth during metamorphosis.

Results: Using live imaging of dbd in Drosophila pupae, we followed its normal development during metamorphosis and the effect of ectopic expression of BrZ3 on this development. After migration of its cell body, dbd extends a growth-cone that grows between two muscle bands followed by branching and turning back on itself to form a compact dendritic bundle. The ectopic expression of the BrZ3 isoform, using the GAL4/UAS system, caused dbd's dendritic tree to transform from its normal, compact, fasciculated form into a comb-like arbor that spread over on the body wall. Time-lapse analysis revealed that the expression of BrZ3 caused the premature extension of the primary dendrite onto immature myoblasts, ectopic growth past the muscle target region, and subsequent elaboration onto the epidermis. To control the timing of expression of BrZ3, we used a temperature-sensitive GAL80 mutant. When BrZ3 expression was delayed until after the extension of the primary dendrite, then a normal arbor was formed. By contrast, when BrZ3 expression was confined to only the early outgrowth phase, then ectopic arbors were subsequently formed and maintained on the epidermis despite the subsequent absence of BrZ3.

Conclusions: The adult arbor of dbd is a highly branched arbor whose branches self-fasciculate to form a compact dendritic bundle. The ectopic expression of BrZ3 in this cell causes a premature extension of its growth-cone, resulting in dendrites that extend beyond their normal muscle substrate and onto the epidermis, where they form a comb-shaped, ectopic arbor. Our quantitative data suggest that new ectopic arbor represents an 'unpacking' of the normally fasciculated arbor onto the epidermis. These data suggest that the nature of their local environment can change dendrite behavior from self-adhesion to self-avoidance.

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Ectopic BrZ3 expression generated a stereotypic ectopic arbor in the adult dbd neuron during metamorphosis. (A, B) Morphology of the dendritic arbor of the dbd neuron in the last larval stage of controls (A: C161-GAL4 > UAS mCD8::GFP) and individuals expressing BrZ3 (B: C161-GAL4 > UAS mCD8:GFP, UAS-BrZ3 dbd [dbd[+BrZ3]]). (C-E) The effects of expressing different Broad isoforms in dbd on its dendritic morphology at the end of metamorphosis. (C) Pharate adult morphology of dbd[+BrZ3]; arrowhead, anti-Broad-Core immunostaining. (D) Morphology of dbd[+BrZ1] is similar to control. (E) Morphology of dbd[+BrZ4] was variable but typically maintained contact with muscles. Bracket: dbd arbor in adjacent muscle grooves. (F) Control pharate adult dbd. (F') Y-projection. (G, H) Examples of dbd[+BrZ3] ectopic combs (brackets) posterior to dorsal abdominal muscles. Loss of anterior dendrites (asterisk) occurred in 25% of dbd[+BrZ3] neurons. (G', H') Y-projections showing the ectopic comb (arrows) was superficial to the muscle layers, veering toward the cuticle. (I, I') MAP1B-like antibody labeled microtubules in the ectopic arbor 'backbone' and some branches (arrowhead): green, anti-CD8; magenta, Futsch; 50 h APF. (J) Control neurons (black) had thicker dendritic bundles than dbd[+BrZ3] neurons (green). The trend line shows a significant negative correlation between dorsal-ventral ectopic arbor height and main bundle thickness among dbd[+BrZ3] (r = -0.49, P < 0.01). Scale bar = 50 μm. Immunostaining except for (I): green, anti-CD8; magenta, phalloidin.
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Figure 4: Ectopic BrZ3 expression generated a stereotypic ectopic arbor in the adult dbd neuron during metamorphosis. (A, B) Morphology of the dendritic arbor of the dbd neuron in the last larval stage of controls (A: C161-GAL4 > UAS mCD8::GFP) and individuals expressing BrZ3 (B: C161-GAL4 > UAS mCD8:GFP, UAS-BrZ3 dbd [dbd[+BrZ3]]). (C-E) The effects of expressing different Broad isoforms in dbd on its dendritic morphology at the end of metamorphosis. (C) Pharate adult morphology of dbd[+BrZ3]; arrowhead, anti-Broad-Core immunostaining. (D) Morphology of dbd[+BrZ1] is similar to control. (E) Morphology of dbd[+BrZ4] was variable but typically maintained contact with muscles. Bracket: dbd arbor in adjacent muscle grooves. (F) Control pharate adult dbd. (F') Y-projection. (G, H) Examples of dbd[+BrZ3] ectopic combs (brackets) posterior to dorsal abdominal muscles. Loss of anterior dendrites (asterisk) occurred in 25% of dbd[+BrZ3] neurons. (G', H') Y-projections showing the ectopic comb (arrows) was superficial to the muscle layers, veering toward the cuticle. (I, I') MAP1B-like antibody labeled microtubules in the ectopic arbor 'backbone' and some branches (arrowhead): green, anti-CD8; magenta, Futsch; 50 h APF. (J) Control neurons (black) had thicker dendritic bundles than dbd[+BrZ3] neurons (green). The trend line shows a significant negative correlation between dorsal-ventral ectopic arbor height and main bundle thickness among dbd[+BrZ3] (r = -0.49, P < 0.01). Scale bar = 50 μm. Immunostaining except for (I): green, anti-CD8; magenta, phalloidin.

Mentions: We examined the effects of expressing Broad isoforms on the growth of dbd, and found the Z3 isoform of Broad (BrZ3) transformed the shape of the adult dbd dendrites from its normal bipolar form into a tree-like arborization. In the larva, BrZ3 misexpression did not interfere with the dendritic morphology of dbd or its association with target muscles (Figure 4A, B; n = 15 neurons in 4 animals). However, during metamorphosis, GAL4C161/UAS-induced expression of BrZ3 resulted in a stereotyped, comb-like arborization in an ectopic position (Figure 4C; n = 45 of 48 neurons). The ectopic arborization was unique to dbd neurons expressing the BrZ3 isoform (dbd[+BrZ3]). The dbd[+BrZ1] neurons were normal (Figure 4D; n = 18 neurons in 7 animals). The dbd[+BrZ4] neurons were highly variable from segment to segment (Figure 4E; n = 31 neurons in 9 animals) and showed multiple short branches that extended from the muscle groove, but they did not consistently produce a stereotyped ectopic arbor. Staining for muscles showed that control arbors did not leave the muscle (Figure 4F), but the dbd[+BrZ3] ectopic arbor extended beyond the posterior ends of the muscles and onto the epidermis (Figure 4G-H'; n = 39 of 42 neurons). Immunostaining against the microtubule-associated protein Futsch showed that that the major axis of the dbd[+BrZ3] ectopic arbor, the comb 'backbone', and some higher-order branches were stabilized with microtubules by 50 h APF (Figure 4I, arrowhead; n = 6). While the general shape and orientation of this comb-like arbor were consistent, its footprint ranged in dorsal-ventral axis (height) from 18 to 181 μm (mean = 97 μm, standard deviation = 49), and in the anteroposterior axis (length) from 1 to 90 μm (mean = 42 μm, standard deviation = 19). Several measures suggest that dbd[+BrZ3] dendrites spread onto the epidermis at the expense of the fasciculated main arbor. Overall, the main dendritic bundle of dbd[+BrZ3] was thinner than in control cells (Figure 4J; P = 0.0002, t-test assuming unequal variances). Within the range of dbd[+BrZ3] morphologies, larger ectopic arbor size was correlated with decreased thickness of the fasciculated arbor (Figure 4J; r = -0.49, P < 0.01). Also, failure of the normal arborization to reach the anterior margin of the muscle was correlated with a larger ectopic arbor size (Figure 4H, asterisk; r = 0.29, P = 0.023). Taken together, these measurements suggest that the dendrites of dbd[+BrZ3] are primarily misplaced rather than overgrowing.


The BTB/POZ zinc finger protein Broad-Z3 promotes dendritic outgrowth during metamorphic remodeling of the peripheral stretch receptor dbd.

Scott JA, Williams DW, Truman JW - Neural Dev (2011)

Ectopic BrZ3 expression generated a stereotypic ectopic arbor in the adult dbd neuron during metamorphosis. (A, B) Morphology of the dendritic arbor of the dbd neuron in the last larval stage of controls (A: C161-GAL4 > UAS mCD8::GFP) and individuals expressing BrZ3 (B: C161-GAL4 > UAS mCD8:GFP, UAS-BrZ3 dbd [dbd[+BrZ3]]). (C-E) The effects of expressing different Broad isoforms in dbd on its dendritic morphology at the end of metamorphosis. (C) Pharate adult morphology of dbd[+BrZ3]; arrowhead, anti-Broad-Core immunostaining. (D) Morphology of dbd[+BrZ1] is similar to control. (E) Morphology of dbd[+BrZ4] was variable but typically maintained contact with muscles. Bracket: dbd arbor in adjacent muscle grooves. (F) Control pharate adult dbd. (F') Y-projection. (G, H) Examples of dbd[+BrZ3] ectopic combs (brackets) posterior to dorsal abdominal muscles. Loss of anterior dendrites (asterisk) occurred in 25% of dbd[+BrZ3] neurons. (G', H') Y-projections showing the ectopic comb (arrows) was superficial to the muscle layers, veering toward the cuticle. (I, I') MAP1B-like antibody labeled microtubules in the ectopic arbor 'backbone' and some branches (arrowhead): green, anti-CD8; magenta, Futsch; 50 h APF. (J) Control neurons (black) had thicker dendritic bundles than dbd[+BrZ3] neurons (green). The trend line shows a significant negative correlation between dorsal-ventral ectopic arbor height and main bundle thickness among dbd[+BrZ3] (r = -0.49, P < 0.01). Scale bar = 50 μm. Immunostaining except for (I): green, anti-CD8; magenta, phalloidin.
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Figure 4: Ectopic BrZ3 expression generated a stereotypic ectopic arbor in the adult dbd neuron during metamorphosis. (A, B) Morphology of the dendritic arbor of the dbd neuron in the last larval stage of controls (A: C161-GAL4 > UAS mCD8::GFP) and individuals expressing BrZ3 (B: C161-GAL4 > UAS mCD8:GFP, UAS-BrZ3 dbd [dbd[+BrZ3]]). (C-E) The effects of expressing different Broad isoforms in dbd on its dendritic morphology at the end of metamorphosis. (C) Pharate adult morphology of dbd[+BrZ3]; arrowhead, anti-Broad-Core immunostaining. (D) Morphology of dbd[+BrZ1] is similar to control. (E) Morphology of dbd[+BrZ4] was variable but typically maintained contact with muscles. Bracket: dbd arbor in adjacent muscle grooves. (F) Control pharate adult dbd. (F') Y-projection. (G, H) Examples of dbd[+BrZ3] ectopic combs (brackets) posterior to dorsal abdominal muscles. Loss of anterior dendrites (asterisk) occurred in 25% of dbd[+BrZ3] neurons. (G', H') Y-projections showing the ectopic comb (arrows) was superficial to the muscle layers, veering toward the cuticle. (I, I') MAP1B-like antibody labeled microtubules in the ectopic arbor 'backbone' and some branches (arrowhead): green, anti-CD8; magenta, Futsch; 50 h APF. (J) Control neurons (black) had thicker dendritic bundles than dbd[+BrZ3] neurons (green). The trend line shows a significant negative correlation between dorsal-ventral ectopic arbor height and main bundle thickness among dbd[+BrZ3] (r = -0.49, P < 0.01). Scale bar = 50 μm. Immunostaining except for (I): green, anti-CD8; magenta, phalloidin.
Mentions: We examined the effects of expressing Broad isoforms on the growth of dbd, and found the Z3 isoform of Broad (BrZ3) transformed the shape of the adult dbd dendrites from its normal bipolar form into a tree-like arborization. In the larva, BrZ3 misexpression did not interfere with the dendritic morphology of dbd or its association with target muscles (Figure 4A, B; n = 15 neurons in 4 animals). However, during metamorphosis, GAL4C161/UAS-induced expression of BrZ3 resulted in a stereotyped, comb-like arborization in an ectopic position (Figure 4C; n = 45 of 48 neurons). The ectopic arborization was unique to dbd neurons expressing the BrZ3 isoform (dbd[+BrZ3]). The dbd[+BrZ1] neurons were normal (Figure 4D; n = 18 neurons in 7 animals). The dbd[+BrZ4] neurons were highly variable from segment to segment (Figure 4E; n = 31 neurons in 9 animals) and showed multiple short branches that extended from the muscle groove, but they did not consistently produce a stereotyped ectopic arbor. Staining for muscles showed that control arbors did not leave the muscle (Figure 4F), but the dbd[+BrZ3] ectopic arbor extended beyond the posterior ends of the muscles and onto the epidermis (Figure 4G-H'; n = 39 of 42 neurons). Immunostaining against the microtubule-associated protein Futsch showed that that the major axis of the dbd[+BrZ3] ectopic arbor, the comb 'backbone', and some higher-order branches were stabilized with microtubules by 50 h APF (Figure 4I, arrowhead; n = 6). While the general shape and orientation of this comb-like arbor were consistent, its footprint ranged in dorsal-ventral axis (height) from 18 to 181 μm (mean = 97 μm, standard deviation = 49), and in the anteroposterior axis (length) from 1 to 90 μm (mean = 42 μm, standard deviation = 19). Several measures suggest that dbd[+BrZ3] dendrites spread onto the epidermis at the expense of the fasciculated main arbor. Overall, the main dendritic bundle of dbd[+BrZ3] was thinner than in control cells (Figure 4J; P = 0.0002, t-test assuming unequal variances). Within the range of dbd[+BrZ3] morphologies, larger ectopic arbor size was correlated with decreased thickness of the fasciculated arbor (Figure 4J; r = -0.49, P < 0.01). Also, failure of the normal arborization to reach the anterior margin of the muscle was correlated with a larger ectopic arbor size (Figure 4H, asterisk; r = 0.29, P = 0.023). Taken together, these measurements suggest that the dendrites of dbd[+BrZ3] are primarily misplaced rather than overgrowing.

Bottom Line: We used the metamorphic remodeling of an identified sensory neuron, the dorsal bipolar dendrite sensory neuron (dbd), to examine the effects of BrZ3 expression on the extent and pattern of dendrite growth during metamorphosis.The adult arbor of dbd is a highly branched arbor whose branches self-fasciculate to form a compact dendritic bundle.These data suggest that the nature of their local environment can change dendrite behavior from self-adhesion to self-avoidance.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Biology, Box 351800, University of Washington, Seattle, WA 98195, USA.

ABSTRACT

Background: Various members of the family of BTB/POZ zinc-finger transcription factors influence patterns of dendritic branching. One such member, Broad, is notable because its BrZ3 isoform is widely expressed in Drosophila in immature neurons around the time of arbor outgrowth. We used the metamorphic remodeling of an identified sensory neuron, the dorsal bipolar dendrite sensory neuron (dbd), to examine the effects of BrZ3 expression on the extent and pattern of dendrite growth during metamorphosis.

Results: Using live imaging of dbd in Drosophila pupae, we followed its normal development during metamorphosis and the effect of ectopic expression of BrZ3 on this development. After migration of its cell body, dbd extends a growth-cone that grows between two muscle bands followed by branching and turning back on itself to form a compact dendritic bundle. The ectopic expression of the BrZ3 isoform, using the GAL4/UAS system, caused dbd's dendritic tree to transform from its normal, compact, fasciculated form into a comb-like arbor that spread over on the body wall. Time-lapse analysis revealed that the expression of BrZ3 caused the premature extension of the primary dendrite onto immature myoblasts, ectopic growth past the muscle target region, and subsequent elaboration onto the epidermis. To control the timing of expression of BrZ3, we used a temperature-sensitive GAL80 mutant. When BrZ3 expression was delayed until after the extension of the primary dendrite, then a normal arbor was formed. By contrast, when BrZ3 expression was confined to only the early outgrowth phase, then ectopic arbors were subsequently formed and maintained on the epidermis despite the subsequent absence of BrZ3.

Conclusions: The adult arbor of dbd is a highly branched arbor whose branches self-fasciculate to form a compact dendritic bundle. The ectopic expression of BrZ3 in this cell causes a premature extension of its growth-cone, resulting in dendrites that extend beyond their normal muscle substrate and onto the epidermis, where they form a comb-shaped, ectopic arbor. Our quantitative data suggest that new ectopic arbor represents an 'unpacking' of the normally fasciculated arbor onto the epidermis. These data suggest that the nature of their local environment can change dendrite behavior from self-adhesion to self-avoidance.

Show MeSH
Related in: MedlinePlus