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Dynein-dynactin complex is essential for dendritic restriction of TM1-containing Drosophila Dscam.

Yang JS, Bai JM, Lee T - PLoS ONE (2008)

Bottom Line: In contrast, compromising dynein/dynactin function did not affect dendritic targeting of two other dendritic markers, Nod and Rdl.Tracing newly synthesized Dscam[TM1] further revealed that compromising dynein/dynactin function did not affect the initial dendritic targeting of Dscam[TM1], but disrupted the maintenance of its restriction to dendrites.The results of this study suggest multiple mechanisms of dendritic protein targeting.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurobiology, University of Massachusetts Medical School, Worcester, MA, USA.

ABSTRACT

Background: Many membrane proteins, including Drosophila Dscam, are enriched in dendrites or axons within neurons. However, little is known about how the differential distribution is established and maintained.

Methodology/principal findings: Here we investigated the mechanisms underlying the dendritic targeting of Dscam[TM1]. Through forward genetic mosaic screens and by silencing specific genes via targeted RNAi, we found that several genes, encoding various components of the dynein-dynactin complex, are required for restricting Dscam[TM1] to the mushroom body dendrites. In contrast, compromising dynein/dynactin function did not affect dendritic targeting of two other dendritic markers, Nod and Rdl. Tracing newly synthesized Dscam[TM1] further revealed that compromising dynein/dynactin function did not affect the initial dendritic targeting of Dscam[TM1], but disrupted the maintenance of its restriction to dendrites.

Conclusions/significance: The results of this study suggest multiple mechanisms of dendritic protein targeting. Notably, dynein-dynactin plays a role in excluding dendritic Dscam, but not Rdl, from axons by retrograde transport.

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Related in: MedlinePlus

Retrograde transport plays a role in restricting Dscam[TM1]::GFP to the somatodendritic domain.(A–D) Transient induction of UAS-Dscam[TM1]::GFP or UAS-mCD8::GFP by TARGET. Prior to induction, GAL80ts fully suppressed the expression at a permissive temperature (A, C). Notably, one hour after heat-shock inactivation of GAL80ts, Dscam[TM1]::GFP was detected only in MB calyx (B) while mCD8::GFP distributed throughout the entire neurons (D). (E–L) Induction of Dscam[TM1]::GFP with or without GluedDN. Following co-induction with dominant-negative Glued, Dscam[TM1]::GFP gradually spread into MB peduncles and axonal lobes (F–H, arrows). In contrast, Dscam[TM1]::GFP was well restricted to the MB calyx in the absence of dominant-negative Glued (I–L). (M) Effects of GluedDN dosage on the misdistribution of Dscam[TM1]::GFP. Dscam[TM1]::GFP could localize in dendrites only (e.g. [E]), dendrites plus peduncles (e.g. [F]), dendrites, peduncles plus proximal portions of axon lobes (e.g. [G]), or from calyx to the tips of axon lobes (e.g. [H]). Note that increasing GlueDN dosage did not accelerate the mislocalization process. Three insertion lines of UAS-GlΔ were examined: UAS-GlΔ84, UAS-GlΔ008m, and UAS-GlΔ020m. All of them were examined individually in 1× Glued-DN. All possible combinations of them were checked in 2× Glued-DN. No statistically significant differences were detected among conditions with distinct insertions or different numbers of insertions.
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pone-0003504-g006: Retrograde transport plays a role in restricting Dscam[TM1]::GFP to the somatodendritic domain.(A–D) Transient induction of UAS-Dscam[TM1]::GFP or UAS-mCD8::GFP by TARGET. Prior to induction, GAL80ts fully suppressed the expression at a permissive temperature (A, C). Notably, one hour after heat-shock inactivation of GAL80ts, Dscam[TM1]::GFP was detected only in MB calyx (B) while mCD8::GFP distributed throughout the entire neurons (D). (E–L) Induction of Dscam[TM1]::GFP with or without GluedDN. Following co-induction with dominant-negative Glued, Dscam[TM1]::GFP gradually spread into MB peduncles and axonal lobes (F–H, arrows). In contrast, Dscam[TM1]::GFP was well restricted to the MB calyx in the absence of dominant-negative Glued (I–L). (M) Effects of GluedDN dosage on the misdistribution of Dscam[TM1]::GFP. Dscam[TM1]::GFP could localize in dendrites only (e.g. [E]), dendrites plus peduncles (e.g. [F]), dendrites, peduncles plus proximal portions of axon lobes (e.g. [G]), or from calyx to the tips of axon lobes (e.g. [H]). Note that increasing GlueDN dosage did not accelerate the mislocalization process. Three insertion lines of UAS-GlΔ were examined: UAS-GlΔ84, UAS-GlΔ008m, and UAS-GlΔ020m. All of them were examined individually in 1× Glued-DN. All possible combinations of them were checked in 2× Glued-DN. No statistically significant differences were detected among conditions with distinct insertions or different numbers of insertions.

Mentions: Transient induction of Dscam[TM1]::GFP in the larval MBs was achieved using the TARGET system, in which GAL4-dependent expression of UAS-transgene is acutely controlled by a temperature-sensitive GAL4 repressor, GAL80ts [50]. At 18°C, GAL4-OK107 was fully suppressed by GAL80ts (Figure 6A and 6C). Following inactivation of GAL80ts by shifting the organisms to higher temperatures (see Experimental Procedures), we could start to detect mCD8::GFP or Dscam[TM1]::GFP in young MB neurons (whose axons occupy core regions of axonal bundles and are weakly labeled by 1D4 mAb [51]) approximately one hour after induction. Since the enrichment of newly synthesized protein in young MB neurons were seen for both mCD8::GFP and Dscam[TM1]::GFP, this phenomenon could possibly result from the expression profile of GAL4-OK107 at the wandering larval stage or the difference in the intrinsic properties of newly derived MB neurons versus mature ones. Notably, while mCD8::GFP was uniformly distributed (Figure 6D), newly synthesized Dscam[TM1]::GFP was consistently located to dendrites (Figure 6B). These observations suggest involvement of selective transport in targeting Dscam[TM1] specifically to the dendrites.


Dynein-dynactin complex is essential for dendritic restriction of TM1-containing Drosophila Dscam.

Yang JS, Bai JM, Lee T - PLoS ONE (2008)

Retrograde transport plays a role in restricting Dscam[TM1]::GFP to the somatodendritic domain.(A–D) Transient induction of UAS-Dscam[TM1]::GFP or UAS-mCD8::GFP by TARGET. Prior to induction, GAL80ts fully suppressed the expression at a permissive temperature (A, C). Notably, one hour after heat-shock inactivation of GAL80ts, Dscam[TM1]::GFP was detected only in MB calyx (B) while mCD8::GFP distributed throughout the entire neurons (D). (E–L) Induction of Dscam[TM1]::GFP with or without GluedDN. Following co-induction with dominant-negative Glued, Dscam[TM1]::GFP gradually spread into MB peduncles and axonal lobes (F–H, arrows). In contrast, Dscam[TM1]::GFP was well restricted to the MB calyx in the absence of dominant-negative Glued (I–L). (M) Effects of GluedDN dosage on the misdistribution of Dscam[TM1]::GFP. Dscam[TM1]::GFP could localize in dendrites only (e.g. [E]), dendrites plus peduncles (e.g. [F]), dendrites, peduncles plus proximal portions of axon lobes (e.g. [G]), or from calyx to the tips of axon lobes (e.g. [H]). Note that increasing GlueDN dosage did not accelerate the mislocalization process. Three insertion lines of UAS-GlΔ were examined: UAS-GlΔ84, UAS-GlΔ008m, and UAS-GlΔ020m. All of them were examined individually in 1× Glued-DN. All possible combinations of them were checked in 2× Glued-DN. No statistically significant differences were detected among conditions with distinct insertions or different numbers of insertions.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2566808&req=5

pone-0003504-g006: Retrograde transport plays a role in restricting Dscam[TM1]::GFP to the somatodendritic domain.(A–D) Transient induction of UAS-Dscam[TM1]::GFP or UAS-mCD8::GFP by TARGET. Prior to induction, GAL80ts fully suppressed the expression at a permissive temperature (A, C). Notably, one hour after heat-shock inactivation of GAL80ts, Dscam[TM1]::GFP was detected only in MB calyx (B) while mCD8::GFP distributed throughout the entire neurons (D). (E–L) Induction of Dscam[TM1]::GFP with or without GluedDN. Following co-induction with dominant-negative Glued, Dscam[TM1]::GFP gradually spread into MB peduncles and axonal lobes (F–H, arrows). In contrast, Dscam[TM1]::GFP was well restricted to the MB calyx in the absence of dominant-negative Glued (I–L). (M) Effects of GluedDN dosage on the misdistribution of Dscam[TM1]::GFP. Dscam[TM1]::GFP could localize in dendrites only (e.g. [E]), dendrites plus peduncles (e.g. [F]), dendrites, peduncles plus proximal portions of axon lobes (e.g. [G]), or from calyx to the tips of axon lobes (e.g. [H]). Note that increasing GlueDN dosage did not accelerate the mislocalization process. Three insertion lines of UAS-GlΔ were examined: UAS-GlΔ84, UAS-GlΔ008m, and UAS-GlΔ020m. All of them were examined individually in 1× Glued-DN. All possible combinations of them were checked in 2× Glued-DN. No statistically significant differences were detected among conditions with distinct insertions or different numbers of insertions.
Mentions: Transient induction of Dscam[TM1]::GFP in the larval MBs was achieved using the TARGET system, in which GAL4-dependent expression of UAS-transgene is acutely controlled by a temperature-sensitive GAL4 repressor, GAL80ts [50]. At 18°C, GAL4-OK107 was fully suppressed by GAL80ts (Figure 6A and 6C). Following inactivation of GAL80ts by shifting the organisms to higher temperatures (see Experimental Procedures), we could start to detect mCD8::GFP or Dscam[TM1]::GFP in young MB neurons (whose axons occupy core regions of axonal bundles and are weakly labeled by 1D4 mAb [51]) approximately one hour after induction. Since the enrichment of newly synthesized protein in young MB neurons were seen for both mCD8::GFP and Dscam[TM1]::GFP, this phenomenon could possibly result from the expression profile of GAL4-OK107 at the wandering larval stage or the difference in the intrinsic properties of newly derived MB neurons versus mature ones. Notably, while mCD8::GFP was uniformly distributed (Figure 6D), newly synthesized Dscam[TM1]::GFP was consistently located to dendrites (Figure 6B). These observations suggest involvement of selective transport in targeting Dscam[TM1] specifically to the dendrites.

Bottom Line: In contrast, compromising dynein/dynactin function did not affect dendritic targeting of two other dendritic markers, Nod and Rdl.Tracing newly synthesized Dscam[TM1] further revealed that compromising dynein/dynactin function did not affect the initial dendritic targeting of Dscam[TM1], but disrupted the maintenance of its restriction to dendrites.The results of this study suggest multiple mechanisms of dendritic protein targeting.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurobiology, University of Massachusetts Medical School, Worcester, MA, USA.

ABSTRACT

Background: Many membrane proteins, including Drosophila Dscam, are enriched in dendrites or axons within neurons. However, little is known about how the differential distribution is established and maintained.

Methodology/principal findings: Here we investigated the mechanisms underlying the dendritic targeting of Dscam[TM1]. Through forward genetic mosaic screens and by silencing specific genes via targeted RNAi, we found that several genes, encoding various components of the dynein-dynactin complex, are required for restricting Dscam[TM1] to the mushroom body dendrites. In contrast, compromising dynein/dynactin function did not affect dendritic targeting of two other dendritic markers, Nod and Rdl. Tracing newly synthesized Dscam[TM1] further revealed that compromising dynein/dynactin function did not affect the initial dendritic targeting of Dscam[TM1], but disrupted the maintenance of its restriction to dendrites.

Conclusions/significance: The results of this study suggest multiple mechanisms of dendritic protein targeting. Notably, dynein-dynactin plays a role in excluding dendritic Dscam, but not Rdl, from axons by retrograde transport.

Show MeSH
Related in: MedlinePlus