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The Abl/enabled signaling pathway regulates Golgi architecture in Drosophila photoreceptor neurons.

Kannan R, Kuzina I, Wincovitch S, Nowotarski SH, Giniger E - Mol. Biol. Cell (2014)

Bottom Line: The Abl effector, Enabled (Ena), selectively labels the cis-Golgi in developing PRs.Finally, we demonstrate that the effects of Abl signaling on Golgi are mediated via regulation of the actin cytoskeleton.Moreover, they raise the possibility that some of the effects of Abl signaling may arise, in part, from alterations of protein trafficking and secretion.

View Article: PubMed Central - PubMed

Affiliation: Axon Guidance and Neural Connectivity Unit, Basic Neuroscience Program, National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, MD 20892.

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Enabled is a marker of the cis-Golgi compartment in wild-type larval photoreceptor (PR) neurons. (A, B) Eye disks of third-instar larvae were analyzed by immunofluorescence with the indicated markers. White boxed region is shown enlarged to the right of the respective panels. Irregular outlines in white highlight a single PR cell body. (A) Projected confocal micrograph of PR clusters stained for Ena (red), cis-Golgi marker GM130 (green), and Elav (to label PR nuclei; blue). Note that Ena-enriched structures in the PR cell body often colabel with the cis-Golgi marker GM130. Scale bar, 5 μm. (B) Projected Z-stacks of PR cluster showing the relative distribution of Ena (red) with trans-Golgi marker dSyx16 (green). Ena structures were juxtaposed with trans-Golgi cisternae. Scale bar, 5 μm. (C) Fluorescence intensity plot of a 3-μm slice (white line in inset) across the cis-Golgi (green) in a single focal plane of a two-color STED micrograph. Note the close agreement between Ena (red) and cis-Golgi (green) at a lateral resolution of ∼100 nm. (D) Immuno–electron micrograph of wild-type eye disk. (D′) Same image with key features marked. Black arrows indicate anti-Ena antibody labeling detected with Nanogold secondary (dense black dots). Open white arrowheads highlight Golgi cisternae. Nuclear envelope (dotted cyan line) and chromatin are indicated for reference. Note that the classic appearance of Golgi as thin, stacked leaflets applies primarily to the medial and trans compartments, whereas the cis-Golgi tends to be thicker and more irregular. Scale bar, 100 nm. Image was collected at 30,000× magnification. (E) Elav-G4>UAS-α-mannosidase II-eGFP. Top row: Tissue stained for GM130 (red), GFP (green), and Elav for PR nuclei (blue). As expected, α-mannosidase II-eGFP colocalized extensively but not perfectly with anti-GM130 (Zuber et al., 2000). Bottom row: Ena in red, GFP in green, and Elav for PR nuclei in blue. Scale bar, 5 μm.
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Figure 1: Enabled is a marker of the cis-Golgi compartment in wild-type larval photoreceptor (PR) neurons. (A, B) Eye disks of third-instar larvae were analyzed by immunofluorescence with the indicated markers. White boxed region is shown enlarged to the right of the respective panels. Irregular outlines in white highlight a single PR cell body. (A) Projected confocal micrograph of PR clusters stained for Ena (red), cis-Golgi marker GM130 (green), and Elav (to label PR nuclei; blue). Note that Ena-enriched structures in the PR cell body often colabel with the cis-Golgi marker GM130. Scale bar, 5 μm. (B) Projected Z-stacks of PR cluster showing the relative distribution of Ena (red) with trans-Golgi marker dSyx16 (green). Ena structures were juxtaposed with trans-Golgi cisternae. Scale bar, 5 μm. (C) Fluorescence intensity plot of a 3-μm slice (white line in inset) across the cis-Golgi (green) in a single focal plane of a two-color STED micrograph. Note the close agreement between Ena (red) and cis-Golgi (green) at a lateral resolution of ∼100 nm. (D) Immuno–electron micrograph of wild-type eye disk. (D′) Same image with key features marked. Black arrows indicate anti-Ena antibody labeling detected with Nanogold secondary (dense black dots). Open white arrowheads highlight Golgi cisternae. Nuclear envelope (dotted cyan line) and chromatin are indicated for reference. Note that the classic appearance of Golgi as thin, stacked leaflets applies primarily to the medial and trans compartments, whereas the cis-Golgi tends to be thicker and more irregular. Scale bar, 100 nm. Image was collected at 30,000× magnification. (E) Elav-G4>UAS-α-mannosidase II-eGFP. Top row: Tissue stained for GM130 (red), GFP (green), and Elav for PR nuclei (blue). As expected, α-mannosidase II-eGFP colocalized extensively but not perfectly with anti-GM130 (Zuber et al., 2000). Bottom row: Ena in red, GFP in green, and Elav for PR nuclei in blue. Scale bar, 5 μm.

Mentions: We found that the Abl antagonist Ena is associated with the cis-Golgi compartment in wild-type Drosophila photoreceptor (PR) neurons (Figure 1, A and C– E). In late third-instar eye imaginal disks, Ena is localized in three subcellular compartments in PR neuronal cell bodies. These are 1) actin-rich apical microvilli-like structures that at later stages develop into mature rhabdomeres; 2) the cortical actin cytoskeleton, which shows diffuse and uniform accumulation of Ena; and 3) distinct perinuclear flattened structures in the cytoplasm. We found strong colabeling of these Ena-enriched flattened structures with GM130, a bona fide marker for the cis-Golgi compartment of the Golgi complex (Figure 1A). Approximately 72.2% ± 1.0 (percent ± SEM) of Ena puncta were associated with GM130 cis-Golgi structures (n = 769 puncta from 11 wild-type disks). Ena structures do not show obvious overlap with other endomembrane compartments, such as early endosomes (Rab5-GFP), late endosomes and lysosomes (Rab11), or centrosomes (CNN–green fluorescent protein [GFP]; unpublished data).


The Abl/enabled signaling pathway regulates Golgi architecture in Drosophila photoreceptor neurons.

Kannan R, Kuzina I, Wincovitch S, Nowotarski SH, Giniger E - Mol. Biol. Cell (2014)

Enabled is a marker of the cis-Golgi compartment in wild-type larval photoreceptor (PR) neurons. (A, B) Eye disks of third-instar larvae were analyzed by immunofluorescence with the indicated markers. White boxed region is shown enlarged to the right of the respective panels. Irregular outlines in white highlight a single PR cell body. (A) Projected confocal micrograph of PR clusters stained for Ena (red), cis-Golgi marker GM130 (green), and Elav (to label PR nuclei; blue). Note that Ena-enriched structures in the PR cell body often colabel with the cis-Golgi marker GM130. Scale bar, 5 μm. (B) Projected Z-stacks of PR cluster showing the relative distribution of Ena (red) with trans-Golgi marker dSyx16 (green). Ena structures were juxtaposed with trans-Golgi cisternae. Scale bar, 5 μm. (C) Fluorescence intensity plot of a 3-μm slice (white line in inset) across the cis-Golgi (green) in a single focal plane of a two-color STED micrograph. Note the close agreement between Ena (red) and cis-Golgi (green) at a lateral resolution of ∼100 nm. (D) Immuno–electron micrograph of wild-type eye disk. (D′) Same image with key features marked. Black arrows indicate anti-Ena antibody labeling detected with Nanogold secondary (dense black dots). Open white arrowheads highlight Golgi cisternae. Nuclear envelope (dotted cyan line) and chromatin are indicated for reference. Note that the classic appearance of Golgi as thin, stacked leaflets applies primarily to the medial and trans compartments, whereas the cis-Golgi tends to be thicker and more irregular. Scale bar, 100 nm. Image was collected at 30,000× magnification. (E) Elav-G4>UAS-α-mannosidase II-eGFP. Top row: Tissue stained for GM130 (red), GFP (green), and Elav for PR nuclei (blue). As expected, α-mannosidase II-eGFP colocalized extensively but not perfectly with anti-GM130 (Zuber et al., 2000). Bottom row: Ena in red, GFP in green, and Elav for PR nuclei in blue. Scale bar, 5 μm.
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Figure 1: Enabled is a marker of the cis-Golgi compartment in wild-type larval photoreceptor (PR) neurons. (A, B) Eye disks of third-instar larvae were analyzed by immunofluorescence with the indicated markers. White boxed region is shown enlarged to the right of the respective panels. Irregular outlines in white highlight a single PR cell body. (A) Projected confocal micrograph of PR clusters stained for Ena (red), cis-Golgi marker GM130 (green), and Elav (to label PR nuclei; blue). Note that Ena-enriched structures in the PR cell body often colabel with the cis-Golgi marker GM130. Scale bar, 5 μm. (B) Projected Z-stacks of PR cluster showing the relative distribution of Ena (red) with trans-Golgi marker dSyx16 (green). Ena structures were juxtaposed with trans-Golgi cisternae. Scale bar, 5 μm. (C) Fluorescence intensity plot of a 3-μm slice (white line in inset) across the cis-Golgi (green) in a single focal plane of a two-color STED micrograph. Note the close agreement between Ena (red) and cis-Golgi (green) at a lateral resolution of ∼100 nm. (D) Immuno–electron micrograph of wild-type eye disk. (D′) Same image with key features marked. Black arrows indicate anti-Ena antibody labeling detected with Nanogold secondary (dense black dots). Open white arrowheads highlight Golgi cisternae. Nuclear envelope (dotted cyan line) and chromatin are indicated for reference. Note that the classic appearance of Golgi as thin, stacked leaflets applies primarily to the medial and trans compartments, whereas the cis-Golgi tends to be thicker and more irregular. Scale bar, 100 nm. Image was collected at 30,000× magnification. (E) Elav-G4>UAS-α-mannosidase II-eGFP. Top row: Tissue stained for GM130 (red), GFP (green), and Elav for PR nuclei (blue). As expected, α-mannosidase II-eGFP colocalized extensively but not perfectly with anti-GM130 (Zuber et al., 2000). Bottom row: Ena in red, GFP in green, and Elav for PR nuclei in blue. Scale bar, 5 μm.
Mentions: We found that the Abl antagonist Ena is associated with the cis-Golgi compartment in wild-type Drosophila photoreceptor (PR) neurons (Figure 1, A and C– E). In late third-instar eye imaginal disks, Ena is localized in three subcellular compartments in PR neuronal cell bodies. These are 1) actin-rich apical microvilli-like structures that at later stages develop into mature rhabdomeres; 2) the cortical actin cytoskeleton, which shows diffuse and uniform accumulation of Ena; and 3) distinct perinuclear flattened structures in the cytoplasm. We found strong colabeling of these Ena-enriched flattened structures with GM130, a bona fide marker for the cis-Golgi compartment of the Golgi complex (Figure 1A). Approximately 72.2% ± 1.0 (percent ± SEM) of Ena puncta were associated with GM130 cis-Golgi structures (n = 769 puncta from 11 wild-type disks). Ena structures do not show obvious overlap with other endomembrane compartments, such as early endosomes (Rab5-GFP), late endosomes and lysosomes (Rab11), or centrosomes (CNN–green fluorescent protein [GFP]; unpublished data).

Bottom Line: The Abl effector, Enabled (Ena), selectively labels the cis-Golgi in developing PRs.Finally, we demonstrate that the effects of Abl signaling on Golgi are mediated via regulation of the actin cytoskeleton.Moreover, they raise the possibility that some of the effects of Abl signaling may arise, in part, from alterations of protein trafficking and secretion.

View Article: PubMed Central - PubMed

Affiliation: Axon Guidance and Neural Connectivity Unit, Basic Neuroscience Program, National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, MD 20892.

Show MeSH
Related in: MedlinePlus