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Lava lamp, a novel peripheral golgi protein, is required for Drosophila melanogaster cellularization.

Sisson JC, Field C, Ventura R, Royou A, Sullivan W - J. Cell Biol. (2000)

Bottom Line: Lva is a coiled-coil protein and, unlike other proteins previously implicated in cellularization or cytokinesis, it is Golgi associated.Biochemical analysis demonstrates that Lva physically interacts with the MMAPs Spectrin and CLIP190.We suggest that Lva and Spectrin may form a Golgi-based scaffold that mediates the interaction of Golgi bodies with microtubules and facilitates Golgi-derived membrane secretion required for the formation of furrows during cellularization.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular, Cell and Developmental Biology, Sinsheimer Labs, University of California at Santa Cruz, Santa Cruz, California 95064, USA. sisson@darwin.ucsc.edu

ABSTRACT
Drosophila cellularization and animal cell cytokinesis rely on the coordinated functions of the microfilament and microtubule cytoskeletal systems. To identify new proteins involved in cellularization and cytokinesis, we have conducted a biochemical screen for microfilament/microtubule-associated proteins (MMAPs). 17 MMAPs were identified; seven have been previously implicated in cellularization and/or cytokinesis, including KLP3A, Anillin, Septins, and Dynamin. We now show that a novel MMAP, Lava Lamp (Lva), is also required for cellularization. Lva is a coiled-coil protein and, unlike other proteins previously implicated in cellularization or cytokinesis, it is Golgi associated. Our functional analysis shows that cellularization is dramatically inhibited upon injecting anti-Lva antibodies (IgG and Fab) into embryos. In addition, we show that brefeldin A, a potent inhibitor of membrane trafficking, also inhibits cellularization. Biochemical analysis demonstrates that Lva physically interacts with the MMAPs Spectrin and CLIP190. We suggest that Lva and Spectrin may form a Golgi-based scaffold that mediates the interaction of Golgi bodies with microtubules and facilitates Golgi-derived membrane secretion required for the formation of furrows during cellularization. Our results are consistent with the idea that animal cell cytokinesis depends on both actomyosin-based contraction and Golgi-derived membrane secretion.

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Lva, α-Spectrin, and CLIP190 colocalize to Golgi bodies. Cellularizing embryos were fixed, labeled with pairs of fluorescent probes, and examined by scanning confocal microscopy. From left to right, matched panels show Lva with F-actin, α-Spectrin, CLIP190, and the Golgi marker p120, respectively. White arrowheads indicate corresponding puncta in matched panels, except in the Lva/F-actin panels, where they indicate puncta at the furrow front. The purple arrowhead in the CLIP190 panel indicates CLIP190 with no corresponding Lva fluorescence. (Inset) Cortical F-actin (green) and MT-inverted baskets (red). Essentially no fluorescence was detected with secondary antibodies alone (data not shown). All views are sagittal. Bar, 5 μm.
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Figure 4: Lva, α-Spectrin, and CLIP190 colocalize to Golgi bodies. Cellularizing embryos were fixed, labeled with pairs of fluorescent probes, and examined by scanning confocal microscopy. From left to right, matched panels show Lva with F-actin, α-Spectrin, CLIP190, and the Golgi marker p120, respectively. White arrowheads indicate corresponding puncta in matched panels, except in the Lva/F-actin panels, where they indicate puncta at the furrow front. The purple arrowhead in the CLIP190 panel indicates CLIP190 with no corresponding Lva fluorescence. (Inset) Cortical F-actin (green) and MT-inverted baskets (red). Essentially no fluorescence was detected with secondary antibodies alone (data not shown). All views are sagittal. Bar, 5 μm.

Mentions: During both cellularization and cytokinesis, the coordination of the F-actin and MT cytoskeletal systems contributes to contraction and/or membrane export (Foe et al. 1993; Field et al. 1999). Although the precise mechanism of this dependency relationship is unknown, MTs may act as tracks for recruiting factors required for the establishment and/or maintenance of the contractile ring, or provide structural stability to the contractile ring late in cytokinesis. During cellularization, long MTs grow out from apically positioned centrosomes down over the surface of nuclei and project their plus ends into the embryos interior, forming “inverted baskets” around each nucleus (Foe et al. 1993). Throughout most of cellularization, the contractile apparatus appears to move down along these MTs as it leads the furrow inward at its tip (see Fig. 4, inset) (Foe et al. 1993). When embryos are treated with drugs that destabilize either F-actin or MTs during cellularization, furrow progression is severely disrupted (Zalokar and Erk 1976; Foe and Alberts 1983). The extent to which these treatments disrupt contraction versus membrane export is unknown.


Lava lamp, a novel peripheral golgi protein, is required for Drosophila melanogaster cellularization.

Sisson JC, Field C, Ventura R, Royou A, Sullivan W - J. Cell Biol. (2000)

Lva, α-Spectrin, and CLIP190 colocalize to Golgi bodies. Cellularizing embryos were fixed, labeled with pairs of fluorescent probes, and examined by scanning confocal microscopy. From left to right, matched panels show Lva with F-actin, α-Spectrin, CLIP190, and the Golgi marker p120, respectively. White arrowheads indicate corresponding puncta in matched panels, except in the Lva/F-actin panels, where they indicate puncta at the furrow front. The purple arrowhead in the CLIP190 panel indicates CLIP190 with no corresponding Lva fluorescence. (Inset) Cortical F-actin (green) and MT-inverted baskets (red). Essentially no fluorescence was detected with secondary antibodies alone (data not shown). All views are sagittal. Bar, 5 μm.
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Related In: Results  -  Collection

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Figure 4: Lva, α-Spectrin, and CLIP190 colocalize to Golgi bodies. Cellularizing embryos were fixed, labeled with pairs of fluorescent probes, and examined by scanning confocal microscopy. From left to right, matched panels show Lva with F-actin, α-Spectrin, CLIP190, and the Golgi marker p120, respectively. White arrowheads indicate corresponding puncta in matched panels, except in the Lva/F-actin panels, where they indicate puncta at the furrow front. The purple arrowhead in the CLIP190 panel indicates CLIP190 with no corresponding Lva fluorescence. (Inset) Cortical F-actin (green) and MT-inverted baskets (red). Essentially no fluorescence was detected with secondary antibodies alone (data not shown). All views are sagittal. Bar, 5 μm.
Mentions: During both cellularization and cytokinesis, the coordination of the F-actin and MT cytoskeletal systems contributes to contraction and/or membrane export (Foe et al. 1993; Field et al. 1999). Although the precise mechanism of this dependency relationship is unknown, MTs may act as tracks for recruiting factors required for the establishment and/or maintenance of the contractile ring, or provide structural stability to the contractile ring late in cytokinesis. During cellularization, long MTs grow out from apically positioned centrosomes down over the surface of nuclei and project their plus ends into the embryos interior, forming “inverted baskets” around each nucleus (Foe et al. 1993). Throughout most of cellularization, the contractile apparatus appears to move down along these MTs as it leads the furrow inward at its tip (see Fig. 4, inset) (Foe et al. 1993). When embryos are treated with drugs that destabilize either F-actin or MTs during cellularization, furrow progression is severely disrupted (Zalokar and Erk 1976; Foe and Alberts 1983). The extent to which these treatments disrupt contraction versus membrane export is unknown.

Bottom Line: Lva is a coiled-coil protein and, unlike other proteins previously implicated in cellularization or cytokinesis, it is Golgi associated.Biochemical analysis demonstrates that Lva physically interacts with the MMAPs Spectrin and CLIP190.We suggest that Lva and Spectrin may form a Golgi-based scaffold that mediates the interaction of Golgi bodies with microtubules and facilitates Golgi-derived membrane secretion required for the formation of furrows during cellularization.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular, Cell and Developmental Biology, Sinsheimer Labs, University of California at Santa Cruz, Santa Cruz, California 95064, USA. sisson@darwin.ucsc.edu

ABSTRACT
Drosophila cellularization and animal cell cytokinesis rely on the coordinated functions of the microfilament and microtubule cytoskeletal systems. To identify new proteins involved in cellularization and cytokinesis, we have conducted a biochemical screen for microfilament/microtubule-associated proteins (MMAPs). 17 MMAPs were identified; seven have been previously implicated in cellularization and/or cytokinesis, including KLP3A, Anillin, Septins, and Dynamin. We now show that a novel MMAP, Lava Lamp (Lva), is also required for cellularization. Lva is a coiled-coil protein and, unlike other proteins previously implicated in cellularization or cytokinesis, it is Golgi associated. Our functional analysis shows that cellularization is dramatically inhibited upon injecting anti-Lva antibodies (IgG and Fab) into embryos. In addition, we show that brefeldin A, a potent inhibitor of membrane trafficking, also inhibits cellularization. Biochemical analysis demonstrates that Lva physically interacts with the MMAPs Spectrin and CLIP190. We suggest that Lva and Spectrin may form a Golgi-based scaffold that mediates the interaction of Golgi bodies with microtubules and facilitates Golgi-derived membrane secretion required for the formation of furrows during cellularization. Our results are consistent with the idea that animal cell cytokinesis depends on both actomyosin-based contraction and Golgi-derived membrane secretion.

Show MeSH
Related in: MedlinePlus