Limits...
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.

Show MeSH

Related in: MedlinePlus

Anti–Lva Fab antibody inhibits furrow progression. (Top) Coomassie stained 10% SDS-polyacrylamide gels show affinity-purified anti–GST IgG and Fab antibodies (left), and affinity-purified anti–Lva IgG and Fab antibodies (right). About 5 μg of each sample was loaded in the presence of β-mercaptoethanol. Arrows indicate intact heavy chains (HC) and light chains (LC), in addition to heavy chain and light chain fragments, HC Fab and LC Fab, respectively. (Bottom) Two Myo-GFP embryos shown 65 min after being injected at the beginning of nuclear cycle 14 with either anti–GST Fab (5 mg/ml, left), or anti–Lva Fab (8.5 mg/ml, right) antibodies. The large Myo-GFP puncta observed in anti–Lva IgG-injected embryos are not observed in anti–Lva Fab antibody-injected embryos. The white arrowheads indicate the injection site. Views are sagittal. Bar, 10 μm.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2169433&req=5

Figure 7: Anti–Lva Fab antibody inhibits furrow progression. (Top) Coomassie stained 10% SDS-polyacrylamide gels show affinity-purified anti–GST IgG and Fab antibodies (left), and affinity-purified anti–Lva IgG and Fab antibodies (right). About 5 μg of each sample was loaded in the presence of β-mercaptoethanol. Arrows indicate intact heavy chains (HC) and light chains (LC), in addition to heavy chain and light chain fragments, HC Fab and LC Fab, respectively. (Bottom) Two Myo-GFP embryos shown 65 min after being injected at the beginning of nuclear cycle 14 with either anti–GST Fab (5 mg/ml, left), or anti–Lva Fab (8.5 mg/ml, right) antibodies. The large Myo-GFP puncta observed in anti–Lva IgG-injected embryos are not observed in anti–Lva Fab antibody-injected embryos. The white arrowheads indicate the injection site. Views are sagittal. Bar, 10 μm.

Mentions: Furrow progression is also inhibited upon injecting monovalent anti–Lva Fab antibody, while injections of anti–GST Fab antibody has no effect (Fig. 7). The relatively uniform effect of the anti–Lva Fab antibody on furrow progression appears to result from the free diffusion of the antibody within the embryo, in contrast to the IgG, which remains concentrated near the site of injection. In addition, the anti–Lva Fab antibody does not induce the formation of Myo-GFP puncta, as seen with the anti–Lva IgG. This observation indicates that the formation of Myo-GFP puncta in anti–Lva IgG-injected embryos is not the cause of the furrowing inhibition.


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)

Anti–Lva Fab antibody inhibits furrow progression. (Top) Coomassie stained 10% SDS-polyacrylamide gels show affinity-purified anti–GST IgG and Fab antibodies (left), and affinity-purified anti–Lva IgG and Fab antibodies (right). About 5 μg of each sample was loaded in the presence of β-mercaptoethanol. Arrows indicate intact heavy chains (HC) and light chains (LC), in addition to heavy chain and light chain fragments, HC Fab and LC Fab, respectively. (Bottom) Two Myo-GFP embryos shown 65 min after being injected at the beginning of nuclear cycle 14 with either anti–GST Fab (5 mg/ml, left), or anti–Lva Fab (8.5 mg/ml, right) antibodies. The large Myo-GFP puncta observed in anti–Lva IgG-injected embryos are not observed in anti–Lva Fab antibody-injected embryos. The white arrowheads indicate the injection site. Views are sagittal. Bar, 10 μm.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2169433&req=5

Figure 7: Anti–Lva Fab antibody inhibits furrow progression. (Top) Coomassie stained 10% SDS-polyacrylamide gels show affinity-purified anti–GST IgG and Fab antibodies (left), and affinity-purified anti–Lva IgG and Fab antibodies (right). About 5 μg of each sample was loaded in the presence of β-mercaptoethanol. Arrows indicate intact heavy chains (HC) and light chains (LC), in addition to heavy chain and light chain fragments, HC Fab and LC Fab, respectively. (Bottom) Two Myo-GFP embryos shown 65 min after being injected at the beginning of nuclear cycle 14 with either anti–GST Fab (5 mg/ml, left), or anti–Lva Fab (8.5 mg/ml, right) antibodies. The large Myo-GFP puncta observed in anti–Lva IgG-injected embryos are not observed in anti–Lva Fab antibody-injected embryos. The white arrowheads indicate the injection site. Views are sagittal. Bar, 10 μm.
Mentions: Furrow progression is also inhibited upon injecting monovalent anti–Lva Fab antibody, while injections of anti–GST Fab antibody has no effect (Fig. 7). The relatively uniform effect of the anti–Lva Fab antibody on furrow progression appears to result from the free diffusion of the antibody within the embryo, in contrast to the IgG, which remains concentrated near the site of injection. In addition, the anti–Lva Fab antibody does not induce the formation of Myo-GFP puncta, as seen with the anti–Lva IgG. This observation indicates that the formation of Myo-GFP puncta in anti–Lva IgG-injected embryos is not the cause of the furrowing inhibition.

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