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Invadolysin: a novel, conserved metalloprotease links mitotic structural rearrangements with cell migration.

McHugh B, Krause SA, Yu B, Deans AM, Heasman S, McLaughlin P, Heck MM - J. Cell Biol. (2004)

Bottom Line: Zymography reveals that a protease activity, present in wild-type larval brains, is missing from homozygous tissue, and we show that IX-14/invadolysin cleaves lamin in vitro.The IX-14/invadolysin protein is predominantly found in cytoplasmic structures resembling invadopodia in fly and human cells, but is dramatically relocalized to the leading edge of migrating cells.Strikingly, we find that the directed migration of germ cells is affected in Drosophila IX-14 mutant embryos.

View Article: PubMed Central - PubMed

Affiliation: Wellcome Trust Centre for Cell Biology, University of Edinburgh, Edinburgh EH9 3JR, Scotland, UK.

ABSTRACT
The cell cycle is widely known to be regulated by networks of phosphorylation and ubiquitin-directed proteolysis. Here, we describe IX-14/invadolysin, a novel metalloprotease present only in metazoa, whose activity appears to be essential for mitotic progression. Mitotic neuroblasts of Drosophila melanogaster IX-14 mutant larvae exhibit increased levels of nuclear envelope proteins, monopolar and asymmetric spindles, and chromosomes that appear hypercondensed in length with a surrounding halo of loosely condensed chromatin. Zymography reveals that a protease activity, present in wild-type larval brains, is missing from homozygous tissue, and we show that IX-14/invadolysin cleaves lamin in vitro. The IX-14/invadolysin protein is predominantly found in cytoplasmic structures resembling invadopodia in fly and human cells, but is dramatically relocalized to the leading edge of migrating cells. Strikingly, we find that the directed migration of germ cells is affected in Drosophila IX-14 mutant embryos. Thus, invadolysin identifies a new family of conserved metalloproteases whose activity appears to be essential for the coordination of mitotic progression, but which also plays an unexpected role in cell migration.

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Abnormal levels of nuclear envelope proteins in l(3)IX-14 mutant cells. Larval brains from wild-type and IX-14 homozygous mutant animals were processed for lamin and otefin detection. (A) Wild-type mitotic cells (arrow) show homogeneous lamin staining as the nuclear lamina becomes dispersed during mitosis. (B) In contrast, both IX-14 alleles have greatly increased lamin staining during mitosis (arrowhead). Bar, 5 μm. (C) Wild-type neuroblasts show distinct nuclear envelope staining similar to lamin. (D) Both IX-14 alleles also show greatly increased otefin staining during mitosis. Protein extracts from wild-type and IX-14 larval brains were probed for Dm0 lamin (E) and otefin (F). α-tubulin (bottom) served as a loading control, and confirmed that the samples were similarly loaded. The levels of both lamin and otefin detectable by immunoblotting was significantly greater in the mutant tissues than in wild-type brains.
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fig3: Abnormal levels of nuclear envelope proteins in l(3)IX-14 mutant cells. Larval brains from wild-type and IX-14 homozygous mutant animals were processed for lamin and otefin detection. (A) Wild-type mitotic cells (arrow) show homogeneous lamin staining as the nuclear lamina becomes dispersed during mitosis. (B) In contrast, both IX-14 alleles have greatly increased lamin staining during mitosis (arrowhead). Bar, 5 μm. (C) Wild-type neuroblasts show distinct nuclear envelope staining similar to lamin. (D) Both IX-14 alleles also show greatly increased otefin staining during mitosis. Protein extracts from wild-type and IX-14 larval brains were probed for Dm0 lamin (E) and otefin (F). α-tubulin (bottom) served as a loading control, and confirmed that the samples were similarly loaded. The levels of both lamin and otefin detectable by immunoblotting was significantly greater in the mutant tissues than in wild-type brains.

Mentions: As higher eukaryotic cells enter mitosis, chromosome condensation and spindle assembly are accompanied by the disassembly of the nuclear envelope. Surprisingly, the majority of IX-14 mitotic cells showed dramatically increased levels of lamin Dm0 (a B-type lamin), compared with wild-type mitotic cells (Fig. 3, A and B). Strikingly, the level of another Drosophila nuclear envelope protein, otefin, also appeared elevated in IX-14 mitotic cells (Fig. 3, C and D). When double immunofluorescence was performed, we observed a simultaneous increase of lamin and otefin in the same cells (unpublished data). We believe this elevation occurs before mitosis as late G2 cells (those positive for mitosis-specific phosphorylation of Serine 10 on histone H3) also exhibited increased lamin and otefin fluorescence.


Invadolysin: a novel, conserved metalloprotease links mitotic structural rearrangements with cell migration.

McHugh B, Krause SA, Yu B, Deans AM, Heasman S, McLaughlin P, Heck MM - J. Cell Biol. (2004)

Abnormal levels of nuclear envelope proteins in l(3)IX-14 mutant cells. Larval brains from wild-type and IX-14 homozygous mutant animals were processed for lamin and otefin detection. (A) Wild-type mitotic cells (arrow) show homogeneous lamin staining as the nuclear lamina becomes dispersed during mitosis. (B) In contrast, both IX-14 alleles have greatly increased lamin staining during mitosis (arrowhead). Bar, 5 μm. (C) Wild-type neuroblasts show distinct nuclear envelope staining similar to lamin. (D) Both IX-14 alleles also show greatly increased otefin staining during mitosis. Protein extracts from wild-type and IX-14 larval brains were probed for Dm0 lamin (E) and otefin (F). α-tubulin (bottom) served as a loading control, and confirmed that the samples were similarly loaded. The levels of both lamin and otefin detectable by immunoblotting was significantly greater in the mutant tissues than in wild-type brains.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2172566&req=5

fig3: Abnormal levels of nuclear envelope proteins in l(3)IX-14 mutant cells. Larval brains from wild-type and IX-14 homozygous mutant animals were processed for lamin and otefin detection. (A) Wild-type mitotic cells (arrow) show homogeneous lamin staining as the nuclear lamina becomes dispersed during mitosis. (B) In contrast, both IX-14 alleles have greatly increased lamin staining during mitosis (arrowhead). Bar, 5 μm. (C) Wild-type neuroblasts show distinct nuclear envelope staining similar to lamin. (D) Both IX-14 alleles also show greatly increased otefin staining during mitosis. Protein extracts from wild-type and IX-14 larval brains were probed for Dm0 lamin (E) and otefin (F). α-tubulin (bottom) served as a loading control, and confirmed that the samples were similarly loaded. The levels of both lamin and otefin detectable by immunoblotting was significantly greater in the mutant tissues than in wild-type brains.
Mentions: As higher eukaryotic cells enter mitosis, chromosome condensation and spindle assembly are accompanied by the disassembly of the nuclear envelope. Surprisingly, the majority of IX-14 mitotic cells showed dramatically increased levels of lamin Dm0 (a B-type lamin), compared with wild-type mitotic cells (Fig. 3, A and B). Strikingly, the level of another Drosophila nuclear envelope protein, otefin, also appeared elevated in IX-14 mitotic cells (Fig. 3, C and D). When double immunofluorescence was performed, we observed a simultaneous increase of lamin and otefin in the same cells (unpublished data). We believe this elevation occurs before mitosis as late G2 cells (those positive for mitosis-specific phosphorylation of Serine 10 on histone H3) also exhibited increased lamin and otefin fluorescence.

Bottom Line: Zymography reveals that a protease activity, present in wild-type larval brains, is missing from homozygous tissue, and we show that IX-14/invadolysin cleaves lamin in vitro.The IX-14/invadolysin protein is predominantly found in cytoplasmic structures resembling invadopodia in fly and human cells, but is dramatically relocalized to the leading edge of migrating cells.Strikingly, we find that the directed migration of germ cells is affected in Drosophila IX-14 mutant embryos.

View Article: PubMed Central - PubMed

Affiliation: Wellcome Trust Centre for Cell Biology, University of Edinburgh, Edinburgh EH9 3JR, Scotland, UK.

ABSTRACT
The cell cycle is widely known to be regulated by networks of phosphorylation and ubiquitin-directed proteolysis. Here, we describe IX-14/invadolysin, a novel metalloprotease present only in metazoa, whose activity appears to be essential for mitotic progression. Mitotic neuroblasts of Drosophila melanogaster IX-14 mutant larvae exhibit increased levels of nuclear envelope proteins, monopolar and asymmetric spindles, and chromosomes that appear hypercondensed in length with a surrounding halo of loosely condensed chromatin. Zymography reveals that a protease activity, present in wild-type larval brains, is missing from homozygous tissue, and we show that IX-14/invadolysin cleaves lamin in vitro. The IX-14/invadolysin protein is predominantly found in cytoplasmic structures resembling invadopodia in fly and human cells, but is dramatically relocalized to the leading edge of migrating cells. Strikingly, we find that the directed migration of germ cells is affected in Drosophila IX-14 mutant embryos. Thus, invadolysin identifies a new family of conserved metalloproteases whose activity appears to be essential for the coordination of mitotic progression, but which also plays an unexpected role in cell migration.

Show MeSH
Related in: MedlinePlus