Gravin, a protein upregulated in wounded endothelial cell monolayers, is localized in the cortical cytoplasm

Research output: Contribution to journalArticle

Abstract

Gravin, a 300 kDa intracellular protein, is downregulated in confluent endothelial cell cultures, but is upregulated at the wound edge in wounded endothelial cell monplayers. Inhibition of gravin expression using antisense oligonucleotides inhibits endothelial wound healing suggesting that this protein may play a role in cell spreading and migration. To better understand how gravin may play a role in cell adhesion and migration, the intracellular distribution of gravin in endothelial cells and its relationship to the cytoskeleton were examined. Cultured human umbilical vein endothelial cells were double-labelled with anti-gravin antibodies and probes for cytoskeletal proteins and examined by fluorescence and confocal microscopy. Fluorescence microscopy showed that gravin had a punctate distribution which did not colocalize with actin filaments, microtubules, or intermediate filaments. Treatment of cells with either cytochalasin D or colchicine disrupted all three cytoskeletal components, but did not significantly alter gravin distribution. Three dimensional reconstruction of images collected by confocal microscopy clearly revealed that gravin was concentrated in the cortical cytoplasm adjacent to both the apical and basal plasma membranes. Immunoelectron microscopy confirmed this with gold labelling being associated with the apical and basal plasma membranes. These data indicate that gravin is not a component of stress fibers, microtubules, or intermediate filaments. However, it is hypothesized that gravin is a membrane skeleton protein that plays a role in endothelial wound healing through interactions with components of either the plasma membrane or the membrane skeleton. Studies are underway to test this hypothesis.

Original languageEnglish (US)
JournalFASEB Journal
Volume11
Issue number3
StatePublished - 1997
Externally publishedYes

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Endothelial cells
endothelial cells
intermediate filaments
Monolayers
Cytoplasm
plasma membrane
Cell membranes
Intermediate Filaments
cytoplasm
Endothelial Cells
Cell Membrane
tissue repair
fluorescence microscopy
Fluorescence Microscopy
Skeleton
Confocal Microscopy
Microtubules
Wound Healing
microtubules
Cell Movement

ASJC Scopus subject areas

  • Agricultural and Biological Sciences (miscellaneous)
  • Biochemistry, Genetics and Molecular Biology(all)
  • Biochemistry
  • Cell Biology

Cite this

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title = "Gravin, a protein upregulated in wounded endothelial cell monolayers, is localized in the cortical cytoplasm",
abstract = "Gravin, a 300 kDa intracellular protein, is downregulated in confluent endothelial cell cultures, but is upregulated at the wound edge in wounded endothelial cell monplayers. Inhibition of gravin expression using antisense oligonucleotides inhibits endothelial wound healing suggesting that this protein may play a role in cell spreading and migration. To better understand how gravin may play a role in cell adhesion and migration, the intracellular distribution of gravin in endothelial cells and its relationship to the cytoskeleton were examined. Cultured human umbilical vein endothelial cells were double-labelled with anti-gravin antibodies and probes for cytoskeletal proteins and examined by fluorescence and confocal microscopy. Fluorescence microscopy showed that gravin had a punctate distribution which did not colocalize with actin filaments, microtubules, or intermediate filaments. Treatment of cells with either cytochalasin D or colchicine disrupted all three cytoskeletal components, but did not significantly alter gravin distribution. Three dimensional reconstruction of images collected by confocal microscopy clearly revealed that gravin was concentrated in the cortical cytoplasm adjacent to both the apical and basal plasma membranes. Immunoelectron microscopy confirmed this with gold labelling being associated with the apical and basal plasma membranes. These data indicate that gravin is not a component of stress fibers, microtubules, or intermediate filaments. However, it is hypothesized that gravin is a membrane skeleton protein that plays a role in endothelial wound healing through interactions with components of either the plasma membrane or the membrane skeleton. Studies are underway to test this hypothesis.",
author = "Bryon Grove",
year = "1997",
language = "English (US)",
volume = "11",
journal = "FASEB Journal",
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TY - JOUR

T1 - Gravin, a protein upregulated in wounded endothelial cell monolayers, is localized in the cortical cytoplasm

AU - Grove, Bryon

PY - 1997

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N2 - Gravin, a 300 kDa intracellular protein, is downregulated in confluent endothelial cell cultures, but is upregulated at the wound edge in wounded endothelial cell monplayers. Inhibition of gravin expression using antisense oligonucleotides inhibits endothelial wound healing suggesting that this protein may play a role in cell spreading and migration. To better understand how gravin may play a role in cell adhesion and migration, the intracellular distribution of gravin in endothelial cells and its relationship to the cytoskeleton were examined. Cultured human umbilical vein endothelial cells were double-labelled with anti-gravin antibodies and probes for cytoskeletal proteins and examined by fluorescence and confocal microscopy. Fluorescence microscopy showed that gravin had a punctate distribution which did not colocalize with actin filaments, microtubules, or intermediate filaments. Treatment of cells with either cytochalasin D or colchicine disrupted all three cytoskeletal components, but did not significantly alter gravin distribution. Three dimensional reconstruction of images collected by confocal microscopy clearly revealed that gravin was concentrated in the cortical cytoplasm adjacent to both the apical and basal plasma membranes. Immunoelectron microscopy confirmed this with gold labelling being associated with the apical and basal plasma membranes. These data indicate that gravin is not a component of stress fibers, microtubules, or intermediate filaments. However, it is hypothesized that gravin is a membrane skeleton protein that plays a role in endothelial wound healing through interactions with components of either the plasma membrane or the membrane skeleton. Studies are underway to test this hypothesis.

AB - Gravin, a 300 kDa intracellular protein, is downregulated in confluent endothelial cell cultures, but is upregulated at the wound edge in wounded endothelial cell monplayers. Inhibition of gravin expression using antisense oligonucleotides inhibits endothelial wound healing suggesting that this protein may play a role in cell spreading and migration. To better understand how gravin may play a role in cell adhesion and migration, the intracellular distribution of gravin in endothelial cells and its relationship to the cytoskeleton were examined. Cultured human umbilical vein endothelial cells were double-labelled with anti-gravin antibodies and probes for cytoskeletal proteins and examined by fluorescence and confocal microscopy. Fluorescence microscopy showed that gravin had a punctate distribution which did not colocalize with actin filaments, microtubules, or intermediate filaments. Treatment of cells with either cytochalasin D or colchicine disrupted all three cytoskeletal components, but did not significantly alter gravin distribution. Three dimensional reconstruction of images collected by confocal microscopy clearly revealed that gravin was concentrated in the cortical cytoplasm adjacent to both the apical and basal plasma membranes. Immunoelectron microscopy confirmed this with gold labelling being associated with the apical and basal plasma membranes. These data indicate that gravin is not a component of stress fibers, microtubules, or intermediate filaments. However, it is hypothesized that gravin is a membrane skeleton protein that plays a role in endothelial wound healing through interactions with components of either the plasma membrane or the membrane skeleton. Studies are underway to test this hypothesis.

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