Revealing the three dimensional architecture of focal adhesion components to explain Ca2 +-mediated turnover of focal adhesions

Shu Jing Chang, Ying Chi Chen, Chi Hsun Yang, Soon Cen Huang, Ho Kai Huang, Chun Chun Li, Hans I.Chen Harn, Wen Tai Chiu

Research output: Contribution to journalArticle

3 Citations (Scopus)

Abstract

Background Focal adhesions (FAs) are large, dynamic protein complexes located close to the plasma membrane, which serve as the mechanical linkages and a biochemical signaling hub of cells. The coordinated and dynamic regulation of focal adhesion is required for cell migration. Degradation, or turnover, of FAs is a major event at the trailing edge of a migratory cell, and is mediated by Ca2 +/calpain-dependent proteolysis and disassembly. Here, we investigated how Ca2 + influx induces cascades of FA turnover in living cells. Methods Images obtained with a total internal reflection fluorescence microscope (TIRFM) showed that Ca2 + ions induce different processes in the FA molecules focal adhesion kinase (FAK), paxillin, vinculin, and talin. Three mutated calpain-resistant FA molecules, FAK-V744G, paxillin-S95G, and talin-L432G, were used to clarify the role of each FA molecule in FA turnover. Results Vinculin was resistant to degradation and was not significantly affected by the presence of mutated calpain-resistant FA molecules. In contrast, talin was more sensitive to calpain-mediated turnover than the other molecules. Three-dimensional (3D) fluorescence imaging and immunoblotting demonstrated that outer FA molecules were more sensitive to calpain-mediated proteolysis than internal FA molecules. Furthermore, cell contraction is not involved in degradation of FA. Conclusions These results suggest that Ca2 +-mediated degradation of FAs was mediated by both proteolysis and disassembly. The 3D architecture of FAs is related to the different dynamics of FA molecule degradation during Ca2 +-mediated FA turnover. General significance This study will help us to clearly understand the underlying mechanism of focal adhesion turnover by Ca2 +.

Original languageEnglish
Pages (from-to)624-635
Number of pages12
JournalBiochimica et Biophysica Acta - General Subjects
Volume1861
Issue number3
DOIs
Publication statusPublished - 2017 Mar 1

Fingerprint

Focal Adhesions
Adhesion
Calpain
Molecules
Talin
Proteolysis
Degradation
Paxillin
Vinculin
Focal Adhesion Protein-Tyrosine Kinases
Fluorescence
Optical Imaging
Cell membranes

All Science Journal Classification (ASJC) codes

  • Biophysics
  • Biochemistry
  • Molecular Biology

Cite this

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title = "Revealing the three dimensional architecture of focal adhesion components to explain Ca2 +-mediated turnover of focal adhesions",
abstract = "Background Focal adhesions (FAs) are large, dynamic protein complexes located close to the plasma membrane, which serve as the mechanical linkages and a biochemical signaling hub of cells. The coordinated and dynamic regulation of focal adhesion is required for cell migration. Degradation, or turnover, of FAs is a major event at the trailing edge of a migratory cell, and is mediated by Ca2 +/calpain-dependent proteolysis and disassembly. Here, we investigated how Ca2 + influx induces cascades of FA turnover in living cells. Methods Images obtained with a total internal reflection fluorescence microscope (TIRFM) showed that Ca2 + ions induce different processes in the FA molecules focal adhesion kinase (FAK), paxillin, vinculin, and talin. Three mutated calpain-resistant FA molecules, FAK-V744G, paxillin-S95G, and talin-L432G, were used to clarify the role of each FA molecule in FA turnover. Results Vinculin was resistant to degradation and was not significantly affected by the presence of mutated calpain-resistant FA molecules. In contrast, talin was more sensitive to calpain-mediated turnover than the other molecules. Three-dimensional (3D) fluorescence imaging and immunoblotting demonstrated that outer FA molecules were more sensitive to calpain-mediated proteolysis than internal FA molecules. Furthermore, cell contraction is not involved in degradation of FA. Conclusions These results suggest that Ca2 +-mediated degradation of FAs was mediated by both proteolysis and disassembly. The 3D architecture of FAs is related to the different dynamics of FA molecule degradation during Ca2 +-mediated FA turnover. General significance This study will help us to clearly understand the underlying mechanism of focal adhesion turnover by Ca2 +.",
author = "Chang, {Shu Jing} and Chen, {Ying Chi} and Yang, {Chi Hsun} and Huang, {Soon Cen} and Huang, {Ho Kai} and Li, {Chun Chun} and Harn, {Hans I.Chen} and Chiu, {Wen Tai}",
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Revealing the three dimensional architecture of focal adhesion components to explain Ca2 +-mediated turnover of focal adhesions. / Chang, Shu Jing; Chen, Ying Chi; Yang, Chi Hsun; Huang, Soon Cen; Huang, Ho Kai; Li, Chun Chun; Harn, Hans I.Chen; Chiu, Wen Tai.

In: Biochimica et Biophysica Acta - General Subjects, Vol. 1861, No. 3, 01.03.2017, p. 624-635.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Revealing the three dimensional architecture of focal adhesion components to explain Ca2 +-mediated turnover of focal adhesions

AU - Chang, Shu Jing

AU - Chen, Ying Chi

AU - Yang, Chi Hsun

AU - Huang, Soon Cen

AU - Huang, Ho Kai

AU - Li, Chun Chun

AU - Harn, Hans I.Chen

AU - Chiu, Wen Tai

PY - 2017/3/1

Y1 - 2017/3/1

N2 - Background Focal adhesions (FAs) are large, dynamic protein complexes located close to the plasma membrane, which serve as the mechanical linkages and a biochemical signaling hub of cells. The coordinated and dynamic regulation of focal adhesion is required for cell migration. Degradation, or turnover, of FAs is a major event at the trailing edge of a migratory cell, and is mediated by Ca2 +/calpain-dependent proteolysis and disassembly. Here, we investigated how Ca2 + influx induces cascades of FA turnover in living cells. Methods Images obtained with a total internal reflection fluorescence microscope (TIRFM) showed that Ca2 + ions induce different processes in the FA molecules focal adhesion kinase (FAK), paxillin, vinculin, and talin. Three mutated calpain-resistant FA molecules, FAK-V744G, paxillin-S95G, and talin-L432G, were used to clarify the role of each FA molecule in FA turnover. Results Vinculin was resistant to degradation and was not significantly affected by the presence of mutated calpain-resistant FA molecules. In contrast, talin was more sensitive to calpain-mediated turnover than the other molecules. Three-dimensional (3D) fluorescence imaging and immunoblotting demonstrated that outer FA molecules were more sensitive to calpain-mediated proteolysis than internal FA molecules. Furthermore, cell contraction is not involved in degradation of FA. Conclusions These results suggest that Ca2 +-mediated degradation of FAs was mediated by both proteolysis and disassembly. The 3D architecture of FAs is related to the different dynamics of FA molecule degradation during Ca2 +-mediated FA turnover. General significance This study will help us to clearly understand the underlying mechanism of focal adhesion turnover by Ca2 +.

AB - Background Focal adhesions (FAs) are large, dynamic protein complexes located close to the plasma membrane, which serve as the mechanical linkages and a biochemical signaling hub of cells. The coordinated and dynamic regulation of focal adhesion is required for cell migration. Degradation, or turnover, of FAs is a major event at the trailing edge of a migratory cell, and is mediated by Ca2 +/calpain-dependent proteolysis and disassembly. Here, we investigated how Ca2 + influx induces cascades of FA turnover in living cells. Methods Images obtained with a total internal reflection fluorescence microscope (TIRFM) showed that Ca2 + ions induce different processes in the FA molecules focal adhesion kinase (FAK), paxillin, vinculin, and talin. Three mutated calpain-resistant FA molecules, FAK-V744G, paxillin-S95G, and talin-L432G, were used to clarify the role of each FA molecule in FA turnover. Results Vinculin was resistant to degradation and was not significantly affected by the presence of mutated calpain-resistant FA molecules. In contrast, talin was more sensitive to calpain-mediated turnover than the other molecules. Three-dimensional (3D) fluorescence imaging and immunoblotting demonstrated that outer FA molecules were more sensitive to calpain-mediated proteolysis than internal FA molecules. Furthermore, cell contraction is not involved in degradation of FA. Conclusions These results suggest that Ca2 +-mediated degradation of FAs was mediated by both proteolysis and disassembly. The 3D architecture of FAs is related to the different dynamics of FA molecule degradation during Ca2 +-mediated FA turnover. General significance This study will help us to clearly understand the underlying mechanism of focal adhesion turnover by Ca2 +.

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U2 - 10.1016/j.bbagen.2017.01.002

DO - 10.1016/j.bbagen.2017.01.002

M3 - Article

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JO - Biochimica et Biophysica Acta - General Subjects

JF - Biochimica et Biophysica Acta - General Subjects

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