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Plasma gelsolin facilitates interaction between β2 glycoprotein I and α5β1 integrin.

Bohgaki M, Matsumoto M, Atsumi T, Kondo T, Yasuda S, Horita T, Nakayama KI, Okumura F, Hatakeyama S, Koike T - J. Cell. Mol. Med. (2011)

Bottom Line: In this study, we identified plasma gelsolin as a protein associated with β(2) GPI by using immunoaffinity chromatography and mass spectrometric analysis.Immunoblot analysis demonstrated that p38 MAPK protein was phosphorylated by monoclonal anti-β(2) GPI antibody treatment, and its phosphorylation was attenuated in the presence of anti-integrin a(5) β(1) antibody.Furthermore, focal adhesion kinase, a downstream molecule of the fibronectin-integrin signalling pathway, was phosphorylated by anti-β(2) GPI antibody treatment.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, Hokkaido University Graduate School of Medicine, Sapporo, Japan.

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Change in intracellular signalling by gelsolin and aCL/β2GPI. (A) Gelsolin and integrin affect phosphorylation of p38 MAPK by anti-β2GPI antibody. RAW264.7 cells were incubated for 2 hrs as indicated after serum-free culture for 16 hrs and then stimulated with aCL/β2GPI (WBCAL1) for 30 min., and then phosphorylation of p38 MAPK was determined by immunoblot analysis using specific antibodies against total-p38 and phospho-p38. (B) Anti-integrin α5β1 antibody inhibits phosphorylation of FAK by aCL/β2GPI. RAW264.7 cells were stimulated with aCL/β2GPI for 10 min. and then phosphorylaion of FAK was determined by immunoblot analysis. (C) aCL/β2GPI increases NF-κB activity. RAW264.7 cells stably expressing kB luciferase reporter were inoculated into a 24-well dish and stimulated as indicated. After stimulation at 37°C for 4 hrs, kB luciferase activity was measured.
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fig05: Change in intracellular signalling by gelsolin and aCL/β2GPI. (A) Gelsolin and integrin affect phosphorylation of p38 MAPK by anti-β2GPI antibody. RAW264.7 cells were incubated for 2 hrs as indicated after serum-free culture for 16 hrs and then stimulated with aCL/β2GPI (WBCAL1) for 30 min., and then phosphorylation of p38 MAPK was determined by immunoblot analysis using specific antibodies against total-p38 and phospho-p38. (B) Anti-integrin α5β1 antibody inhibits phosphorylation of FAK by aCL/β2GPI. RAW264.7 cells were stimulated with aCL/β2GPI for 10 min. and then phosphorylaion of FAK was determined by immunoblot analysis. (C) aCL/β2GPI increases NF-κB activity. RAW264.7 cells stably expressing kB luciferase reporter were inoculated into a 24-well dish and stimulated as indicated. After stimulation at 37°C for 4 hrs, kB luciferase activity was measured.

Mentions: We previously reported that p38-MAPK was phosphorylated in RAW264.7 cells stimulated by human monoclonal aCL/b2GPI [22]. To determine whether a cell surface complex including gelsolin activates RAW264.7 cells, we investigated the phosphorylation of p38-MAPK. Stimulation to RAW264.7 cells by aCL (WBCAL1) showed that p38-MAPK phosphorylation was not induced by plasma gelsolin alone but was induced by aCL/β2GPI stimulation and was further enhanced by plasma gelsolin plus aCL/β2GPI stimulation (Fig. 5A). However, anti-integrin α5β1 antibody attenuated phosphorylation of p38-MAPK by plasma gelsolin plus aCL/β2GPI stimulation (Fig. 5A). These findings indicate that aCL/β2GPI caused phosphorylation of p38-MAPK in collaboration with gelsolin and integrin on the cell surface. Furthermore, to determine the effect on downstream molecules such as focal adhesion kinase FAK, the phosphorylation of FAK by aCL/β2GPI was investigated. Stimulation of aCL/β2GPI and plasma gelsolin resulted in an increased level of phosphorylation of FAK, whereas anti-integrin α5β1 antibody attenuated the phosphorylation of FAK (Fig. 5B). Taken together, the results suggest that anti-β2GPI antibody affects the integrin signalling including its downstream signal molecule FAK, followed by activation of p38-MAPK.


Plasma gelsolin facilitates interaction between β2 glycoprotein I and α5β1 integrin.

Bohgaki M, Matsumoto M, Atsumi T, Kondo T, Yasuda S, Horita T, Nakayama KI, Okumura F, Hatakeyama S, Koike T - J. Cell. Mol. Med. (2011)

Change in intracellular signalling by gelsolin and aCL/β2GPI. (A) Gelsolin and integrin affect phosphorylation of p38 MAPK by anti-β2GPI antibody. RAW264.7 cells were incubated for 2 hrs as indicated after serum-free culture for 16 hrs and then stimulated with aCL/β2GPI (WBCAL1) for 30 min., and then phosphorylation of p38 MAPK was determined by immunoblot analysis using specific antibodies against total-p38 and phospho-p38. (B) Anti-integrin α5β1 antibody inhibits phosphorylation of FAK by aCL/β2GPI. RAW264.7 cells were stimulated with aCL/β2GPI for 10 min. and then phosphorylaion of FAK was determined by immunoblot analysis. (C) aCL/β2GPI increases NF-κB activity. RAW264.7 cells stably expressing kB luciferase reporter were inoculated into a 24-well dish and stimulated as indicated. After stimulation at 37°C for 4 hrs, kB luciferase activity was measured.
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Related In: Results  -  Collection

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fig05: Change in intracellular signalling by gelsolin and aCL/β2GPI. (A) Gelsolin and integrin affect phosphorylation of p38 MAPK by anti-β2GPI antibody. RAW264.7 cells were incubated for 2 hrs as indicated after serum-free culture for 16 hrs and then stimulated with aCL/β2GPI (WBCAL1) for 30 min., and then phosphorylation of p38 MAPK was determined by immunoblot analysis using specific antibodies against total-p38 and phospho-p38. (B) Anti-integrin α5β1 antibody inhibits phosphorylation of FAK by aCL/β2GPI. RAW264.7 cells were stimulated with aCL/β2GPI for 10 min. and then phosphorylaion of FAK was determined by immunoblot analysis. (C) aCL/β2GPI increases NF-κB activity. RAW264.7 cells stably expressing kB luciferase reporter were inoculated into a 24-well dish and stimulated as indicated. After stimulation at 37°C for 4 hrs, kB luciferase activity was measured.
Mentions: We previously reported that p38-MAPK was phosphorylated in RAW264.7 cells stimulated by human monoclonal aCL/b2GPI [22]. To determine whether a cell surface complex including gelsolin activates RAW264.7 cells, we investigated the phosphorylation of p38-MAPK. Stimulation to RAW264.7 cells by aCL (WBCAL1) showed that p38-MAPK phosphorylation was not induced by plasma gelsolin alone but was induced by aCL/β2GPI stimulation and was further enhanced by plasma gelsolin plus aCL/β2GPI stimulation (Fig. 5A). However, anti-integrin α5β1 antibody attenuated phosphorylation of p38-MAPK by plasma gelsolin plus aCL/β2GPI stimulation (Fig. 5A). These findings indicate that aCL/β2GPI caused phosphorylation of p38-MAPK in collaboration with gelsolin and integrin on the cell surface. Furthermore, to determine the effect on downstream molecules such as focal adhesion kinase FAK, the phosphorylation of FAK by aCL/β2GPI was investigated. Stimulation of aCL/β2GPI and plasma gelsolin resulted in an increased level of phosphorylation of FAK, whereas anti-integrin α5β1 antibody attenuated the phosphorylation of FAK (Fig. 5B). Taken together, the results suggest that anti-β2GPI antibody affects the integrin signalling including its downstream signal molecule FAK, followed by activation of p38-MAPK.

Bottom Line: In this study, we identified plasma gelsolin as a protein associated with β(2) GPI by using immunoaffinity chromatography and mass spectrometric analysis.Immunoblot analysis demonstrated that p38 MAPK protein was phosphorylated by monoclonal anti-β(2) GPI antibody treatment, and its phosphorylation was attenuated in the presence of anti-integrin a(5) β(1) antibody.Furthermore, focal adhesion kinase, a downstream molecule of the fibronectin-integrin signalling pathway, was phosphorylated by anti-β(2) GPI antibody treatment.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, Hokkaido University Graduate School of Medicine, Sapporo, Japan.

Show MeSH
Related in: MedlinePlus