Characterization of the protease activity that cleaves the extracellular domain of beta-dystroglycan.
Dystroglycan (DG) complex, composed of alphaDG and betaDG, provides a link between the extracellular matrix (ECM) and cortical cytoskeleton. Although the proteolytic processing of betaDG was reported in various physiological and pathological conditions, its exact mechanism remains unknown. In this study, we addressed this issue using the cell culture system of rat schwannoma cell line RT4. We found that the culture medium of RT4 cells was enriched with the protease activity that degrades the fusion protein construct of the extracellular domain of betaDG specifically. This activity was suppressed by the inhibitor of matrix metalloproteinase-2 (MMP-2) and MMP-9, but not by the inhibitors of MMP-1, MMP-3, MMP-8, and MMP-13. Zymography and RT-PCR analysis showed that RT4 cells secreted MMP-2 and MMP-9 into the culture medium. Finally, active MMP-2 and MMP-9 enzymes degraded the fusion protein construct of the extracellular domain of betaDG. These results indicate (1) that RT4 cells secrete the protease activity that degrades the extracellular domain of betaDG specifically and (2) that MMP-2 and MMP-9 may be involved in this process.
Zhong
Di
D
Department of Neurology and Neuroscience, Teikyo University School of Medicine 2-11-1 Kaga, Itabashi-ku, Tokyo 173-8605, Japan.
Saito
Fumiaki
F
Saito
Yuko
Y
Nakamura
Ayami
A
Shimizu
Teruo
T
Matsumura
Kiichiro
K
D000818:Animals; D002478:Cells, Cultured; D017364:Collagenases / Q000378:metabolism; D017077:Culture Media, Conditioned / Q000378:metabolism; D003599:Cytoskeleton / Q000378:metabolism; D049030:Dystroglycans / Q000378:metabolism*; D010450:Endopeptidases / Q000378:metabolism*; D005109:Extracellular Matrix / Q000378:metabolism*; D020781:Matrix Metalloproteinase 1 / Q000378:metabolism; D053509:Matrix Metalloproteinase 13; D020778:Matrix Metalloproteinase 2 / Q000378:metabolism; D019278:Matrix Metalloproteinase 3 / Q000378:metabolism; D020784:Matrix Metalloproteinase 8 / Q000378:metabolism; D020780:Matrix Metalloproteinase 9 / Q000378:metabolism; D020782:Matrix Metalloproteinases / Q000378:metabolism*; D051381:Rats; D020133:Reverse Transcriptase Polymerase Chain Reaction
D017077:Culture Media, Conditioned; D049030:Dystroglycans; D010450:Endopeptidases; D017364:Collagenases; D053509:Matrix Metalloproteinase 13; D020782:Matrix Metalloproteinases; C507083:Mmp13 protein, rat; D019278:Matrix Metalloproteinase 3; D020778:Matrix Metalloproteinase 2; D020784:Matrix Metalloproteinase 8; D020780:Matrix Metalloproteinase 9; D020781:Matrix Metalloproteinase 1