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Biosynthesis of α2(IV) and α1(IV) chains of collagen IV and interactions with matrix metalloproteinase‐9
Author(s) -
Toth Marta,
Sado Yoshikazu,
Ninomiya Yoshifumi,
Fridman Rafael
Publication year - 1999
Publication title -
journal of cellular physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.529
H-Index - 174
eISSN - 1097-4652
pISSN - 0021-9541
DOI - 10.1002/(sici)1097-4652(199907)180:1<131::aid-jcp15>3.0.co;2-s
Subject(s) - immunoprecipitation , ht1080 , microbiology and biotechnology , transfection , biochemistry , biotinylation , cell culture , chemistry , enzyme , monoclonal antibody , in vitro , biology , antibody , gene , genetics , immunology
In vitro binding studies with latent matrix metalloproteinase‐9 (pro‐MMP‐9) have revealed the existence of nondisulfide‐bonded α2(IV) chains on the cell surface capable of forming a high‐affinity complex with the enzyme. Here we investigated the biosynthesis and cellular distribution of α2(IV) and α1(IV) chains in breast epithelial (MCF10A and MDA‐MB‐231) and fibrosarcoma (HT1080) cells by pulse‐chase analysis followed by immunoprecipitation with chain‐specific monoclonal antibodies (mAb). These studies showed that whereas the α1(IV) chain remained in the intracellular compartment, nondisulfide‐bonded α2(IV) chains were secreted into the media in a stable form. Consistently, only α2(IV) was detected on the cell surface by surface biotinylation or indirect immunofluorescence. In agreement with the pulse‐chase analysis, media subjected to coprecipitation experiments with pro‐MMP‐9 or pro‐MMP‐9‐affinity chromatography followed by immunoblotting with chain‐specific mAbs resulted in the detection of α2(IV). A preferential secretion of nondisulfide‐bonded α2(IV) chains was also observed in CHO‐K1 cells transiently transfected with full‐length mouse α2(IV) or α1(IV) cDNAs. However, a complex of mouse α1(IV) with pro‐MMP‐9 was coprecipitated with exogenous enzyme from lysates of CHO‐K1 cells transfected with mouse α1(IV), suggesting that under overexpression conditions the enzyme can also interact with the α1(IV) chain. Collectively, these studies further demonstrate the interactions of pro‐MMP‐9 with collagen IV chains and a unique processing and targeting of nondisulfide‐bonded α2(IV) chains that may play a role in the surface/matrix association of pro‐MMP‐9. J. Cell. Physiol. 180:131–139, 1999. © 1999 Wiley‐Liss, Inc.