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Characterization of cell-adhesive proteins secreted by cell lines growing in protein- and lipid-free synthetic medium: mouse L.P3 cells secrete a procollagen molecule with potent cell-attachment activity.

Abstract：

:We have previously shown that mouse L.P3 cells secrete fibronectin and a novel protein, gelatin-non-binding and heparin-binding cell-adhesive protein (GNCP). Here, we screened and characterized cell-adhesive proteins in the conditioned media (CM's) of a series of cell lines growing in a protein- and lipid-free synthetic medium (P3 cell lines). Although cell-attachment activity of the CM's ranged from undetectable to highly significant, fractionation with affinity columns revealed the presence of significant cell-attachment activity in all CM's. Using cell-attachment assay and immunoassay on blotted filters, various cell-adhesive proteins were detected in the CM's, and most of them were identified as fibronectin, laminin, vitronectin, and collagen. GNCP-like proteins were detected in the CM's of HeLa.P3, JTC-16.P3, L.P3, and JTC-12.P3. There was no relationship between the origin of the cell lines and the cell-adhesive proteins secreted. GNCP purified from L.P3-CM was separated into 120-, 140-, 150-, and 160-kDa proteins on SDS-PAGE, which were judged to be a type of mouse type I procollagen from the following results: (1) they were digested by collagenase, (2) pepsin treatment converted the 150- and 160-kDa proteins into 120- and 140-kDa proteins, (3) they were recognized by anti-type I collagen antiserum, and (4) amino-terminal sequence of the pepsin-digested 140-kDa protein had significant homology with type I collagen. GNCP showed half-maximum activity of cell attachment at 0.03 micrograms/ml, indicating that GNCP was a cell-adhesive protein with an extremely high specific activity compared to other known cell-adhesive proteins.

journal_name

Cell Struct Funct

journal_title

Cell structure and function

authors

Morita H,Yasumitsu H,Watanabe Y,Miyazaki K,Umeda M

doi

10.1247/csf.18.61

subject

Has Abstract

pub_date

1993-02-01 00:00:00

pages

61-72

issue

eissn

0386-7196

issn

1347-3700

journal_volume

pub_type

杂志文章

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Characterization of cell-adhesive proteins secreted by cell lines growing in protein- and lipid-free synthetic medium: mouse L.P3 cells secrete a procollagen molecule with potent cell-attachment activity.