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Clonal variation of expression of the genes coding for plasminogen activators, their inhibitors and the urokinase receptor in HT1080 sarcoma cells
Author(s) -
Laug Walter E.,
Wang Kai,
Mundi Rita,
Rideout William,
Kruithof Egbert K. O.,
Bogenmann Emil
Publication year - 1992
Publication title -
international journal of cancer
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.475
H-Index - 234
eISSN - 1097-0215
pISSN - 0020-7136
DOI - 10.1002/ijc.2910520224
Subject(s) - biology , microbiology and biotechnology , urokinase receptor , plasminogen activator , ht1080 , gene , clone (java method) , blot , gene expression , cell culture , northern blot , lysis , messenger rna , biochemistry , genetics
The human sarcoma cell line HT1080 was found, by in situ hybridization, to consist of cells expressing various levels of urokinase (uPA) and tissue type (tPA) plasminogen activator (PA) suggesting clonal variation of expression of these genes. Colonies originating from single HT1080 cells were, therefore, established and screened for PA activity using a fibrin agarose overlay. Colonies inducing lysis (clone C + and H + ) or no lysis (clones B − and M − ) were isolated and tested for mRNA levels of uPA, tPA, uPA receptor (uPAR) and the 3 PA inhibitors (PAI), PAI‐1, PAI‐2 and protease‐nexin I. The different clones revealed considerable variation of expression of the different PA and PAI genes, with lysis‐inducing clones expressing mainly the PA genes, whereas non‐lysing clones demonstrated higher expression of the PAI genes. Amplification or loss of specific genes was excluded by Southern blotting. The protein levels of cellular and secreted PA and PAI determined by ELISA and Western blots demonstrated a pattern similar to that observed for PA and PAI mRNA concentrations, suggesting clonal differences either on the level of transcription or in RNA processing and/or stability. Due to complex interactions between PA and PAI, neither mRNA nor protein levels of the different genes were predictive for the amount of functional PA activity present in the supernatant or on the cell surface of the different clones. Receptor‐bound uPA activity was found to be considerably higher in lysis‐inducing than in non‐lysing clones and the activity was dependent on neutralization by PAI‐1 rather than on the level of uPAR mRNA. © 1992 Wiley‐Liss, Inc.