ProEGF cytoplasmic domain (proEGFcyt) is a novel regulator of cathepsin-L secretion and elastinolytic activity in human thyroid carcinoma cells

Abstract

We and others have shown that the cytoplasmic domains of EGF-like ligands have important biological functions. Employing stable transfectants of the human follicular thyroid carcinoma cell line FTC-133 over-expressing the proEGF cytoplasmic domain (FTC-133-proEGFcyt) we show here that proEGFcyt causes a transcriptional up-regulation of cathepsin- (cath-) B and -D and affects the processing of cath-L. Cath-L has strong elastinolytic activity and was the only cathepsin to be secreted by FTC-133-proEGFcyt transfectants. ProEGFcyt significantly impaired the ability of FTC-133 to migrate through elastin matrices when compared with FTC-133 stably transfected with the empty plasmid and a natural proEGFsplice mutant construct. Decreased migration through elastin matrix resulted from decreased secretion of cath-L in FTC-133-proEGFcyt. A similar reduction in elastinolytic activity was observed when FTC-133 control or proEGFsplice form transfectants were incubated with a specific cath-L inhibitor suggesting that this elastinolytic activity detected was largely mediated by cath-L. Down-regulation of cath-L in FTC-133-proEGFcyt involved the upregulation of the t-SNARE component SNAP25 as determined by siRNA knock-down of SNAP25 mRNA. Incubation of FTC-133-proEGFcyt with soluble EGF reversed the effects of proEGFcyt and this antagonistic EGF action was mediated by the EGFR. In summary, we provide first molecular evidence for the suppressive role of proEGFcyt on thyroid carcinoma cell elastinolytic activity and invasiveness and identify novel opposing modular functions of soluble EGF and the cytoplasmic domain of human proEGF.