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Proteolytic Signal Crosstalk in the Prostate Cancer Microenvironment: PC3 Cell Metalloproteinases and Autocrine‐paracrine‐fibroblast Regulation of Proteinase‐activated Receptors (PARs)
Author(s) -
Hollenberg Morley,
Mihara Koichiro
Publication year - 2021
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.2021.35.s1.05158
Subject(s) - paracrine signalling , autocrine signalling , microbiology and biotechnology , biology , cell culture , cancer cell , cancer research , receptor , chemistry , cancer , biochemistry , genetics
Tumour‐associated fibroblasts(TAFs) and non‐tumour epithelial cells (NT‐EPCs) are known to respond to tumour‐derived microenvironment agonists. We hypothesized that prostate cancer‐derived PC3 cells can signal to TAFs and NT‐EPCs by generating proteinases that can regulate proteinase‐activated receptors(PARs). We thus expressed Dual‐Tagged N‐terminal‐mCherry/C‐terminal‐YFP PAR1 in PC3 cells and in a non‐tumour prostate epithelial cell line, RWPE. N‐terminal N‐luciferase(LUC)‐tagged PARs 1,2 and 4 were expressed in WI38 fibroblasts. Upon live cell imaging, intact expressed dual‐tagged PARs appear ‘yellow’; cleaved dual‐tagged‐PARs appear green (loss of N‐terminal mCherry). Cleavage of N‐LUC‐PARs expressed in WI38 cells releases a N‐LUC fluorescence signal into the supernatant. Imaging showed that PC3 cell‐expressed dual‐tagged PAR1 appeared ‘green’, demonstrating autocrine PAR cleavage. Addition of a general MMP inhibitor(GM6001) blocked PC3‐expressed dual‐tagged‐PAR1 cleavage (yellow receptor) as did CRISPR‐elimination of PC3‐expressed MMP2. PC3‐derived supernatants cleaved tagged PAR1 expressed in RWPE cells (green). PC3‐derived supernatants also cleaved all of WI38 cell expressed N‐LUC‐PARs 1, 2 & 4, releasing fluorescence from the cells. We conclude that prostate cancer‐derived PC3 cells produce PAR‐regulating proteinases, including MMP2, that can regulate tumour and non‐tumour cell PAR signalling by an autocrine and paracrine mechanism in a tumour microenvironment.