Modulation of human stromelysin 3 promoter activity and gene expression by human breast cancer cells
A Ahmad1, J F Marshall, P Basset
1Division of Oncology, United Medical and Dental School, London, UK. ahmad@icrf.icnet.uk
Abstract:
The matrix-degrading enzyme family of matrix-metalloproteinases (MMPs) has been implicated in the process of tumour metastasis. Cellular protein and RNA localisation techniques have been used to show that, whilst several MMP genes are expressed in both cancer and stromal cells, stromelysin 3 is expressed only in stromal fibroblasts adjacent to cancer cells. Immunohistochemical and in situ hybridisation evidence suggests that neoplastic cells can stimulate stromal cell MMP production either in a paracrine fashion or by a cell-cell contact mechanism. Using 2 different lengths of the human stromelysin 3 (ST3) gene 5' flanking sequence cloned upstream of luciferase and CAT reporter genes, we now show that human breast cancer cells can directly activate the ST3 promoter. The putative response element in the ST3 promoter, which lies between 0.46 and 3.4 kb upstream of the transcription start site, is able to effect a 2- to 3-fold increase in downstream gene expression. We further show that this transcriptional up-regulation definitely occurs via a paracrine, and possibly via a cell-cell contact, mechanism. Confirmation that this ST3 promoter activation results in ST3 gene induction of a similar magnitude was shown using Northern blotting of stimulated fibroblasts. Our data provide further evidence that cancer cells can induce fibroblast MMP expression and help to explain the in vivo expression pattern of ST3 in breast cancer.
Insights
Cancer cells stimulate stromal fibroblasts to produce matrix-metalloproteinases (MMPs), specifically stromelysin 3 (ST3). This study identifies a mechanism where cancer cells directly activate the ST3 gene promoter in adjacent stromal cells, driving MMP expression crucial for tumor metastasis.
Area of Science:
- Oncology
- Molecular Biology
- Biochemistry
Background:
- Matrix-metalloproteinases (MMPs) are enzymes involved in tumor metastasis.
- Stromelysin 3 (ST3) is notably expressed in stromal fibroblasts near cancer cells, suggesting a role in tumor microenvironment modulation.
Purpose of the Study:
- To investigate the mechanism by which cancer cells influence stromal cell MMP production, focusing on stromelysin 3.
- To determine if human breast cancer cells can directly activate the ST3 gene promoter and elucidate the signaling pathways involved.
Main Methods:
- Cloning of human stromelysin 3 (ST3) gene 5' flanking sequences upstream of luciferase and CAT reporter genes.
- Utilizing reporter gene assays to assess promoter activity in response to cancer cell co-culture.
- Employing Northern blotting to confirm ST3 gene induction in stimulated fibroblasts.
Main Results:
- Human breast cancer cells directly activate the ST3 promoter, leading to a 2- to 3-fold increase in downstream gene expression.
- Transcriptional up-regulation of ST3 occurs via paracrine signaling and potentially cell-cell contact mechanisms.
- Activated ST3 promoter activity correlates with increased ST3 gene expression in fibroblasts, confirming the induction.
Conclusions:
- Cancer cells can induce fibroblast MMP expression, providing a molecular basis for the observed in vivo expression patterns of ST3 in breast cancer.
- The findings elucidate a key interaction in the tumor microenvironment where cancer cells orchestrate stromal cell activity to facilitate metastasis.
