Transcriptional Down-regulation of the Metastasis-inducing S100A4 (p9Ka) in Benign but Not in Malignant Rat Mammary Epithelial Cells by GC-factor

Abstract

The S100-related calcium-binding protein S100A4 (p9Ka) is expressed at a low level in rat mammary epithelial cells from normal mammary gland and benign mammary tumors. In transgenic mice, expressed rat S100A4 transgenes co-operate with the activated c-erbB-2 oncogene, neu, to form metastatic mammary tumors. Elevated levels of S100A4 (p9Ka) in cultured benign rat or mouse mammary epithelial cells are associated with the induction of metastatic capability. A cis-acting sequence related to the consensus recognition sequence of GC-factor, 1,300 base pairs upstream of the start site of transcription of the rat S100A4 gene, acts as a cis-acting inhibitor of transcription of the S100A4 (p9Ka) gene in a low S100A4 (p9Ka)-expressing benign rat mammary epithelial cell line, but not in highly expressing rat mammary epithelial cell lines. There is an inverse relationship between the level of S100A4 (p9Ka) mRNA and the level of GC-factor mRNA in a range of rat mammary cell lines. The results suggest a novel mechanism for regulating the expression of the mRNA encoding an S100 protein.S100A4 (p9Ka) is a member of the S100 family of calciumbinding proteins first identified in a cultured myoepitheliallike cell line, Rama 29 (1). The S100A4 (p9Ka) gene has been isolated and its nucleotide sequence determined (2). Elevation of the level of S100A4 (p9Ka) in benign, non-metastatic rat (3) and mouse (4) cells induces metastatic capability in some of the cells. Although mice containing multiple copies of a rat (5) or mouse (6) S100A4 transgene show no observable phenotype, mice which contain cells that express both the rat S100A4 transgenes and the activated neu oncogene, or mouse S100A4 transgenes in a genetic background that yields primary mammary tumors, succumb to metastatic mammary tumors (6, 7), strongly suggesting that overexpression of S100A4 in mammary tumors can yield the metastatic phenotype. Some rat and mouse cultured mammary epithelial cell lines of high metastatic potential contain elevated levels of S100A4, when compared with their non-metastatic counterparts (8, 9).

Network-based analysis of calcium-binding protein genes identifies Grp94 as a target in human oral carcinogenesis

“…A number of Ca 2 þ -binding proteins are thought to be implicated in establishing the malignant and metastatic phenotypes of various tumours (Chen et al, 1997;Takenaga et al, 1997;Van Ginkel et al, 1998), and recent studies have reported altered expression of several Ca 2 þ -binding protein genes in a wide range of human malignancies (Bustin et al, 2001;Pietas et al, 2002;Lakshmikuttyamma et al, 2004;Imazawa et al, 2005). Regarding oral squamous cell carcinomas (OSCCs), aberrant expression of S100 Ca 2 þ -binding protein families seen in tumour sites has been correlated with cancer cell invasion or metastasis (Moriyama-Kita et al, 2004;Tsai et al, 2005).…”

Differential Reactivity of the RatS100A4(p9Ka) Gene to Sodium Bisulfite Is Associated with Differential Levels of the S100A4 (p9Ka) mRNA in Rat Mammary Epithelial Cells

“…The rat S100A4(p9Ka) gene (12,20) does not contain a CpG island located between 2,696 base pairs upstream and 2,474 base pairs downstream of the start site of transcription, in common with the human (30) and mouse (14) S100A4 genes. FIG.…”

“…The intron region, between ϩ135 and ϩ312 bp, contains a positive cis-acting regulatory element found in the first intron of the mouse (16) and rat (29) S100A4 genes. The upstream regulatory region corresponds to DNA between Ϫ1,404 bp and Ϫ1,227 bp 5Ј of the transcriptional start site and contains the repressive GC-factor regulatory sequence element that inhibits transcription of S100A4(p9Ka) mRNA in the low S100A4(p9Ka)-expressing Rama 37CL-A3 cells (12).…”

“…The TATA box region contains DNA between Ϫ113 and ϩ36 base pairs (12,20) relative to the start site of transcription. The intron region, between ϩ135 and ϩ312 bp, contains a positive cis-acting regulatory element found in the first intron of the mouse (16) and rat (29) S100A4 genes.…”

“…In view of the metastasisinducing properties of S100A4 in epithelial cells, it is important to understand mechanisms whereby its level is kept low in these cells. The rat S100A4(p9Ka) gene contains a cis-acting sequence that binds a GC factor-like repressor protein and exerts a repressive effect on the transcription of the S100A4(p9Ka) gene in low S100A4(p9Ka)-expressing rat benign mammary tumor cells but not in high-expressing metastatic or myoepithelial-like rat mammary epithelial cells (12). Hypermethylation is also believed to play a role in the transcriptional silencing of the mouse (mts1) and human (CAPL) S100A4 genes (13)(14)(15).…”