The new cohort of EMT genes are activated by benzo(a)pyrene and/or SATB1 during the malignant transformation of breast epithelial cells

Tumor metastasis, the migration of tumor cells from a primary site to distant organs, is the leading cause of death among cancer patients. Despite significant advancements in cancer research, much remains to be learned about how tumor cells acquire invasive properties at the molecular level. Here, we investigated the transcriptional program that enables breast epithelial cells to develop highly invasive malignant phenotypes. We focused on genes that promote epithelial-to mesenchymal transition (EMT), a process critical for both embryogenesis and cancer cell invasion, and we compiled a cohort of 339 genes associated with EMT functions with many identified in recent literature. Using an immortalized, non-malignant breast epithelial cell line, MCF10A, we analyzed changes in the transcriptional activity of these genes through Global Run-On Sequencing (GRO-seq) and RNA-seq following stable SATB1 transduction. SATB1 is a nuclear protein with genome-organizing functions that promotes metastatic cancer, including breast cancer. SATB1-expressing MCF10A cells acquire both tumorigenic and metastatic properties. We identified a total of 98 EMT promoting genes that were upregulated in SATB1-expressing MCF10A cells compared to control cells. Additionally, we investigated the effects of the chemical carcinogen benzo[a]pyrene (B[a]P), known to induce malignancy in MCF10A cells. Our findings revealed that B[a]P significantly upregulated 54 EMT genes after 3 hours. By day 3, this number increased to 72 genes, with 36 genes being common between those elevated by SATB1. Notably, the significant effects of B[a]P on EMT gene expression observed in MCF10A cells were largely diminished in SATB1 expressing MCF10A cells, suggesting that the pre-existing SATB1-regulated transcriptional program is dominant. Our results indicate that, despite the distinct causes of breast cancer, both the chemical carcinogen B[a]P and SATB1 activate many common EMT genes during the malignant transformation of breast epithelial cells. This suggests a key network of gene expression that drives the malignant phenotypes of these cells. We investigated the changes in the transcriptional program of breast epithelial cells using the MCF10A-1 cell line by stably expressing SATB1 compared to control cells, or by exposing MCF10A-1 cells to benzo[a]pyrene (B[a]P) compared to non-treated controls. We focused on studying a particular group of genes that function in promoting epithelial-to-mesenchymal transition (EMT), which is essential during embryogenesis as well as during cancer metastasis, as it promotes cell motility. We compiled a gene set consisting of 300 EMT genes from recent literature and from gene ontology (GO) lists, and we studied which of these genes are activated or upregulated when MCF10A-1 cells undergo malignant transformation and acquire metastatic phenotypes. We employed a method called GRO-seq, which detects nascent transcripts genome-wide to capture the early transcriptional response to environmental stimuli, as well as RNA-seq, which measures accumulated levels of mRNAs.