RNA-sequencing of isogenic primary, pre-malignant immortalized, and Ras-transformed human mammary epithelial cells

Understanding changes in gene expression during tumor initiation and progression is critical to understanding how genetic alterations drive malignancy. We used a genetically defined cell culture model to study the progression of normal human mammary epithelial cells (HMECs) to malignancy. Primary HMECs were immortalized through the expression of hTERT, p53DD, cyclin D1, CDK4R24C and c-MYCT58A. This immortalization conferred limitless replicative potential as well as migratory capacity. These pre-malignant cells were subsequently HRASG12V transformed, which converted the immortalized cells to a fully tumorigenic state with significantly increased invasive capacity. We analyzed the cells using RNA-sequencing, and we report dramatic mRNA expression changes during the pre-malignant immortalization of primary cells, and very few mRNA expression changes occurring during oncogenic Ras transformation. RNA signatures in pre-malignant immortalized and Ras-transformed cells are consistent with previously reported epithelial-to-mesenchymal transition (EMT) signatures. RNA-sequencing in biological triplicate of three cell types (Primary HMECs, Immortalized HMECs, Transformed HMECs); Illumina HiSeq 2000 125bp PE (1 replicate) and Illumina HiSeq 4000 150bp PE (2 replicates)