PROJECT SUMMARY Currently there are no effective treatments that can cure metastatic breast cancer. The epithelial-to- mesenchymal transition (EMT) is a critical cancer cell plasticity and dedifferentiation program by which epithelial cells acquire pro-migratory and invasive mesenchymal properties. To initiate the EMT program, cancer cells need to receive pro-EMT signals, such as TGF from either neighboring tumor cells or from the microenvironment (surrounding stroma). Our research group made the unexpected finding that during TGFβ mediated EMT, Pol I rDNA transcription is: (1) driven by Snail, a transcription factor known to play a central role in orchestrating the mesenchymal Pol II dependent gene expression program required for cellular invasiveness and metastatic spread; (2) a concurrent loss of Myc occurs at rDNA genes; and (3) Snail forms a complex with METTL5 to enable that the newly made rRNA are m6A modified by METTL5 before the rRNA is incorporated into the mature ribosome; (4) Snail also forms a complex with METTL3 and may therefore direct m6A modifications of selective mRNAs; (5) the m6A marked mRNAs enables recruitment of the non-canonical DAP5/eIF3d/METTL3 translation complex to drive selective m6A marked mRNA translation in the mesenchymal state; and (6) METTL5 is required for execution of the EMT program, as silencing of METTL5 prevents cells from undergoing EMT. The focus of this grant application is to gain a deeper molecular understanding of Pol I rDNA transcription, METTL5 m6A modified rRNA and ribosomes and why selective DAP5/eIF3d mediated translational control is essential for orchestrating breast cancer cell plasticity in EMT, its role in breast cancer metastatic progression, and how Snail orchestrates and coordinates the m6A rRNA/mRNA axis. This knowledge will ultimately inform how targeting Pol I machinery, specialized ribosomes and translation control could represent a novel therapy specifically targeting the plastic, non-proliferating and chemo-resistant EMT cells fueling tumor reoccurrence and metastasis.