PROJECT SUMMARY Multiple Myeloma (MM) is an incurable cancer of differentiated B lymphocytes (or “plasma cells”) which generally presents late in life and has a median life expectancy of ~4-7 years from time of diagnosis. MM cells are unusual among most other cell lineages, including other types of cancer, in that they synthesize large amounts of a single, nonfunctional protein. Despite this unique signature, the regulation of the protein synthesis machinery of the cell, the ribosome, remains poorly studied in MM. Recent evidence has advanced our understanding of a stress response cascade triggered downstream of the ribosome, which ranges from ribosome-mediated quality control (RQC) of single mRNAs to a “ribotoxic” stress response and global translational shutdown when fully activated. Importantly, preliminary data demonstrate that among 375 cancer cell lines, MM cells are among those that express the highest baseline levels of an upstream factor in this cascade, EDF1. Together, this suggests that RQC and the ribotoxic stress response may be generally activated at baseline in MM cells compared to other cell types. Beyond RQC and the ribotoxic stress response, MM cells are also differentially sensitive to inhibition of specific components the translational machinery of the cell, including all three subunits of the eukaryotic initiation factor 4F complex. Literature evidence and preliminary data suggest that MM cells may be hyper-dependent on the eIF4F complex to translate specific subsets of mRNAs which promote their proliferation or survival, but the identity of these mRNAs and mechanistic insight into why they are particularly dependent on eIF4F levels, both generally and in MM, is unclear. In this proposal, I seek to connect these ideas in order to understand how MM cells modulate the translational machinery to meet their unique protein synthesis demands. The central hypothesis of this proposal is that translation is broadly dysregulated in MM to allow high rates of protein synthesis and the translation of specific mRNAs which promote survival and proliferation. I propose to address this hypothesis through the following specific Aims: Aim 1: Characterize translation, quality control, and the ribotoxic stress response in MM; Aim 2: Define the eIF4F-sensitive translational landscape in MM; Aim 3: polysome-associated CAGE-seq (paCAGE) to interrogate translational control by 5'UTRs in MM. These aims will be achieved through a combination of biochemical and sequencing approaches in immortalized MM cell lines, including the development of a new technology which will be essential to identify 5'UTR sequence motifs within eIF4F-sensitive mRNAs. This work will be significant because it may reveal new therapeutic targets in multiple myeloma while elucidating important ribosomal biology which may be more generally involved in other model systems or disease states.