rRNA modification, ribosome composition and specialized translation in glioblastoma. - ABSTRACT Glioblastoma (GBM) is the most common and most aggressive brain tumor type among adults, with a 14- month average patient survival. Despite all genomic efforts in recent years, novel therapies based on common GBM mutations and alterations have been disappointing. This proposal focuses on Ribosome biogenesis, a multi-step process that includes rRNA transcription, processing and modification, generation, and assembly of all 80S ribosome components. In cancer cells, quantitative and qualitative differences in ribosome biogenesis components and regulators ultimately contribute to changes in translation that enhance oncogenic processes. Targeting ribosome biogenesis has emerged as a promising therapeutic option, especially in highly aggressive tumors that demand increased and specialized protein production, such as GBM. To effectively use ribosome biogenesis inhibitors in cancer therapy, we must first identify differences in rRNA transcription, processing, and modification between normal and cancer cells and assess the contribution of different ribosome biogenesis components to tumor development and therapy response. Aim 1. snoRNAs are the main players in rRNA modification, and they function by guiding snoRNPs to specific positions in rRNA via base pairing. We used a dedicated RNAseq platform to investigate differences in snoRNA expression between glioma stem cells (GSCs), GBM, Astrocytes, Neuronal Precursor Cells (NPCs), and brain. We established that GSCs display a unique snoRNA profile. We propose that such differences in snoRNA expression create a distinct pattern of rRNA modification that influences ribosome composition in GSCs. We will isolate active ribosomes from glioma stem cells (GSCs), astrocytes, and neuronal precursor cells (NPCs) and employ a specialized genomic method (Nm-Mut-seq) to identify differences in their rRNA 2'O methylation profiles and Mass Spectroscopy to determine stochiometric variations in their ribosome composition. Next, we will perform a comparative proteomics study to measure protein synthesis in GSCs vs. astrocytes and NPCs. Aim 2. Poly(ADP-ribosyl)ation (PARylation) contributes to the integrity of protein complexes implicated in ribosome biogenesis. We identified the RNA binding protein SERBP1 as a novel oncogenic factor in GBM. Our recent genomics and proteomics studies established that SERBP1 interacts preferentially with rRNA transcription/modification regulators and PARP1, the main factor in protein PARylation. We showed that SERBP1 promotes PARylation and associates preferentially with PARylated proteins and PAR-binders. We propose that SERBP1 functions as a novel player in ribosome biogenesis in glioblastoma. We will determine if SERBP1 influences ribosome composition in GSCs by impacting rRNA modification and the activity of factors implicated in ribosome biogenesis. Next, we will determine the value of combining SERBP1 targeting and ribosome biogenesis inhibition as a therapeutic strategy.
