Non-coding RNAs in oncogene-induced biomolecular condensates

Most cellular biochemical processes occur in confined compartments. This type of spatial organization allows the molecular composition of the reaction chambers to be optimized such that very specific biochemical reactions can proceed in a highly efficient fashion. Many subcellular compartments are surrounded by membranes. In contrast, so-called "membrane-less organelles" can form through the biomolecular condensation of proteins and nucleic acids with specific biochemical and biophysical properties. For example, proteins with disordered regions that lack a clearly defined three-dimensional structure are often part of biomolecular condensates. However, it is not well understood how long RNA molecules with regulatory functions that do not code for proteins contribute to the formation of biomolecular condensates. Cancer is often triggered by fusion proteins that arise from mutation-induced chromosomal rearrangements. Because fusion proteins are often found in biomolecular condensates, we hypothesize that there are cancer-specific biomolecular condensates. While these structures are not expected to be found in normal cells, they should be important for cancer cell growth. Furthermore, long, non-protein-coding RNAs can be specifically produced in cancer cells. However, it is not known whether and how these cancer-specific regulatory RNAs regulate the formation, stability and composition of oncogenic biomolecular condensates. We will investigate the role of cancer-specific regulatory RNAs in the context of oncogenic biomolecular condensates. We will use bioinformatic analysis to identify cancer-specific non-protein coding RNAs. Using the CRISPR/Cas9 technology to visualize fusion proteins together with modern microscopy, we will investigate whether these RNAs interact with oncogenic fusion proteins in the context of biomolecular condensates in cancer cells. We will also investigate how the loss of cancer- specific regulatory RNAs affects the formation and stability of oncogenic biomolecular condensates, and how the resulting cancer-specific regulatory processes are affected. To this end, we will inactivate selected RNAs using the CRISPR/Cas9 technology and study the effects using microscopy and high-throughput sequencing. The data will improve our understanding of the poorly characterized concept of oncogenic biomolecular condensation and the role of regulatory RNAs in this context. The results will contribute to the development of strategies for targeted inactivation of oncogenic biomolecular condensates.

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