Characterization of novel RNA-binding proteins in cancer

RNA-binding proteins (RBPs) are key players in the post-transcriptional regulation of gene expression, regulating each step of RNA metabolism, from synthesis to decay through a dynamic association. Accordingly, the repertoire of new non-canonical RBPs has consistently grown in the last few years. Given that post-transcriptional events play pivotal roles in the adaption of cells to the local microenvironment, it is common that perturbations of RBP-networks can lead to cancer through mechanisms that are still poorly understood. In this context, we investigated the role played by TRAP1, a molecular chaperone whose role in cancer has been extensively described, and its predicted interacting-partner Protein Syndesmos (SDOS). SDOS, also known as Nudt16l1, is a paralog of the catalytic nuclear Nudt16p family of proteins that has been predicted to lack the decapping activity. This work demonstrates that SDOS interacts with TRAP1, as shown by co-immunoprecipitation and proximity ligation assays. Moreover, SDOS associates with actively translating polyribosomes and takes part to stress granules, being involved in the downmodulation of mRNA translation. By both polynucleotide kinase (PNK) assay and small-scale RNA interactome capture, we demonstrated, for the first time, that SDOS and TRAP1 are novel, non-canonical RBPs. Consequently, we have characterized the RNA-binding properties of SDOS and TRAP1, by combining three high-throughput approaches: i) individual nucleotide cross-linking and immunoprecipitation (iCLIP) sequencing, to identify direct RNA targets; ii) Ribosome profiling sequencing, to identify differentially translated targets and iii) Gene expression analysis, to identify differentially expressed genes. Combination of these analyses allowed us to identify several crucial regulated pathways and, among those, we focused our attention on a small subset of genes responsible for ciliopathies, a class of rare diseases caused by defects in primary cilia. Among them we confirmed TMEM107, a ciliary transition zone protein, as directly bound at RNA level by SDOS, as demonstrated by RNA-immunoprecipitation analysis. Moreover, TMEM107 translational regulation by SDOS was demonstrated by western blot and qPCR assays. Taken together these findings suggest that SDOS might regulate primary cilia formation. Intriguingly, a new area of research is emerging linking cilia to cancer, suggesting the existence of a bridge between SDOS and TRAP1 functions and related diseases.