Grammatikakis, I. | bioRxiv Science

bioRxiv2022

Matrin3 regulates cell proliferation and spindle dynamics by regulating CDC14B alternative splicing

Matrin3 is an RNA-binding protein that affects diverse RNA-related processes, including mRNA splicing. While Matrin3 has been intensively studied in neurodegenerative diseases, its function in cancer remains unclear. Here, we discovered Matrin3-mediated regulation of mitotic spindle dynamics in colorectal cancer (CRC) cells. We identified bound and regulated Matrin3-target RNAs transcriptome-wide in CRC cells and found that Matrin3 broadly modulates mRNA splicing patterns. Among the top Matrin3 targets, we focused on CDC14B and found that Matrin3 loss increased inclusion of an exon containing a premature termination codon into the CDC14B transcript and simultaneous down-regulation of the standard CDC14B transcript. Selective knockdown of the CDC14B standard transcript phenocopied Matrin3 knockdown and exhibited reduced proliferation and defects in mitotic spindle formation, suggesting that CDC14B is a key downstream effector of Matrin3. Collectively, these data reveal an important role for the Matrin3/CDC14B axis in control of CRC cell proliferation and mitotic spindle formation.

molecular biology↗

bioRxiv2022

HNRNPH1 destabilizes the G-quadruplex structures formed by G-rich RNA sequences that regulate the alternative splicing of an oncogenic fusion transcript

In the presence of physiological monovalent cations, thousands of RNA G-rich sequences can form parallel G-quadruplexes (G4s) unless RNA-binding proteins inhibit, destabilize, or resolve the formation of such secondary RNA structures. Here, we have used a disease-relevant model system to investigate the biophysical properties of the RNA-binding protein HNRNPH1s interaction with G-rich sequences. We demonstrate the importance of two EWSR1-exon 8 G-rich regions in mediating the exclusion of this exon from the oncogenic EWS-FLI1 transcripts expressed in a subset of Ewing sarcomas, using complementary analysis of tumor data, long-read sequencing, and minigene studies. We determined that HNRNPH1 binds the EWSR1-exon 8 G-rich sequences with low nM affinities irrespective of whether in a non-G4 or G4 state but exhibits different kinetics depending on RNA structure. Specifically, HNRNPH1 associates and dissociates from G4-folded RNA faster than the identical sequences in a non-G4 state. Importantly, we demonstrate using gel shift and spectroscopic assays that HNRNPH1, particularly the qRRM1-qRRM2 domains, destabilizes the G4s formed by the EWSR1-exon 8 G-rich sequences in a non-catalytic fashion. Our results indicate that HNRNPH1s binding of G-rich sequences favors the accumulation of RNA in a non-G4 state and that this contributes to its regulation of RNA processing.

molecular biology↗