mRNA splicing in midbody is essential for proper cell abscission in mitosis

Midbody is a transient structure assembled in the last step of mitosis and is currently conceptualized as a structural remnant subject to degradation following cytokinesis. However, our findings indicate that midbody regulation is more complicated than original thought. Knockdown spliceosome proteins in cells showed a multinuclear phenotype and abnormal midbody. Spliceosome and ribosome components are enriched in midbody. We further found that mRNA splicing and protein translation occurs in the midbody during last step of mitosis, which are necessary events for proper abscission of nascent daughter cells. Midbody spliced and translated PRC1 and NINL are essential for proper midbody function. Either PRC1 or NINL knockdown also showed a multinuclear phenotype and abnormal midbody. Treating cancer cells with both chemotherapy and spliceosome inhibitors dramatically lowers the concentration of both drugs, avoiding more severe side effects. Our study discovers a novel regulating mechanism for mitosis and provides a new strategy for cancer therapy. Overall design: two plates of cells were incubated with full culture medium containing 500µM biotin-phenol, while one plate of cells was supplied with 100nM PlaB until cells entered cytokinesis, which usually took approximately 40 minutes. Following this, 1mM H2O2 was added to all three plates of cells for 3 minutes. The reaction was stopped by replacing the medium with quenching buffer (10 mM sodium ascorbate, 10 mM sodium azide, and 5 mM Trolox in Dulbecco's Phosphate Buffered Saline [DPBS]). Cells were washed three times with quenching buffer and collected. Total RNAs were extracted using Trizol. For the enrichment of biotinylated RNAs, total RNAs were mixed with 20µL of Dynabeads MyOne Streptavidin Beads C1, rotating overnight at 4°C, and eluted as previously described. Eluted RNAs were purified using GeneJet RNA Concentration Kits (ThermoFisher) and the RNA concentration was determined using the Agilent 2100 Bioanalyzer.