The Hox transcription factor Ultrabithorax binds RNA and regulates co-transcriptional splicing through an interplay with RNA polymerase II

ABSTRACT Transcription Factors (TFs) play a pivotal role in cell fate decision by coordinating distinct gene expression programs. Although most TFs act at the DNA regulatory layer, few TFs can bind RNA and modulate mRNA splicing. Yet, the mechanistic cues underlying TFs function in splicing remain elusive. Focusing on the Drosophila Hox TF Ultrabithorax (Ubx), our work shed light on a novel layer of Ubx function at the RNA level. Transcriptome and genome-wide binding profiles in embryonic mesoderm and Drosophila cells indicate that Ubx regulates mRNA expression and splicing to promote distinct functions in defined cellular contexts. Ubx modulates splicing via its DNA-binding domain, the Homeodomain (HD). Our results demonstrate a new RNA-binding ability of Ubx in cells and in vitro . Notably, the N51 amino acid of the HD, which mediates Ubx-DNA interaction, is non-essential for Ubx-RNA interaction in vitro but is required in vivo . We find that the N51 amino acid is necessary to mediate interaction between Ubx and the active form of the RNA Polymerase II (Pol II S2Phos) in Drosophila cells. By combining molecular and imaging approaches, our results reveal that Ubx mediates elongation-coupled splicing via a dynamic interplay with active Pol II and chromatin binding. Overall, our work uncovered a novel role of the Hox TFs at the mRNA regulatory layer. This could be an essential function for other classes of TFs to control cell diversity.