Alternative mRNA splicing controls functions of the human H3K27 demethylase UTX/KDM6A

The UTX/KDM6A gene encodes the UTX histone H3K27 demethylase, which plays an important role in mammalian development and is frequently mutated in cancers and particularly, in urothelial cancers. Despite many studies on UTX proteins, variations in mRNA splicing have been overlooked. Here, we employ long-read Nanopore sequencing to present a comprehensive analysis of UTX/KDM6A splicing events in different human cell lines and in tissue samples from both bladder cancer and normal bladder epithelium. We found that the central region of UTX mRNAs encoded by exons 12 to 17 undergoes extensive alternative splicing. Upto half of all stable mRNAs (8-48% in bladder tissues and 18-58% in cell lines) are represented by the UTX canonical isoform lacking exon 14 encoding a nuclear localization signal. We found that exon 14-containing isoforms exclusively localize to the nucleus, unlike cytonuclear localization of the canonical isoform. Chromatin association was also higher for exon 14-containing isoform compared to the canonical UTX. In both cases, UTX localized mainly to the active regions of the genome. Using quantitative mass spectrometry, we found that all UTX isoforms integrated into the MLL3 and MLL4 complexes. Sub-stoichiometric interactions were observed for the PR-DUB histone H2A deubiquitination and MiDAC histone deacetylase complexes. Interestingly, one of the novel UTX isoforms, which lacks exons 14 and 16, fails to interact with PR-DUB and MiDAC complex members. In conclusion, our study shows that UTX mRNAs undergo extensive alternative splicing events that control the subcellular localization of UTX proteins and their interactions with other chromatin regulatory complexes.