HNRNPH1-mediated splicing events regulate transcript variant composition and the organization of the 5'UTR.

HNRNPH1 is a regulator of alternative splicing, but few studies have defined the splicing events it mediates. Here, we used short- and long-read RNA sequencing to interrogate the transcriptome-wide effects of HNRNPH1 depletion and its regulation of specific splicing events. Differential alternative splicing analysis revealed effects on the transcriptome that involved all splice event categories. We confirmed HNRNPH1's regulation of a splicing event involving -exons 18a and 18b that encode distinct TCF3 transcription factor isoforms. Extending this finding, we present evidence that in neuroblastoma, is a MYCN target, potentially explaining the higher levels of and exon 18a transcript variants in this tumor type. Analysis of two skipped exon events determined that HNRNPH1 regulates the splicing of exons encoding part of the EIF4G1 translation initiation factor's N-terminus and an exon included in the 5'UTR of specific transcript variants encoding the mitotic kinase AURKA. Using reporter constructs, we show this 5'UTR exon enhances expression, suggesting HNRNPH1 could contribute to regulating AURKA protein levels. Our findings highlight HNRNPH1's roles in regulating the expression of proteins with diverse cellular functions.