RT Journal
A1 Choquet, Karine
A1 Chaumont, Louis-Philippe
A1 Bache, Simon
A1 Baxter-Koenigs, Autum R.
A1 Churchman, L. Stirling
T1 Genetic regulation of nascent RNA maturation revealed by direct RNA nanopore sequencing
JF Genome Research 
JO Genome Research 
YR 2025 
FD April 01 
VO 35 
IS 4 
SP 712 
OP 724 
DO 10.1101/gr.279203.124 
UL http://genome.cshlp.org/content/35/4/712.abstract 
AB Quantitative trait loci analyses have revealed an important role for genetic variants in regulating alternative splicing (AS) and alternative cleavage and polyadenylation (APA) in humans. Yet, these studies are generally performed with mature mRNA, so they report on the outcome rather than the processes of RNA maturation and thus may overlook how variants directly modulate pre-mRNA processing. The order in which the many introns of a human gene are removed can substantially influence AS, while nascent RNA polyadenylation can affect RNA stability and decay. However, how splicing order and poly(A) tail length are regulated by genetic variation has never been explored. Here, we used direct RNA nanopore sequencing to investigate allele-specific pre-mRNA maturation in 12 human lymphoblastoid cell lines. We find frequent splicing order differences between alleles and uncover significant single-nucleotide polymorphism (SNP)-splicing order associations in 17 genes. This includes SNPs located in or near splice sites as well as more distal intronic and exonic SNPs. Moreover, several genes showed allele-specific poly(A) tail lengths, many of which also have a skewed allelic abundance ratio. HLA class I transcripts, which encode proteins that play an essential role in antigen presentation, show the most allele-specific splicing orders, which frequently co-occur with allele-specific AS, APA, or poly(A) tail length differences. Together, our results expose new layers of genetic regulation of pre-mRNA maturation and highlight the power of long-read RNA sequencing for allele-specific analyses.