Intron location and sequence modulate gene expression in Yarrowia lipolytica

Introns are widespread among eukaryotic genomes. While intron-containing genes often show higher expression than genes lacking introns, the intron features influencing gene expression remain largely elusive. Here, we systematically characterize the intron landscape of Yarrowia lipolytica, an oleaginous yeast that is increasingly used as a microbial cell factory. Transcriptome analysis across 12 environments identified 2421 introns in 1430 genes, including 1302 newly discovered introns and 479 newly annotated intron-containing genes. We find that intron-containing genes exhibit higher and more stable expression across conditions and identify six key intron features, including the 5' splice motif, 3' splice motif, branch point motif, distance from branch point to 3' splice site, GC content, and intron size, that influence splicing efficiency and gene expression. A linear regression model based on these features robustly captures the intron's effect on gene expression, enabling us to select and test 55 different introns that modulate expression of a reporter gene by 200-fold. Moreover, we demonstrate that intron effects are robust across genomic contexts and identify a previously uncharacterized intron, I3, that strongly enhances gene expression and protein production. Together, our results provide new fundamental insights and open new avenues for using introns as regulatory elements.