Long-read sequencing-based atlas of tissue-specific expression of Drp1 transcript variants

Dynamin-related protein 1 (Drp1), encoded by DNM1L, is essential for mitochondrial fission, but its functional roles remain unclear due to isoform-specific effects from alternative splicing. Short-read RNA sequencing fails to resolve full-length isoforms involving distant exons, limiting our understanding. Here, we applied targeted long-read sequencing to profile full-length DNM1L transcripts in human left ventricle and iPSC-derived cardiomyocytes, recovering all annotated isoforms with conserved expression patterns and isoforms 1-4 being most abundant. Functional assays revealed that isoform abundance does not predict enzymatic activity. Extending this to six mouse tissues, we identified distinct, tissue-enriched expression profiles. Functional rescue in Drp1-knockout mouse embryonic fibroblasts showed isoform-dependent differences in mitochondrial fission. Isoforms lacking the A-insert (e.g., b and d) robustly rescued fission, while isoforms enriched in brain or muscle showed only partial rescue, suggesting exons 2 and 3 negatively regulate Drp1 activity. Our cross-species atlas integrates long-read transcriptomics with functional validation, revealing how isoform diversity underpins tissue-specific mitochondrial dynamics and physiological roles of Drp1.