Long-read single-cell RNA sequencing uncovers cell-type specific transcript regulation in COVID-19

SARS-CoV-2 infection leads to extensive host transcriptomic changes, but the role of alternative splicing in shaping the immune response remains underexplored. Here, we present the first application of long-read single-cell RNA sequencing on nasopharyngeal swabs from COVID-19 patients and healthy controls to resolve transcript-level changes across cell types. Our analysis identified major epithelial cell types and pronounced immune infiltration, with cell-type annotations concordant with those from short-read data. By enabling isoform-level resolution, our nanopore sequencing approach revealed cell-type specific alternative splicing, undetectable with short-read sequencing. For example, although gene-level expression of the key immune and apoptosis regulators, IFNAR2 and FAIM, did not differ between COVID-19 patients and healthy controls we identified marked shifts in isoform usage. Between moderate and critical cases, we observed cell-type specific differential transcript usage in the T cell signaling kinase FYN and the immune-regulatory transcription factor IRF2. As some of these splicing alterations yield functionally distinct isoforms, we hypothesize that alternative splicing modulates immune signaling and apoptosis, fine-tuning the host response to SARS-CoV-2 infection. Our study demonstrates the unique power of long-read single-cell transcriptomics to uncover isoform-resolved regulatory changes, offering novel insights into the role of alternative splicing in shaping immune responses to viral infections.