<p dir="ltr">Hepatocellular carcinoma (HCC) is one of the most prevalent cancers in the world. In recent years, RNA modifications, such as N6-methyladenosine (m6A), has emerged as important players in cancer progression. They have been reported to regulate various RNA processes, such as splicing, nuclear export, mRNA stability and translation. However, the role of many RNA modifications in HCC development has remained largely unknown. Pseudouridine, an isomer of uridine, is an abundant RNA modifcation, which is present in mRNA, rRNA, tRNA and other non-coding RNA. In humans, pseudouridylation is catalyzed by 13 pseudouridine synthases (PUSs). In this study, we employed bisulfite-induced deletion sequencing (BID-seq), a novel sequencing technique allowing transcriptome-wide detection of pseudouridine at single-base resolution, to investigate the landscape of RNA pseudouridylation in HCC. Our integrated analysis has revealed the dysregulations and clinical significance of pseudouridine in HCC. Using a multi-omics approach, we also investigated the role of pseudouridine in mRNA translation. The dataset includes 1) numerical data for plotting figures in the study, 2) omics data in the study and 3) Western blot and cell colony images in the study.</p>