DDX53 binds a wide range of RNA molecules in human TCam-2 cells (eCLIP data)

DDX53 (DEAD-box helicase 53, known also as CAGE) is an intronless gene on the X chromosome, which expression shows strong testis and brain specificity. Additionally, it belongs to the group of cancer-testis (CT) antigens, with most studies to date focusing on its role in cancer. Previous reports of rare variants within the DDX53 in infertile men have provided a rationale for our study of DDX53 in the context of human spermatogenesis. By using the human seminoma cell line (TCam-2) as an in vitro male germline model, we aimed to investigate the function and molecular targets of DDX53 protein. Our eCLIP and RNA-seq data show that DDX53 protein binds a wide repertoire of RNA targets, drives transcriptome changes in human cells, and is involved in alternative splicing of RNA transcripts. Further, we examined the localization of DDX53 using confocal microscopy, revealing that it is mainly located in the cytoplasm, but also in the nucleus of TCam-2 cells. Our data indicate that DDX53 protein acts as a regulator of RNA metabolism in human cells. Collectively, we show that DDX53 participates in transcriptome regulation (including splicing) in male germ cells and exhibits transcriptome-wide RNA interactions, but its wider biological role remains to be clarified. Overall design: Two biological replicates of TCam-2 cells expressing the DDX53-FLAG were used in eCLIP-seq performed by Eclipse BioInnovations (USA). Cells were crosslinked, then lysed, RNase I treated, and protein-RNA complexes were immunoprecipitated with anti-FLAG antibody coupled to magnetic beads. The eCLIP experiment was performed by Eclipse BioInnovations' standard RBP-eCLIP pipeline.