Minimal edit, maximal effect: extended survival SCID pigs with single amino acid deletion [RNA-Seq]
Severe combined immunodeficiency (SCID) represents a critical area of research for understanding human immune disorders and developing therapeutic interventions. While murine models have provided valuable insights, their translational relevance is limited by physiological differences from humans. Here, we established and characterized a novel RAG2 biallelic knock out (RAG2bKO) model using TALEN-mediated gene editing, which demonstrates extended viability in specific pathogen-reduced facilities while maintaining complete immunodeficiency. Our model exhibits a precise in-frame deletion of a single histidine residue (p.His109del) in the RAG2 protein, resulting in structural destabilization and functional impairment. Comprehensive phenotypic analysis revealed complete thymic aplasia, splenic hypoplasia, and ablation of mature T and B lymphocytes. RNA-seq and H3K4me3 ChIP-seq approaches, including transcriptomic and epigenomic analyses, demonstrated conserved dysregulation of immune pathways across lymphoid tissues, with significant suppression of adaptive immune pathways and compensatory upregulation of innate immune mechanisms. The RAG2bKO pigs showed extended survival up to one year in controlled environments, representing a significant improvement over existing models. This large animal model provides a robust platform for studying SCID pathogenesis, evaluating long-term therapeutic interventions, Overall design: To investigate the transcriptomic consequences of RAG2 deficiency, we performed RNA sequencing on lymphoid tissues (lymph node, spleen and thymus) from RAG2 biallelic knockout (RAG2bKO) pigs and their wild-type (WT) littermates.