Genome-wide Characterization of Pm-Rel DNA-binding Landscape in Pearl Oyster Pinctada fucata martensii.

Pm-Rel, a key member of the NF-κB family, has been implicated in transplantation-induced immune responses in Pinctada fucata martensii; however, its genome-wide DNA-binding profile and associated regulatory features remain poorly characterized. Here, we generated a Pm-Rel-specific antibody and performed chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-seq) in pearl oyster hemocytes and identified 4,894 Pm-Rel-associated binding peaks and annotated 3,711 nearby genes. Functional enrichment analyses showed that these genes were significantly enriched in apoptosis and immune-associated pathways, including the tumor necrosis factor (TNF), apoptosis, and JAK-STAT signaling pathway. Motif analysis identified a highly enriched κB-like motif together with additional enriched sequence elements within Pm-Rel-associated regions. Representative sequences, including GGGGAATTCC, CTAAAAATAG, and AGTCGGAATT, were subsequently evaluated using dual-luciferase reporter assays. Combined expression of Pm-Rel and Pm-NF-κB activated reporter constructs containing the selected sequences, with the AGTCGGAATT-containing reporter showing the highest activity under the tested conditions. Collectively, this study provides a genome-wide characterization of Pm-Rel DNA-binding features in P. f. martensii, offering genomic insights into NF-κB-mediated immune and cellular regulatory processes in mollusks.