Juvenile-specific expression of a phospholipase A2 isozyme gene in the venom gland of Protobothrops flavoviridis.

Venom composition and gene expression in the habu snake Protobothrops flavoviridis (P. flavoviridis), native to the southwestern islands of Japan, vary by habitat and may reflect regional dietary differences. Because juveniles and adults feed on different prey, we hypothesized that venom composition undergoes age-dependent changes during growth. To test this hypothesis, we compared venom proteins from juvenile and adult snakes by two-dimensional gel electrophoresis (2-DE) and identified a protein spot uniquely present in juvenile venom. Mass spectrometry revealed that this protein is encoded by the phospholipase A2 (PLA2) gene pgPLA1b/2b, previously regarded as a pseudogene because its product is absent from adult venom. Reverse transcription-PCR (RT-PCR) analysis of venom glands from snakes of different ages showed that pgPLA1b/2b is expressed in 0- and 1-year-old individuals, barely detectable at age 2, and absent in adults, whereas other PLA2 isozyme genes remain constitutively expressed regardless of age. These findings demonstrate that pgPLA1b/2b is the only venom PLA2 isozyme gene exhibiting ontogenetic regulation. Since PLA2 isozyme genes form gene clusters in the P. flavoviridis genome, the age-dependent expression of only pgPLA1b/2b is of particular interest. Our study provides molecular evidence that venom composition in P. flavoviridis is developmentally regulated through juvenile-specific expression of a venom gene, likely reflecting ecological and dietary transitions during growth. Future studies should elucidate the transcriptional mechanisms underlying this regulation.