Stage-Specific Expression and Perinuclear Enrichment of γ-Tubulin During Tomont Development and Theront Morphogenesis in Cryptocaryon irritans

Abstract

Cryptocaryon irritans is a lethal parasitic ciliate of marine fishes whose tomont stage is marked by repeated nuclear division and the morphogenesis of infective theronts, both dependent on dynamic microtubule reorganization. To investigate cytoskeleton-associated regulation during this proliferative process, we cloned and characterized the γ-tubulin gene (Ci-TUBG) and γ-tubulin complex protein 3 gene (Ci-TUBGCP3) from C. irritans. Sequence analysis showed that Ci-TUBG contains a canonical dual tubulin domain architecture and is highly conserved relative to homologs from Ichthyophthirius multifiliis and Tetrahymena thermophila. Structural modeling and molecular docking suggested a stable putative interaction interface between Ci-TUBG and Ci-TUBGCP3. Spatiotemporal expression analysis and immunofluorescence revealed stage-dependent upregulation and pronounced perinuclear enrichment of Ci-TUBG during tomont division, with persistent perinuclear localization in theronts. RNA interference showed that Ci-TUBG knockdown markedly reduced tomont hatching rate, decreased theront yield, and caused defects in theront morphogenesis accompanied by abnormal nuclear development. These findings indicate that Ci-TUBG is essential for tomont division and theront morphogenesis in C. irritans and provide a basis for understanding cytoskeleton-associated developmental regulation in this parasitic ciliate.