Identification and characterization of proteins involved in stolon adhesion in the highly invasive fouling ascidian Ciona robusta.

Adhesive ascidians have caused serious biofouling problems and huge economic losses in marine ecosystems. However, adhesion mechanisms, particularly on functional proteins involved in ascidian adhesion, remain largely unexplored. Here, we identified 26 representative stolon proteins from the highly invasive fouling ascidian Ciona robusta using the proteomics approach. The uncharacterized stolon proteins were rich in adhesion-related conserved domains. Real-time quantitative PCR further revealed specific expressions of these uncharacterized protein genes in stolon tissue, suggesting their potential roles in stolon adhesion.> A recombinant vWFA domain-containing uncharacterized protein, ascidian stolon protein 1 (ASP-1), was successfully expressed in a baculovirus-insect cell system and purified in vitro. Coating experiment showed that tyrosinase-modified ASP-1 could absorb to glass and organic glass stronger than unmodified ASP-1, while only modified ASP-1 could absorb to aluminum foil. Quartz crystal microbalance analysis also showed the increase in absorption ability of ASP-1 after modification. In addition, abundant 3,4-l-dihydroxyphenylalanine (DOPA) in modified protein was detected by nitroblue tetrazolium staining. These results suggest that ASP-1 be involved in ascidian DOPA-dependent and material-selective adhesion. Overall, this study provides insight into molecular mechanisms of C. robusta stolon adhesion, and findings here are expected to be conductive to develop strategies against biofouling caused by ascidians.