Recently, diatom biosilica-associated insoluble organic matrices have been discovered that exhibit characteristic nanopatterns and are believed to be key components of the cellular machinery for silica morphogenesis. Elucidating the functions of these organic matrices requires a detailed understanding of their biomolecular compositions and the mechanism of their assembly. In this project we aim to gain such an understanding using the cingulin containing organic microrings from T. pseudonana as a model system (see Figure). Using biochemical and molecular genetic tools (including RNAi) three key questions will be addressed: (i) What are the similarities and differences in protein composition of silica forming insoluble organic matrices from different diatom species? (ii) How does the binding of soluble components affect the silica forming activities of the organic microrings? (iii) What is the in vivo role of cingulins regarding assembly of the organic microrings, and morphogenesis of the girdle band silica? The research in this project is expected to identify novel generic and species specific components of the insoluble organic matrices, and provide first insight into the mechanism of their assembly. Through thea in vivo incorporation of fluorescently tagged matrix proteins, and subsequent analysis by high-resolution fluorescence microscopy (in collaboration with SP5), this project will also provide the basis for obtaining a detailed picture of the structure of the organic microrings with nearly single molecule resolution. The results from these experimental approaches are expected to reveal the role that the insoluble organic microrings and their interactions with soluble biomineralization components play in the morphogenesis of diatom silica.