Silaffins-Driven Genetic Engineering of Diatom Cell Walls: Insight into Biosilica Morphology and Nanomaterial Design

Abstract

AbstractDiatoms synthesize silica cell walls (frustules) with genetically encoded morphologies, ranging from nanopatterns to micropatterns, that far exceed current synthetic chemistry. Silaffins, a family of phosphoproteins undergoing complex post-translational modifications, have been isolated from frustules and shown to facilitate and regulate biosilica formationin vitrowith long-chain polyamines. However, their particular role in frustule morphogenesis and functionality remains unclear. In this study, functions of two representativesilaffins,TpSil1andTpSil3, were investigated in the model organismThalassiosira pseudonanausing gene overexpression and CRISPR/Cas9-mediated knockout approaches. Due to high sequence homology,TpSil2was concurrently disrupted inTpSil1knockout strains, while the homozygous knockout ofTpSil3proved to be lethal. Quantitative morphological analysis revealed distinct yet complementary roles: TpSil3 governs both microscale overall size and mesoscale features, including macropore (fultoportula) density and mesopore (cribrum pore) pattern, whereas TpSil1/2 exclusively contribute to macropore morphogenesis and mesopore density. Overexpression ofsilaffinsincreased silica deposition, while knockouts exhibited reduced silicification but enhanced cell growth and photosynthetic efficiency. Furthermore, these genetic modifications significantly influenced the physicochemical and optical properties of bulk frustules, potentially enhancing the hemostatic, catalytic and photonic performances, thereby positioning them as versatile candidates for a wide range of biotechnological and industrial applications. Collectively, our findings elucidate the distinct roles ofTpSil1/2andTpSil3in diatom physiology and frustule morphology, highlighting a promising pathway for engineering nanostructured silica materials with tailored properties through synthetic biology.