Monday April 3rd, 4pm CEST

Anne Jantschke^*, Iddo Pinkas⁺, Andreas Schertel⁺, Prof. Lia Addadi⁺ and Prof. Steve Weiner⁺

Biomineralization pathways in calcifying dinoflagellates: uptake, storage in MgCaP rich bodies and formation of the shell

^*Biomineralization and Crystallography, Institute for Geosciences, JGU Mainz, Germany
⁺Structural Biology, Weizmann Institute of Science, Rehovot, Israel

During the last decades, significant progress has been made in understanding biomineral formation in model microalgae like diatoms and coccolithophores. In contrast, the mechanisms that control the intricate mineral construction in calcareous dinoflagellates are practically unknown. Our main objectives are to gain insight into their mineral architecture, subcellular structures that may play a role in biomineralization, and calcite morphogenesis. Two representative members of calcareous dinoflagellates were investigated using cryo-electron microscopy (cryo SEM and cryo FIB SEM) in combination with various spectroscopic techniques (FT-IR, Raman, Fluorescence, EDS) enabling an investigation of cells as close as possible to the natural state.
Calcite formation occurs via multiple independent nucleation events inside the so-called outer matrix. Based on 2D and 3D cryo-electron microscopic datasets it is shown that individual calcite crystals grow with preferred orientation into a dense reticular network resulting in a highly regular, porous calcite shell.
In both species, vacuoles containing crystalline inclusions were observed using cryo-SEM. So far, crystalline deposits were assigned to calcite assuming that they are involved in shell formation. Surprisingly, using in situ Raman spectroscopic imaging these crystalline inclusions could be identified as anhydrous guanine in the biogenic β-form using their low-wavenumber Raman signature.
Interestingly, live-cell fluorescence imaging with Calcein-AM, cryo-sectioning and cryo-EDS show the presence of small MgCaP-rich mineral bodies within the same vacuolar enclosures. 3D cryo-FIB-SEM imaging of a calcifying cell shows a remarkably large number (353) of these bodies distributed in the cell volume. Out of these bodies, 52 (~15%) are located between the two inner organic layers of the outer matrix. We suggest that these MgCaP-rich bodies are being secreted into the outer matrix and are part of a Ca-concentrating or transport mechanism.
Based on our results a new dinoflagellate biomineralization model was developed which includes the active uptake of Ca²⁺, temporary deposition in MgCaP-precursor bodies, extrusion into the outer matrix and transformation into low Mg-calcite.