Amorphous calcium carbonate (ACC) is a precursor phase of calcite in the formation of the sea urchin larval spicule. The goal of this research is to study the formation and stabilization mode of this transient phase. We first characterized the mineralogy of the spicules from the sea urchin Strongylocentrotus purpuratus. We then examined the role of the macromolecules extracted from the spicules at different growth stages in the formation of transient ACC in vitro.The biogenic amorphous transient phase is shown to be both structurally and compositionally different from the known stable ACC phases. It does not contain bound water, and is thus the first dehydrated ACC phase to be detected. The macromolecules that were extracted at early stages of spicule growth, when the amorphous content of the biogenic mineral is high, induced the formation of transient ACC in vitro in the presence of magnesium ions. In contrast, the macromolecules extracted at a later stage, when the spicules are completely crystalline, induced the formation of single crystals of low magnesian calcite. We therefore deduce that the macromolecules from the sea urchin larval spicules together with magnesium ions, mediate the transient formation of ACC as a precursor to calcite. These observations may well provide novel ideas for improved materials synthesis.

During its embryonic development, the sea urchin embryo forms an endoskeletal calcitic spicule. This instance of biomineralization is experimentally accessible and also offers the advantage of occurring within a developmental context. Here we investigate the time course of appearance and localization of two proteins among the four dozen that constitute the protein matrix of the skeletal spicule. SM50 and SM30 have been studied in some detail, and polyclonal antisera have been prepared against them (C. E. Killian and F. H. Wilt, 1996, J. Biol. Chem. 271, 9150-9159). Using these antibodies we describe here the localization and time course of accumulation of these two proteins in Strongylocentrotus purpuratus, both in the intact embryo and in micromere cultures. We also investigate the disposition of the matrix proteins, SM50, SM30, and PM27, in the three-dimensional spicule by studying changes in protein localization during experimental manipulation of isolated skeletal spicules. We conclude that SM50, PM27, and SM30 probably play different roles in biomineralization, based on their localization and patterns of expression. It is unlikely that these proteins are solely structural elements within the mineral. SM50 and PM27 may play a role in defining the extracellular space in which spicule deposition occurs, while SM30 may play a role in secretion of spicule components. Finally, we report on the effects of serum on expression of some primary mesenchyme-specific proteins in micromere cultures; withholding serum severely depresses accumulation of SM30 but has only modest effects on the accumulation of other proteins.

No abstract

Skeleton formation involves secretion of massive amounts of mineral precursor, usually a calcium salt, and matrix proteins, many of which are deposited on, or even occluded within, the mineral. The cell biological underpinnings of this secretion and subsequent assembly of the biomineralized skeletal element is not well understood. We ask here what is the relationship of the trafficking and secretion of the mineral and matrix within the primary mesenchyme cells of the sea urchin embryo, cells that deposit the endoskeletal spicule. Fluorescent labeling of intracellular calcium deposits show mineral precursors are present in granules visible by light microscopy, from whence they are deposited in the endoskeletal spicule, especially at its tip. In contrast, two different matrix proteins tagged with GFP are present in smaller post-Golgi vesicles only seen by electron microscopy, and the secreted protein are only incorporated into the spicule in the vicinity of the cell of origin. The matrix protein, SpSM30B, is post-translationally modified during secretion, and this processing continues after its incorporation into the spicule. Our findings also indicate that the mineral precursor and two well characterized matrix proteins are trafficked by different cellular routes.

No abstract