Ultrastructural localization of proteins involved in sea urchin biomineralization.

Three skeletal tissues of the adult echinoid Paracentrotus lividus (the pedicellaria primordium, the test, and the tooth) were immunolabeled with three sera raised against the total mineralization organic matrix and two specific matrix proteins (SM30 and SM50) from the embryo of the echinoid Strongylocentrotus purpuratus. Two conventional chemical fixation protocols and two high-pressure freezing/freeze-substitution protocols were tested. One conventional protocol is recommended for its good preservation of the ultrastructure, and one high-pressure freezing/freeze-substitution protocol is recommended for its good retention of antigenicity. Immunolabeling was obtained in the three adult tissues. It was confined to the active skeleton-forming cells and to the structured organic matrix. The results indicate that the matrix proteins follow the classical routes of secretory protein assembly and export and suggest that SM30 and SM50 are a part of the tridimensional network formed by the organic matrix before the onset of mineralization. They show that the genetic program of part of skeletogenesis is conserved among different calcification models and developmental stages.     

Immunoelectron microscopic studies of glycosaminoglycans in the metaphyseal bone trabeculae of growing rats

Summary The types and distribution of glycosaminoglycans (GAGs) were studied immunocytochemically in osteoid, mineralized bone matrix, and cartilage matrix of growing rat metaphyseal bone after aldehyde fixation and EDTA demineralization, using four monoclonal antibodies (mAbs 1-B-5, 2-B-6, 3-B-3 and 5-D-4). These mAbs specifically recognize epitopes in non-sulphated chondroitin (C0-S); chondroitin 4-sulphate (C4-S) and dermatan sulphate (DS); chondroitin 6-sulphate (C6-S) and C0-S; and keratan sulphate (KS) respectively. In osteoid, all mAbs except 1-B-5 weakly stained matrix material on and between collagen fibrils, and moderately stained organic material corresponding to bone nodules, which are known sites of mineralization. However, the staining of osteoid abruptly decreased at the mineralization front; weak staining was confined mostly to the organic material of bone nodules in mineralized bone matrix, with very weak or no staining of the rest of the bone matrix. This staining progressively decreased toward the mineralized cartilage matrix and became negative. The mineralized cartilage matrix and lamina limitans reacted strongly with all mAbs except 5-D-4. These results indicate that osteoid contains sulphated proteoglycans containing C4-S and/or DS, C6-S and KS, and subsequent bone matrix mineralization appears to require accumulation of these macromolecules within bone nodules and eventual loss of these substances for complete mineralization, whereas proteoglycans containing C0-S, C4-S and/or DS, and C6-S, still exist in mineralized cartilage matrix and lamina limitants.     

Evidence of low molecular weight components in the organic matrix of the reef building coral, Stylophora pistillata

Biominerals contain both inorganic and organic components. Organic components are collectively termed the organic matrix, and this matrix has been reported to play a crucial role in mineralization. Several matrix proteins have been characterized in vertebrates, but only a few in invertebrates, primarily in Molluscs and Echinoderms. Methods classically used to extract organic matrix proteins eliminate potential low molecular weight matrix components, since cut-offs ranging from 3.5 to 10 kDa are used to desalt matrix extracts. Consequently, the presence of such components remains unknown and these are never subjected to further analyses. In the present study, we have used microcolonies from the Scleractinian coral Stylophora pistillata to study newly synthesized matrix components by labelling them with 14C-labelled amino acids. Radioactive matrix components were investigated by a method in which both total organic matrix and fractions of matrix below and above 5 kDa were analyzed. Using this method and SDS-PAGE analyses, we were able to detect the presence of low molecular mass matrix components (<3.5 kDa), but no free amino acids in the skeletal organic matrix. Since more than 98% of the 14C-labelled amino acids were incorporated into low molecular weight molecules, these probably form the bulk of newly synthesized organic matrix components. Our results suggest that these low molecular weight components may be peptides, which can be involved in the regulation of coral skeleton mineralization.     

Structural Organisation of the Cusps of the Radular Teeth of the Chiton Plaxiphora albida

Abstract The structure, morphology and organisation of the cusps of the major lateral radula teeth of the chiton Plaxiphora albida have been examined using light, transmission and scanning electron microscopy, together with energy dispersive X-ray analysis and Mössbauer spectroscopy. In this chiton species, both the anterior and posterior surfaces of the major lateral teeth are composed of magnetite, which is indicated to be non-stoichiometric and associated with some maghemite, together with small amounts of phosphorus and silicon. This outer layer surrounds an inner core region of the tooth, which only reaches the surface through a small window zone on the anterior surface and which contains large amounts of iron and phosphorus presumably in the form of iron(III) phosphate. The organic matrix, on which the teeth are constructed, consists of a zone of densely packed fine fibres at the surface of the tooth, underlain by larger fibres which become sparser deeper into the cusp. The core region is characterized by the presence of densely packed short fibres. In contrast to the situation found in most other species of chiton, large fibres of the organic matrix extend throughout the region of magnetite mineralization, leading to the suggestion that the matrix exerts more control over the mineralization of magnetite than has previously been thought.     

Soil C and N dynamics within a precipitation gradient in Mediterranean eucalypt plantations

Soil C and N dynamics were evaluated in five eucalypt plantations within a precipitation gradient (500–2,000 mm) in Portugal. Soil physical and chemical properties, total and labile (particulate organic matter, hydrolyzable, hot water soluble and microbial) soil C and N pools, and C and N mineralization were measured to characterize the C and N dynamics and their controlling factors within this gradient. Contents of total and labile soil organic C and N were positively correlated with the mean annual precipitation. A similar relationship was observed for net N mineralization (anaerobic and long-term aerobic incubation), gross N mineralization (¹⁵N isotope dilution technique) and C mineralization. In contrast, rates of C and N mineralization (per unit of C and N) were higher in the driest sites due to their higher proportion of particulate organic matter C. Net and gross N mineralization were strongly correlated and showed similar controlling factors (mean annual precipitation, total and labile C and N and extractable P contents), suggesting that net N mineralization during long-term aerobic incubation reflects gross N transformations. Although, gross NO₃–N production and gross NO₃–N immobilization were observed in all sites, net nitrification in the drier sites was not observed in the first weeks of the study. Our results suggest that, under Mediterranean conditions, mean annual precipitation is the major factor determining the C and N dynamics in soils with Eucalyptus plantations.     

Structure of the nacreous organic matrix of a bivalve mollusk shell examined in the hydrated state using cryo-TEM

During mollusk shell formation, the mineral phase forms within an organic matrix composed of beta-chitin, silk-like proteins, and acidic glycoproteins rich in aspartic acid. The matrix is widely assumed to play an important role in controlling mineralization. Thus, understanding its structure is of prime importance. Cryo-transmission electron microscopy (Cryo-TEM) studies of the matrix of the bivalve Atrina embedded in vitrified ice show that the interlamellar sheets are composed mainly of highly ordered and aligned beta-chitin fibrils. The silk, which is quantitatively an important component of the matrix, could not be imaged within the sheets. Organic material was, however, observed between sheets. We infer that this is the location of the silk. As this material reveals no regular structure, we suggest that at least prior to mineralization the silk is in the form of a hydrated gel. This is supported by cryo-TEM structural observations of an artificial assembly of beta-chitin with and without silk. This view of the nacreous organic matrix significantly changes previous models of the matrix structure and hence hypotheses pertaining to the mechanisms by which mineral formation occurs.     

Persistence of cartilage proteoglycan and link protein during matrix-induced endochondral bone development: An immunofluorescent study

Monospecific antibodies to cartilage proteoglycan monomer and link protein were employed with immunofluorescence microscopy to determine the tissue distribution of these constituents during matrix-induced endochondral bone development. Subcutaneous implantation of demineralized diaphyseal bone matrix resulted in new endochondral bone formation. On Day 3, the implant consisted of mesenchymal tissue which did not contain any demonstrable cartilage-related proteoglycan or link protein. With the onset of early chondrogenesis on Day 5, cartilage proteoglycan monomer and link protein were first localized together in the cartilage matrix, particularly around chondrocytes in territorial sites. Progressively more staining around cells was observed at Days 7 and 9. On Day 9, when mineralization was first observed, there was no evidence of a net loss of these molecules prior to mineralization of the cartilage matrix. On Day 11 and thereafter, bone formation was observed by appositional growth on calcified cartilage spicules. Whereas the osteoblasts and bone matrix were devoid of any staining for cartilage proteoglycan and link components, the residual, partly mineralized cartilage spicules still reacted with antibodies to cartilage proteoglycan monomer and link protein in territorial sites, but in reduced amounts, indicating a loss of these molecules associated with a loss of hypertrophic chondrocytes. Since mineral prevented the access of Fab' antibody subunits, demineralization after fixation was routinely employed. The results reveal that cartilage proteoglycan monomer and link protein are present around chondrocytes in hyaline cartilage during the early stages of endochondral bone formation and that there is no net loss of these molecules prior to mineralization of this cartilage matrix as was previously thought.     

Proteins and saccharides of the sea urchin organic matrix of mineralization: characterization and localization in the spine skeleton

Properties of the echinoderm skeleton are under biological control, which is exerted in part by the organic matrix embedded in the mineralized part of the skeleton. This organic matrix consists of proteins and glycoproteins whose carbohydrate component is specifically involved in the control mechanisms. The saccharide moiety of the organic matrix of the spines of the echinoid Paracentrotus lividus was characterized using enzyme-linked lectin assays (ELLAs). O-glycoproteins, different types of complex N-glycoproteins, and terminal sialic acids were detected. Sialic acids are known to interact with Ca ions and could play an important role in the mineralization process. Some of the carbohydrate components detected by ELLAs as well as two organic matrix proteins (SM30 and SM50) were localized within different subregions of the spine skeleton using field-emission scanning electron microscopy. The mappings show that some of these components are not homogeneously distributed in the different skeletal subregions. For example, some N-glycoproteins were preferentially located in the putative amorphous subregion of the skeleton, whereas some O-glycoproteins were localized in the subregion where skeletal growth is inhibited. These results suggest that the biological control exerted on the skeletal properties can be partly modulated by local differences in the organic matrix composition.     

The penetration of exogenous tracers through the enameloid organ of developing teleost fish teeth

In order to determine whether exogenous materials permeate to the forming tooth enameloid matrix, teleost species were injected intramuscularly with horseradish peroxidase (HRP) or myoglobin, or; intracardially with lanthanum nitrate or HRP, then killed a predetermined intervals post-injection. Tooth bearing bones were processed for transmission electron microscopy. At the enameloid matrix formation stage, capillaries associated with the enameloid organ were few in number and rarely fenestrated. Both organic tracers reached the matrix at cervical but not coronal, regions of the teeth in all species examined. Lanthanum was rarely observed extravascularly and never extended to the enameloid matrix at the secretion stage. At the enameloid mineralization stage, fenestrated capillaries were closely associated with the outer dental epithelial cells (ODE). All tracers were observed in the plasma membrane invaginations of the ODE. Only intracardially injected HRP compromised the apical intercellular junctions of the inner dental epithelial cells (IDE) to reach the mineralizing enameloid Lanthanum did not extend past the ODE-IDE cell junctions. It is concluded that the close association of mineralization stage fenestrated capillaries with the highly invaginated ODE cells result in increased tracer penetration compared to the secretory stage. The deeper penetration of the organic tracers, compared with lanthanum, between mineralization stage IDE cells may be due to longer in vivo circulation of the former material. The apical junctions of mineralization stage IDE cells, however, remained impermeable to the organic tracers. The absence of mineral in secretory stage enameloid mineral could not be due to specialized cell junctions preventing access of molecules to the matrix. It is suggested that controlling factors other than cellular permeability initiate enameloid mineralization.     

Soil nitrogen dynamics along a gradient of long-term soil development in a Hawaiian wet montane rainforest

I analyzed the rates of net N mineralization and nitrification of soils from seven sites in a Hawaiian wet montane forest. The sites differ in age, ranging from 400 to 4,100,000 yr, but are comparable in other variables (all at 1200 miasl with 4000 mm or more mean annual rainfall), and the chronosequence simulated a development of soils from basaltic lava. Soils were incubated for 20 days at 17.5 °C, which is nearly equivalent to a mean field air temperature of the sites, and at an elevated temperature of 25.5 °C under three treatments: 1) field-wet without amendments, 2) air dried to a permanent wilting point, and 3) fertilized with phosphate (NaH₂PO₄) at the rate of 50 g P per g dry soil. Both mineralization and nitrification rates varied significantly among the sites at the field temperature (p<.00001). Fractions of the mineralized organic matter (indexed by the N produced per g organic C) increased sharply from the youngest to the 5000-yr site before declining abruptly to a near constant value from the 9000 to the 1,400,000-yr sites. Total organic C in the top soils (<15 cm deep) increased almost linearly with age across the sites. Consequently, net NH₄- and NO₃-N produced on an area basis (g m^-2 20 d^-1) increased sharply from 0.2 in the youngest site to 1.2 in the 5000-yr site, then both became depressed once but steadily increased again. The fraction of organic matter mineralized, and the net N turnover rates were outstandingly high in the oldest site where a large amount of organic matter was observed; the topsoil organic matter which was used in this analysis appeared to be highly labile, whereas the subsurface organic matter could be relatively recalcitrant. As suggested by earlier workers, the initial increase in N turnover seemed to correspond to the increasing quantity of N in the soils through atmospheric deposition and biological fixation. The later decline in fraction of organic matter mineralized seemed to relate to increasing soil C/N ratios, increasingly recalcitrant organic matter, and poorer soil drainage with age. The elevated temperature treatment produced significantly higher amounts of N mineralization, except for the youngest site where N was most limiting, and for two sites where soil waterlogging might be severe. P fertilization invariably resulted in slower N turnovers, suggesting that soil microbes responded to added P causing N immobilization. The youngest site did not significantly respond to added P. The magnitude of immobilization was higher in older than in younger soils, suggesting that P more strongly limits microbial populations in the older soils.     

A simple model for estimating the contribution of nitrogen mineralization to the nitrogen supply of crops from a stabilized pool of soil organic matter and recent organic input

A simple model was developed to estimate the contribution of nitrogen (N) mineralization to the N supply of crops. In this model the soil organic matter is divided into active and passive pools. Annual soil mineralization of N is derived from the active pool. The active pool comprises stabilized and labile soil organic N. The stabilized N is built up from accumulated inputs of fresh organic N during a crop rotation but the labile N is a fraction of total N added, which mineralizes faster than the stabilized N. The passive pool is considered to have no participation in the mineralization process. Mineralization rates of labile and stabilized soil organic N from different crop residues decomposing in soil were derived from the literature and were described by the first-order rate equation dN/dt =-K*N, where N is the mineralizable organic N from crop residues andK is a constant. The data were groupedK ₁ by short-term (0–1 year) andK ₂ by long-term (0–10 years) incubation. Because the range of variation inK ₂ was smaller than inK ₁ we felt justified in using an average value to derive N mineralization from the stabilized pool. The use of a constant rate ofK ₁ was avoided so net N mineralization during the first year after addition is derived directly from the labile N in the crop residues. The model was applied to four Chilean agro-ecosystems, using daily averages of soil temperature and moisture. The N losses by leaching were also calculated. The N mineralization varied between 30 and 130 kg N ha^–1 yr^–1 depending on organic N inputs. Nitrogen losses by leaching in a poorly structured soil were estimated to be about 10% of total N mineralized. The model could explain the large differences in N- mineralization as measured by the potential N mineralization at the four sites studied. However, when grassland was present in the crop rotation, the model underestimated the results obtained from potential mineralization.     

Preservation of living organic structures in unicellular and multicellular organisms in the fossil state

It is known that early postmortem mineralization is a phenomenon underlying the preservation of organic structures such as bacterial cells, biofilms, soft bodies, cells, vessels, and slightly mineralized integuments of multicellular animals in the fossil state. Bacteria, the activity of which creates conditions for mineralization of organic matter, are a driving force behind this process. The most widely known types of postmortem mineralization that are mediated by bacteria are phosphatization, silicification, pyritization, carbonatization, iron-manganese mineralization, and the formation of clay minerals, which either replace organic structures or cover them with thin films.     

Transmission microscopy of freeze dried,unstained epiphyseal cartilage of the guinea pig

Summary The question of the initial mineralization in the epiphyseal plate has been investigated to date in specimens prepared by conventional electron microscopical techniques. As conventional techniques can produce artifacts, either a loss of mineral substance or a secondary nucleation, the mineralization process was investigated using freeze dried, vacuum embedded growth cartilage which was neither contrasted nor stained and which had a very short contact with water.The prevailing theory that the first mineralization begins within extracellular matrix vesicles and that the mineralization outside these vesicles is a secondary process was confirmed. Mineralized matrix vesicles were found in the fully mineralized long septa down to the opening zone. In several cases a mineralization could be observed in those transverse septa in which organic substance was present between the cells. The typical radial arrangement of the apatitic needles and platelets in the matrix vesicles could be explained by the formation of a mineralization in an ionotropic gel, the orientation of the matrix macromolecules to be produced by a vectorial influx of calcium ions and phosphate groups coming from different directions. Thin strands of mineral substance with low contrast, which follow the direction of the longitudinal septum, were assumed to be the mineralized collagen fibrils. In several needles dot-like formations were seen and the distance between the middle of neighbouring dots was found to lie mainly in the range 30–56 Å, while the lateral separation distance between the middle of closely packed parallel chains and needles was found to lie mainly in the range 30-42 Å. Parallel periodic structures which could be visualized in apatitic chains and needles 20–40 Å in diameter were assumed to be the 8.2 Å-(100)-lattice planes of apatite, being an indication that these formations already possess criteria of the apatite lattice.We express our thanks to the Deutsche Forschungsgemeinschaft for financial support and to Dr. A. Boyde, London, for valuable discussions.     

Effect of temperature on submerged macrophyte litter decomposition within sediments from a large shallow and subtropical freshwater lake

In shallow aquatic systems, the majority of organic matter mineralization occurs in the sediments. Several factors including temperature control mineralization rates, however, the underlying causes of the effects are not well understood in subtropical lakes. In this study, we determined the influence of temperature on organic matter degradation by taking sediments from four sites in a subtropical large shallow freshwater lake, and monitoring organic matter composition and enzymes in microcosm experiments at five temperatures from 5 to 40°C. Following a three-month incubation, it was found that the mineralization of submerged plants in sediments was strongly influenced by temperature. Removal efficiency of total organic carbon in sediments ranged from 4.3 to 22.6% at 5°C, and reached 46.7–55.5% at 40°C. In addition, the removal efficiency of organic matter and the relative recalcitrant carbon decomposition depended on sediment type. For sediments in the site located in the lake center, recalcitrant and labile carbon decomposition had equivalent responses to the different temperatures. For sediments with dominance of submerged macrophytes, the humic acids were low even at high temperature. Thus, the annual deposition of plant litter in sediments favored organic carbon decomposition rather than humification.     

Do stress-whitening and optical clearing of collagenous tissue occur by the same mechanism?

Bone is biphasic with an organic matrix and an inorganic mineral component. As we age bone's susceptibility to fracture increases. It has been shown that there is no change in mean mineralization with aging, but bone nevertheless becomes less tough. This aging effect is therefore likely related to the organic phase.     

Decomposition of soil organic matter from boreal black spruce forest: environmental and chemical controls

Black spruce forests are a dominant covertype in the boreal forest region, and they inhabit landscapes that span a wide range of hydrologic and thermal conditions. These forests often have large stores of soil organic carbon. Recent increases in temperature at northern latitudes may be stimulating decomposition rates of this soil carbon. It is unclear, however, how changes in environmental conditions influence decomposition in these systems, and if substrate controls of decomposition vary with hydrologic and thermal regime. We addressed these issues by investigating the effects of temperature, moisture, and organic matter chemical characteristics on decomposition of fibric soil horizons from three black spruce forest sites. The sites varied in drainage and permafrost, and included a “Well Drained” site where permafrost was absent, and “Moderately well Drained” and “Poorly Drained” sites where permafrost was present at about 0.5 m depth. Samples collected from each site were incubated at five different moisture contents (2, 25, 50, 75, and 100% saturation) and two different temperatures (10°C and 20°C) in a full factorial design for two months. Organic matter chemistry was analyzed using pyrolysis gas chromatography-mass spectrometry prior to incubation, and after incubation on soils held at 20°C, 50% saturation. Mean cumulative mineralization, normalized to initial carbon content, ranged from 0.2% to 4.7%, and was dependent on temperature, moisture, and site. The effect of temperature on mineralization was significantly influenced by moisture content, as mineralization was greatest at 20°C and 50–75% saturation. While the relative effects of temperature and moisture were similar for all soils, mineralization rates were significantly greater for samples from the “Well Drained” site compared to the other sites. Variations in the relative abundances of polysaccharide-derivatives and compounds of undetermined source (such as toluene, phenol, 4-methyl phenol, and several unidentifiable compounds) could account for approximately 44% of the variation in mineralization across all sites under ideal temperature and moisture conditions. Based on our results, changes in temperature and moisture likely have similar, additive effects on in situ soil organic matter (SOM) decomposition across a wide range of black spruce forest systems, while variations in SOM chemistry can lead to significant differences in decomposition rates within and among forest sites.     

Skeletal formation in the modern but ultraconservative chaetetid spongeSpirastrella (Acanthochaetetes) wellsi (demospongiae,porifera)

Summary The modern hadromerid coralline spongeSpirastrella (Acanthochaetetes) wellsi exhibits a unique secondary high-Mg calcite (>19 mol % MgCO₃) basal skeleton. The basal skeleton is constructed of bundles of elongated crystals more or less tangentially orientated. The initial formation of these crystals is controlled by soluble highly acidic aspartic and glutamic-rich (40%) macromolecules. The skeletal mineralization occurs in four different loci: in the top of the calicles, at the tabulae, on collagenous anchor fibres, and within closed spaces between the tabulae. The clicle walls are formed on the uppermost top of the basal skeleton as a continuous process. Based on long term stainings with Ca²⁺-chelating fluorochroms (calcein, chlorotetracyclines) the growth rate of this sponge is extremely low with ca. 50–100μm/a. The skeletal formation takes places outside the sponge, within a narrow zone (300–500 nm) between the basopinacoderm and the mature basal skeleton. The sponge produces thread-like folded templates (‘spaghetti fibres’) of 0,5–2 μm size, the shape controlling insoluble organic matrix. These templates become mineralized in a first step as MgCO₃, then are stretched. A soluble organic matrix is also secreted, and remains are included inside the mineralized skeleton. This organic matrix consists of in a complex mixture containing small very acidic proteins (5, 13, 31 KD; 40% Asp and Glu and therefore most probably Ca²⁺-binding) and high molecular weight glycoproteins among several other organic compounds. The mature crystals are high-Mg calcites. During calcification large cells with large reserve granules (LCG) are always present in a tight connection with the basopinacoderm. These cells form also the collagenous anchor fibres. Primary tabulae are formed by a non-collagenous organic sheet. Calcification happens only when LCG cells are enriched on the organic sheet. Randomly oriented high-Mg calcite crystals are growing on the collagenous anchor fibres. The same type of the mineralization is observed within the spaces of the tabulae. This particular case of mineralization is controlled by decaying sponge tissue (ammonification). The δ¹³C values are in equilibrium with the ambient sea water and vary between +3.2 and +2.8 ‰. The mode of mineralization of the basal skeleton can be described as biologically induced resp. matrix mediated.     

Role of intestinal mucus in crystal biogenesis: an electron-microscopical,diffraction and X-ray microanalytical study

Summary In the posterior intestine of the sea-water eel, mucus plays an important role in biocrystallization of calcium ions. By means of transmission and scanning electron microscopy associated with X-ray microanalysis and X-ray diffraction it has been possible to determine the role of mucous fibers as nucleation sites. Biocrystallization occurs in 2 steps: (1) Calcification of mucus. As soon as mucus is excreted in the intestinal lumen, it is loaded with calcium, as shown by lanthanum affinity and X-ray microanalysis on freeze-dried tissues. (2) Genesis of crystals. Needleshaped crystallites build up in coalescent spherites in the intestinal lumen near the microvilli. Genesis occurs as follows: (a) crystallite mineralization by nucleation in an organic matrix composed of glycoproteinaceous mucous fibers, followed by the appearance of spherites; (b) coalescence in spherites and association of spherites in rhombohedra; (c) extrusion of organic material during the final step of crystallization.     

The roles of autophosphorylation and phosphorylation in the life of osteopontin

Osteopotin is a secreted glycosylated phosphoprotein found in bone and other normal and malignant tissues. Osteopontin can be autophosphorylated on tyrosine residues and can also be phosphorylated on serine and threonine residues by several protein kinases. Autophosphorylation of osteopontin may generate sites for specific interactions with other proteins on the cell surface and/or within the extracelluar matrix. These interactions of osteopontin are thought to be essential for bone mineralization and function. The polyaspartic acid motif of osteopontin, in combination with neighboring sequences that include serine residues phosphorylated by protein kinases, could fold and assemble into a molecular structure that participates in the mineralization of the bone matrix.     

Localization of calcium and phosphorus in early predentin-matrix components by electron spectroscopic imaging (ESI)-analysis in rat molars

Summary The subcellular distribution of the inorganic elements calcium (Ca) and phosphorus (P) was studied in the first-formed dentin matrix during initial mineralization in neonatal rat molars. This most peripheral matrix region is comprised of a proteoglycan-rich ground substance, interwoven by a collagenous network, matrix vesicles, aperiodic fibrils derived from the dental basal lamina, and apical odontoblastic cell processes. All matrix components may possibly serve as templets for mineral deposition during initial calcification of first-formed mantle dentin and predentin. By means of the very sensitive ESI-analysis we studied the subcellular localization of Ca and P and their possible association with distinct organic extracellular matrix components and odontoblasts. Ca-signals were found in the ground substance, at striated collagen fibrils and plasma membranes of odontoblasts in the cuspal early matrix region, but occurred only sparsely in the ground substance of the more distal matrix region where odontoblast processes attach to aperiodic fibrils of the dental basal lamina. Ca was generally absent in matrix vesicles. In contrast, P-signals were found in matrix vesicles, at aperiodic fibrils and at the plasma membranes of odontoblasts. Ca and P co-localized at striated collagen fibrils (type I or II). These results suggest that striated collagen fibrils might serve as primary deposition sites for calcium phosphate during early biological calcification of organic extracellular macromolecules.