Localization of bone sialoprotein (BSP) to Golgi and post-Golgi secretory structures in osteoblasts and to discrete sites in early bone matrix.

Abstract

Bone sialoprotein (BSP), a bone matrix-enriched glycoprotein containing the Arg-Gly-Asp (RGD) motif and endowed with cell binding properties, was localized in osteoblasts and early bone matrix of developing rat bone at the ultrastructural level. Preliminary light microscopic observations indicated that intracellular labelling was restricted to a paranuclear dot corresponding to the "negative Golgi image" of classical histology. The same pattern was observed whether antisera against the fully glycosylated protein or a peptide antiserum to a stretch of amino acids in human BSP sequence were employed. At the EM level, we obtained labeling over the Golgi area of osteoblasts but not over the rER. The labeling was concentrated over distensions of the trans Golgi and over pro-secretory granules. In the matrix, BSP was distributed in a non-random manner. The label was concentrated over spherical aggregates of finely fibrillar material corresponding to the sites of early mineral deposition (so-called "mineralization nodules"). Such BSP-positive foci were seen both close to and away from the cell surface. The predominant association of BSP with Golgi and post-Golgi secretory structures and its absence from rER, as well as the reproducibility of the same pattern of localization with different antisera, might indicate a slow transit of the protein through the Golgi, not necessarily associated with protein glycosylation.

Calcospherulites11Calcospherulites (calco: calcium salt+spherulite: spherical crystalline body); calcium-containing, spherical bodies have also been referred to as calcified microspheres, mineral clusters, crystal ghost aggregates, calcification nodules, calcospherites, and small calcified spheres. isolated from the mineralization front of bone induce the mineralization of type I collagen

“…Several other groups have reported the presence of discrete spherical or ovoid shaped entities at the mineralization front of bone having similar structural and elemental characteristics to what is described in the current study [11][12][13][14][15][16][17][18][19]32,33]. A unique contribution of the present study to this existing literature is the isolation and initial biochemical characterization of these mineralization front structures.…”

“…Another finding from these tests revealed that calcospherulites incubated in type I collagen gels take on an ultrastructural appearance similar to that of crystal ghost aggregates observed in vivo [9]. Since both our isolated calcospherulites and crystal ghost aggregates [13] contain BSP, then this ultrastructural similarity may not be coincidental and might suggest a structure/function identity. Altogether, the similarities between our in vitro results and observations made from in vivo samples suggest that calcospherulites likely mediate the mineralization of osteoid at the mineralization front of bone tissue in vivo.…”

“…Many other investigators have reported the presence of calcified, spherical-shaped structures along the mineralization front of bone [11][12][13][14][15][16][17][18][19]. Recently, we have reported that calcified spherulites tend to cluster in select matrix areas during periods of rapid bone formation, and that this focal process is defined by an extracellular matrix assembly of bone acidic glycoprotein-75 (BAG-75) 1 referred to as biomineralization foci (BMF) [20,21].…”

“…Recently, we have reported that calcified spherulites tend to cluster in select matrix areas during periods of rapid bone formation, and that this focal process is defined by an extracellular matrix assembly of bone acidic glycoprotein-75 (BAG-75) 1 referred to as biomineralization foci (BMF) [20,21]. One of the bone matrix proteins associated with these spherulites, bone sialoprotein (BSP) [13,16], has been shown to nucleate apatite crystals in vitro [22]. It is not known how these spherulites form or how they interact with fibrillar collagen, which stems, in part, from a lack of a method to release and recover these structures specifically from the mineralization front of bone [17].…”

Altered Cbfal expression and biomineralization in an osteosarcoma cell line

“…These were small needle-like crystals associated with fibrillar extracellular matrix deposits and intracellular spherical structures, with an X-ray diffraction pattern resembling human bone [15]. This pattern resembled the early stages in mineral formation in primary osteoblast cultures [2,4,9] and normal rat bone sections [3], and their chemical composition was bio-apatitic in nature, resembling normal bone in crystal size and calcium phosphate ratio [15]. Furthermore, when UMR cells were experimentally transplanted in animals they formed mineralized bone trabeculae at ectopic sites [I 11.…”