Immunohistochemical Similarities and Differences between Amelogenin and Tuftelin Gene Products During Tooth Development

Abstract

SUMMARYAmelogenins and tuftelins are highly specialized proteins secreted into the developing enamel matrix during mammalian enamel formation. Both tuftelins and amelogenins have been associated with various functions during nucleation and maturation of the developing enamel matrix. In this study we conducted experiments to investigate whether tuftelins and portions of the amelogenin molecule were deposited and processed in spatially distinguished portions of the developing enamel matrix, using antibodies specific against tuftelin or amelogenins. The amelogenin antibodies were raised against recombinant and native amelogenins and also included an antibody against a polypeptide encoded by amelogenin exon 4. To compare spatial expression patterns of enamel protein epitopes, 3-day postnatal mouse molar tooth organs were processed for paraffin histology and cut into serial sections. Adjacent sections were exposed to antibodies against either tuftelin or various amelogenin epitopes. To investigate age-related changes of enamel protein expression, amelogenin and tuftelin antibodies were applied to tooth organs of developmental stages E19 and 1, 3, 5, 7, 9 and 11 postnatal days. Tuftelin was detected within the odontoblast processes during earlier stages of development (E19 and 1 day postnatal), whereas during later stages (3-11 days) it was recognized in a portion of the enamel layer adjacent to the dentine-enamel junction. In contrast, all four antibodies against amelogenins reacted with parts of the ameloblast cytoplasm and the entire enamel layer. Using immunohistochemistry, we were not able to detect any differences in the spatial distribution of the four amelogenin epitopes investigated. The spatial differences in the distribution of amelogenin and tuftelin as observed in this study may be intepreted as an indication of functional differences between both proteins during early enamel biomineralization.

Membranes, minerals, and proteins of developing vertebrate enamel

“…Although some of these recently identified enamel proteins have been localized in tissues outside of teeth, they might be of relevance to the mechanisms of enamel formation (Deutsch et al, 1991;ZeichnerDavid et al, 1995ZeichnerDavid et al, , 1997MacDougall et al, 1998). Tuftelin, the nonamelogenin enamel protein investigated in this study, has been sequenced and characterized (Deutsch, 1989;Deutsch et al, 1991;ZeichnerDavid et al, 1995ZeichnerDavid et al, , 1997Diekwisch et al, 1997;Mao et al, 2001;Satchell et al, 2002).…”

Laminin α2 Is Essential for Odontoblast Differentiation Regulating Dentin Sialoprotein Expression

“…We used antibodies directed against laminin ␣2 (4H8 -2, Alexis) (12), ameloblastin (AMBN) (17), amelogenin (AMEL) (18,19), and DSP (20) (provided by Yoshihiko Yamada). The primary antibodies were detected using fluorescein isothiocyanate or Cy-3-conjugated secondary antibodies (Jackson ImmunoResearch).…”

Identification of CCAAT/Enhancer-binding Protein α as a Transactivator of the Mouse Amelogenin Gene

“…Amelogenins are essential to the proper regulation of enamel mineralization. Amelogenin expression is ameloblast-specific and developmentally regulated at the temporal and spatial levels (1)(2)(3)(4)(5)(6)(7)(8). A 2263-nucleotide proximal promoter element from the mouse X-chromosomal amelogenin gene has been demonstrated by transgenic mouse analysis to direct the expression of a reporter gene in a temporal and spatial pattern that is essentially identical to that of the endogenous amelogenin gene (5).…”