DEGRADATION OF HAMSTER AMELOGENINS DURING SECRETORY STAGE ENAMEL FORMATION IN ORGAN-CULTURE

Citation
Aljj. Bronckers et al., DEGRADATION OF HAMSTER AMELOGENINS DURING SECRETORY STAGE ENAMEL FORMATION IN ORGAN-CULTURE, Matrix biology, 14(7), 1995, pp. 533-541
Citations number
34
Categorie Soggetti
Biology,"Cell Biology
Journal title
ISSN journal
0945053X
Volume
14
Issue
7
Year of publication
1995
Pages
533 - 541
Database
ISI
SICI code
0945-053X(1995)14:7<533:DOHADS>2.0.ZU;2-U
Abstract
Increasing amelogenin heterogeneity during pre-eruptive enamel formati on has been explained by proteolytic cleavage of a parent amelogenin, differences in posttranslational modicifactions, translation of multip le alternative spliced mRNA transcripts or combinations of these possi bilities. We investigated the possibility of proteolytic degradation o f amelogenins during secretory amelogenesis by pulse-labelling ameloge nins with [H-3]proline followed by a pulse chase, all under organ cult ure conditions. The results indicate that during pulse chase, hamster molar tooth explants rapidly released substantial amounts of the radio activity into the culture medium, as non-trichloroacetic-acid precipit able, noncollagenous H-3-activity at the expense of radioactivity asso ciated with the proteins in the enamel space. Simultaneously, there wa s a continuous mineralization of the forming enamel in vitro as shown by an increase in total calcium content of the explants. Western blott ing, microdissection studies and fluorography of radiolabelled matrix proteins after SDS-PAGE indicated that after an 8-h labelling, three r adioactive amelogenin species could be extracted from forming enamel, one prominent species of molecular mass 26 kDa and two less prominent ones of 28 and 22 kDa. During pulse chase more amelogenin bands with l ower molecular mass became apparent, a pattern similar to that observe d in vivo. Examination of amelogenin blots with the glycan assay showe d that none of the hamster amelogenins stained for carbohydrate. We co nclude that changes in the amelogenin profiles during enamel developme nt of cultured hamster explants are similar to those observed in vivo. Although the formation of differently sized amelogenins from alternat ively spliced mRNA transcripts cannot be ruled out, the in vitro pulse -chase data obtained in this study suggest that amelogenin degradation per se is probably responsible for most of the changes seen in the am elogenin profiles.