Amelogenin-cytokeratin 14 interaction in ameloblasts during enamel formation

The GMp motif is found in the N-terminal region of CK14, a differentiation marker for ameloblasts. The binding affinity of CK14 and amelogenin was con firmed by dosimetric binding of CK14 to recombinant amelogenin (rM179), and to the tyrosine-rich amelogenin polypeptide. The specific binding site for CK14 was identified in the amelogenin trityrosyl motif peptide (ATMP) of t yrosine-rich amelogenin polypeptide and specific interaction between CK14 a nd [H-3]ATMP was confirmed by Scatchard analysis. Blocking rM179 with GlcNA c, GMp, or CK14 with ATMP abrogates the CK14-amelogenin interaction. CK14 f ailed to bind to ATMP when the third proline was substituted with threonine , as in some cases of human X-linked amelogenesis imperfecta or when tyrosy l residues were substituted with phenylalanine. Morphometry of developing t eeth distinguished three phases of enamel formation; growth initiation phas e (days 0-1), prolific growth phase (days 1-7), and growth cessation phase (post-day 7). Confocal microscopy revealed co-assembly of CK14/amelogenin i n the perinuclear region of ameloblasts on day 0, migration of the co-assem bled CK14/amelogenin to the apical region of the ameloblasts from day 1, re aching a peak on days 3-5, and a collapse of the co-assembly. Autoradiograp hy with [H-3]ATMP and [H-3]GMp corroborated the dissociation of the co-asse mbly at the ameloblast Tomes' process. It is proposed that CK14 play a chap eron role for nascent amelogenin polypeptide during amelogenesis.