Endocytosis and Enamel Formation

Abstract

Enamel formation requires consecutive stages of development to achieve its characteristic extreme mineral hardness. Mineralization depends on the initial presence then removal of degraded enamel proteins from the matrix via endocytosis. The ameloblast membrane resides at the interface between matrix and cell. Enamel formation is controlled by ameloblasts that produce enamel in stages to build the enamel layer (secretory stage) and to reach final mineralization (maturation stage). Each stage has specific functional requirements for the ameloblasts. Ameloblasts adopt different cell morphologies during each stage. Protein trafficking including the secretion and endocytosis of enamel proteins is a fundamental task in ameloblasts. The sites of internalization of enamel proteins on the ameloblast membrane are specific for every stage. In this review, an overview of endocytosis and trafficking of vesicles in ameloblasts is presented. The pathways for internalization and routing of vesicles are described. Endocytosis is proposed as a mechanism to remove debris of degraded enamel protein and to obtain feedback from the matrix on the status of the maturing enamel.

Keywords: Rab proteins; amelogenesis; clathrin; endocytic trafficking; endocytosis; pinocytosis.

Figure 1

Immunocytochemical preparation illustrating the distribution of gold labeled amelogenin over various compartments of ameloblasts from the secretion stage in mouse incisors. Lysosomes appear variably labeled. Multivesicular bodies are often intensely labeled (mvb+). An unlabeled multivesicular body (mvb−) and an unlabeled dark lysosome (dl−) are shown. The Golgi apparatus (G) shows some labeling by gold particles, × 24,875. Bar = 0.5 μm. Permission to reprint from: Application of High-Resolution Immunocytochemistry To the Study of the Secretory, Resorptive, and Degradative Functions of Ameloblasts by Nanci et al. (1987a).

Figure 2

Sites of secretion and endocytosis of the ameloblast membrane. (A) The secretory ameloblasts forms a Tomes' process from the apical membrane with a proximal portion and a distal portion. The proximal portion is associated with formation of interrod enamel, the distal portion with rod enamel formation. Vesicle fusion can be observed on the surface membrane adjacent to the rod growth site. Many vesicles fuse (secretion) or originate (endocytosis) from membrane infoldings found on the proximal portion and the distal portion. (B) In the maturation stage, degraded enamel proteins are internalized by ameloblasts. Ameloblasts modulate between smooth-ended and ruffle bordered membranes. In 80% of the maturation stage, ameloblasts are ruffle-ended with deep membrane invaginations. Degraded enamel proteins from the enamel matrix permeate the area between convoluted tubules and are resorbed via vesicles.

Figure 3

Early and mid secretory stage ameloblasts in mandibular mouse incisors. All procedures involving animals were reviewed and approved by the IACUC committee at the University of Michigan and all relevant guidelines were followed. Handling of animals and tissues was followed according to protocols described earlier (Smith et al., 2016). (A) The Tomes' processes are not fully developed in early secretory stage ameloblasts. Enamel crystallite ribbons in the initial interrod layer are oriented perpendicular to dentin and the apical ameloblast membrane. (B) In mid secretory ameloblasts pinocytotic vesicles are observed along the non-secreting surface of the Tomes' process and laterally between adjacent ameloblasts below the level of the apical junctional complex. In the distal portion of the Tomes process membrane infoldings form a complex network and are in close relationship to vesicles. D, dentin; E, enamel; mvb, multivesicular body; acw, apical cell web; rER, rough endoplasmic reticulum; m, mitochondria; db, dense body (lysosome); im, membrane infolding; dTP, distal portion of Tomes' process; IRG, interrod growth site; pTP, proximal portion of Tomes' process; sg, secretory granule; p, pinocytotic vesicle.

Figure 4

Vesicular trafficking pathways during endocytosis. Schematic diagram showing the general pathways of internalized material. Molecules are taken up into vesicles via various types of endocytosis (receptor-mediated, phagocytosis, micropinocytosis, pinocytosis). Vesicles (coated and non-coated) fuse with EE, the first cellular sorting station, where the cargo is distributed to MVB/LE, GA or back to the plasma membrane (recycling pathway) via their respective Rab proteins. Cargo which is destined for degradation in lysosomes, is transported from EE to lysosomes via MVB/LE. At MVB/LE vesicles can still enter the recycling pathway via GA. Decreasing pH in the EE, MVB/LE, and lysosome are indicated by different shades of blue. EE, early endosome; ER, endoplasmatic reticulum; GA, Golgi apparatus; L, lysosome; MVB/LE, multivesicular body/late endosome; N, nucleus.