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Cellular and Molecular Life Sciences :... Jan 2007Proteins of the developing enamel matrix include amelogenin, ameloblastin and enamelin. Of these three proteins amelogenin predominates. Protein-protein interactions are... (Comparative Study)
Comparative Study
Proteins of the developing enamel matrix include amelogenin, ameloblastin and enamelin. Of these three proteins amelogenin predominates. Protein-protein interactions are likely to occur at the ameloblast Tomes' processes between membrane-bound proteins and secreted enamel matrix proteins. Such protein-protein interactions could be associated with cell signaling or endocytosis. CD63 and Lamp1 are ubiquitously expressed, are lysosomal integral membrane proteins, and localize to the plasma membrane. CD63 and Lamp1 interact with amelogenin in vitro. In this study our objective was to study the molecular events of intercellular trafficking of an exogenous source of amelogenin, and related this movement to the spatiotemporal expression of CD63 and Lamp1 using various cell lineages. Exogenously added amelogenin moves rapidly into the cell into established Lamp1-positive vesicles that subsequently localize to the perinuclear region. These data indicate a possible mechanism by which amelogenin, or degraded amelogenin peptides, are removed from the extracellular matrix during enamel formation and maturation.
Topics: Amelogenesis; Amelogenin; Animals; Antigens, CD; Biological Transport; Cell Line; DNA Primers; Dogs; Fluorescent Antibody Technique; Genetic Vectors; Green Fluorescent Proteins; Humans; Immunohistochemistry; Lysosomal-Associated Membrane Protein 1; Mice; Platelet Membrane Glycoproteins; Reverse Transcriptase Polymerase Chain Reaction; Tetraspanin 30; Transport Vesicles
PubMed: 17187173
DOI: 10.1007/s00018-006-6429-4 -
Ecotoxicology and Environmental Safety Oct 2022Sulphur dioxide (SO) and fluoride are among the most common environmental pollutants affecting human health, and both co-exist in areas predominantly consuming coal. It...
Sulphur dioxide (SO) and fluoride are among the most common environmental pollutants affecting human health, and both co-exist in areas predominantly consuming coal. It is vital to analyse the combined toxicity of SO and fluoride, and their effects on health and the underlying mechanisms of their co-exposure have not yet been adequately assessed. In the present study, we used ICR mice and LS8 cells to investigate the toxicity of SO and fluoride exposure to the enamel, alone or in combination. Factorial design analysis was used to reveal the combined toxicity in vitro and in vivo. Co-exposure to SO and fluoride exacerbated enamel injury, resulting in more severe hypomineralization of incisor, and enamel structure disorders in mice, and could induce the accumulation of protein residue in the matrix of the enamel. Amelogenin expression was increased upon exposure to SO and fluoride, but enamel matrix proteases were not affected. Consistent with our in vivo results, co-exposure of SO and fluoride aggravated amelogenin expression in LS8 cells, and increased the YAP and RUNX2 levels. Co-exposure to SO and fluoride resulted in greater toxicity than individual exposure, both in vitro and in vivo, indicating that residents of areas exposed to SO and fluoride may have an increased risk of developing enamel damage.
Topics: Amelogenin; Animals; Coal; Core Binding Factor Alpha 1 Subunit; Environmental Pollutants; Fluorides; Humans; Incisor; Mice; Mice, Inbred ICR; Peptide Hydrolases; Signal Transduction; Sulfur Dioxide; Up-Regulation; YAP-Signaling Proteins
PubMed: 36155332
DOI: 10.1016/j.ecoenv.2022.114106 -
JOM (Warrendale, Pa. : 1989) Jun 2021Amelogenin is the most abundant matrix protein guiding hydroxyapatite formation in enamel, the durable bioceramic tissue that covers vertebrate teeth. Here, we sought to...
Amelogenin is the most abundant matrix protein guiding hydroxyapatite formation in enamel, the durable bioceramic tissue that covers vertebrate teeth. Here, we sought to refine structure-function for an amelogenin domain based on data showing a 42 amino acid amelogenin-derived peptide (ADP7) mimicked formation of hydroxyapatite similar to that observed for the full-length mouse 180 amino acid protein. In mice, we used CRISPR-Cas9 to express only ADP7 by the native amelogenin promoter. Analysis revealed ADP7 messenger RNA expression in developing mouse teeth with the formation of a thin layer of enamel. , ADP7 peptide partially replaced the function of the full-length amelogenin protein and its several protein isoforms. Protein structure-function relationships identified through assays can be deployed in whole model animals using CRISPR-Cas9 to validate function of a minimal protein domain to be translated for clinical use as an enamel biomimetic.
PubMed: 34456537
DOI: 10.1007/s11837-021-04687-x -
Journal of Structural Biology Dec 2020Amelogenin, a protein critical to enamel formation, is presented as a model for understanding how the structure of biomineralization proteins orchestrate biomineral... (Review)
Review
Amelogenin, a protein critical to enamel formation, is presented as a model for understanding how the structure of biomineralization proteins orchestrate biomineral formation. Amelogenin is the predominant biomineralization protein in the early stages of enamel formation and contributes to the controlled formation of hydroxyapatite (HAP) enamel crystals. The resulting enamel mineral is one of the hardest tissues in the human body and one of the hardest biominerals in nature. Structural studies have been hindered by the lack of techniques to evaluate surface adsorbed proteins and by amelogenin's disposition to self-assemble. Recent advancements in solution and solid state nuclear magnetic resonance (NMR) spectroscopy, atomic force microscopy (AFM), and recombinant isotope labeling strategies are now enabling detailed structural studies. These recent studies, coupled with insights from techniques such as CD and IR spectroscopy and computational methodologies, are contributing to important advancements in our structural understanding of amelogenesis. In this review we focus on recent advances in solution and solid state NMR spectroscopy and in situ AFM that reveal new insights into the secondary, tertiary, and quaternary structure of amelogenin by itself and in contact with HAP. These studies have increased our understanding of the interface between amelogenin and HAP and how amelogenin controls enamel formation.
Topics: Amelogenin; Amino Acid Sequence; Animals; Biomineralization; Dental Enamel Proteins; Durapatite; Humans; Hydrogen-Ion Concentration; Protein Conformation
PubMed: 32979496
DOI: 10.1016/j.jsb.2020.107630 -
Journal of Dental Research Apr 2011Gene expression profiles of human ameloblastoma microdissected cells were characterized with the purpose of identifying genes and their protein products that could be... (Comparative Study)
Comparative Study
Gene expression profiles of human ameloblastoma microdissected cells were characterized with the purpose of identifying genes and their protein products that could be targeted as diagnostic and prognostic markers as well as for potential therapeutic interventions. Five formalin-fixed, decalcified, paraffin-embedded samples of ameloblastoma were subjected to laser capture microdissection, linear mRNA amplification, and hybridization to oligonucleotide human 41,000 RNA arrays and compared with universal human reference RNA, to determine the gene expression signature. Assessment of the data by Significance Analysis of Microarrays (SAM) and cluster analysis showed that 38 genes were highly expressed (two-fold increase) in all samples, while 41 genes were underexpressed (two-fold reduction). Elements of the sonic hedgehog pathway and Wingless type MMTV integration site family were validated by immunohistochemistry. We have identified the expression of multiple genes and protein products that could serve as potential diagnostic, prognostic, and therapeutic targets.
Topics: Ameloblastoma; Amelogenin; Biomarkers, Tumor; Calbindin 2; Dental Enamel Proteins; Extracellular Matrix Proteins; Gene Expression Profiling; Genomics; Hedgehog Proteins; Humans; Kallikreins; Lasers, Semiconductor; Matrix Metalloproteinase 20; Microdissection; Neoplasm Proteins; Nucleic Acid Hybridization; Oligonucleotide Array Sequence Analysis; Patched Receptors; Proteins; RNA, Messenger; RNA, Neoplasm; Receptors, Cell Surface; Receptors, G-Protein-Coupled; S100 Calcium Binding Protein G; Smoothened Receptor; Wnt Proteins
PubMed: 21282726
DOI: 10.1177/0022034510391791 -
Frontiers in Physiology 2014Epithelially-derived ameloblasts secrete extracellular matrix proteins including amelogenin, enamelin, and ameloblastin. Complex intermolecular interactions among these...
Epithelially-derived ameloblasts secrete extracellular matrix proteins including amelogenin, enamelin, and ameloblastin. Complex intermolecular interactions among these proteins are believed to be important in controlling enamel formation. Here we provide in vitro and in vivo evidence of co-assembly and co-localization of ameloblastin with amelogenin using both biophysical and immunohistochemical methods. We performed co-localization studies using immunofluorescence confocal microscopy with paraffin-embedded tissue sections from mandibular molars of mice at 1, 5, and 8 days of age. Commercially-available ameloblastin antibody (M300) against mouse ameloblastin residues 107-407 and an antibody against full-length recombinant mouse (rM179) amelogenin were used. Ameloblastin-M300 clearly reacted along the secretory face of ameloblasts from days 1-8. Quantitative co-localization was analyzed (QCA) in several configurations by choosing appropriate regions of interest (ROIs). Analysis of ROIs along the secretory face of ameloblasts revealed that at day 1, very high percentages of both the ameloblastin and amelogenin co-localized. At day 8 along the ameloblast cells the percentage of co-localization remained high for the ameloblastin whereas co-localization percentage was reduced for amelogenin. Analysis of the entire thickness on day 8 revealed no significant co-localization of amelogenin and ameloblastin. With the progress of amelogenesis and ameloblastin degradation, there was a segregation of ameloblastin and co-localization with the C-terminal region decreased. CD spectra indicated that structural changes in ameloblastin occurred upon addition of amelogenin. Our data suggest that amelogenin-ameloblastin complexes may be the functional entities at the early stage of enamel mineralization.
PubMed: 25120489
DOI: 10.3389/fphys.2014.00274 -
Clinical and Experimental Dental... Apr 2022We set out to investigate whether a hybrid stem-like p-EMT phenotype develops during murine molar enamel development in vivo.
OBJECTIVES
We set out to investigate whether a hybrid stem-like p-EMT phenotype develops during murine molar enamel development in vivo.
SETTING AND SAMPLE POPULATION
Histology specimens incorporating molar tooth buds harvested from mice at post-natal day 4 (P4) were included in our studies.
MATERIALS AND METHODS
We employed double immunofluorescence staining to analyze the simultaneous expression of the epithelial marker E-cadherin and the mesenchymal marker N-cadherin in histology specimens with tooth buds harvested from P4 mice. Moreover, we evaluated the expression of the core master stem cell markers Oct4 and Sox2, as well as the mature ameloblast marker amelogenin.
RESULTS
Here we document the co-expression of E-cadherin and N-cadherin in a sub-population of pre-ameloblasts in the inner enamel epithelium suggestive of the presence of a hybrid epithelial/mesenchymal phenotype resulting from p-EMT. Moreover, the core stem cell factors Oct4 and Sox2 colocalized with E-cadherin expressing pre-ameloblasts, whereas the mesenchymal marker N-cadherin was expressed specifically by amelogenin-positive mature secretory ameloblasts.
CONCLUSIONS
The differentiation of E-cadherin-positive pre-ameloblasts towards N-cadherin-positive mature secretory ameloblasts transition through a previously unidentified epithelial/mesenchymal stage derived through p-EMT, co-expressing the master transcription factors Oct4 and Sox2.
Topics: Ameloblasts; Amelogenin; Animals; Cadherins; Dental Enamel; Epithelial-Mesenchymal Transition; Mice
PubMed: 35182458
DOI: 10.1002/cre2.543 -
Frontiers in Physiology 2016
PubMed: 27667974
DOI: 10.3389/fphys.2016.00374 -
Journal of Dental Research Dec 2021The nanofibrous nature and its intricate structural organization are the basis for the extraordinary ability of sound enamel to outlive masticatory forces at minimal... (Review)
Review
The nanofibrous nature and its intricate structural organization are the basis for the extraordinary ability of sound enamel to outlive masticatory forces at minimal failure rates. Apatite nanofibers of several hundreds of micrometers to possibly millimeters in length originate during the secretory stage of amelogenesis as 2-nm-thin and 15-nm-wide ribbons that develop and grow in length under the guidance of a dynamic mixture of specialized proteins, the developing enamel matrix (DEM). A critical role in the unidirectional and oriented growth of enamel mineral ribbons has been attributed to amelogenin, the major constituent of the DEM. This review elaborates on recent studies on the ability of ribbon-like assemblies of amelogenin to template the formation of an amorphous calcium phosphate precursor that transforms into apatite mineral ribbons similar to the ones observed in developing enamel. A mechanistic model of the biological processes that drive biomineralization in enamel is presented in the context of a comparative analysis of enamel mouse models and earlier structural data of the DEM emphasizing a regulatory role of the matrix metalloproteinase 20 in mineral deposition and the involvement of a process-directing agent for the templated mineral growth directed by amelogenin nanoribbons.
Topics: Amelogenesis; Amelogenin; Animals; Dental Enamel; Dental Enamel Proteins; Matrix Metalloproteinase 20; Mice; Nanotubes, Carbon
PubMed: 34009057
DOI: 10.1177/00220345211012925 -
Journal of Oral Biosciences 2011Before a tooth erupts into the oral cavity, the mineralized enamel and dentin layers begin to develop. During these early stages of enamel formation, an abundant group...
Before a tooth erupts into the oral cavity, the mineralized enamel and dentin layers begin to develop. During these early stages of enamel formation, an abundant group of proteins known as amelogenins are secreted by ameloblast cells within the developing tooth. These proteins are required for the enamel layer to reach its normal thickness and attain its intricate structure. Human patients with amelogenin gene mutations have a condition referred to as amelogenesis imperfecta, and we have analyzed human gene defects so that we can recreate them in mice. We have generated mice with a null amelogenin mutation where no amelogenin is produced, mice that over-express normal and mutated amelogenins, and over-expressors have been mated to null mice for rescue experiments. Because there are at least 15 messages that are alternatively spliced from a single amelogenin primary RNA transcript, these approaches have begun to reveal the functions of individual amelogenin proteins during enamel development. Finally, amelogenins are processed by carefully regulated proteolytic digestion leading to many additional amelogenin peptides and it is likely that protein function is altered during this developmental process. We have also had some surprises, as one of our mouse models develops odontogenic tumors, and we know now that some of the amelogenins are expressed in other regions of the body outside of the oral cavity, and may have a role in signal transduction.
PubMed: 22215951
DOI: 10.2330/joralbiosci.53.248