The purpose of this study was to establish transgenic mouse lines as a tool to investigate the function of amelogenin during mineralization by causing ectopic production of amelogenin and studying its effect. The mouse amelogenin (mAme) was cloned from a 16-day-old whole mouse embryo cDNA library and was determined to be "full-length" mouse amelogenin (with a complete coding region) by comparison with the mouse amelogenin reported previously by Snead et al. (1985) and Lau et al. (1992). The overexpression construct contained: (1) the rat osteocalcin (OC) promoter (1.8 kb); (2) the adenovirus splicing casettes, including introgenic (Int) sequence (0.3 kb); (3) the full-length mAme cDNA (0.8 kb); and (4) the polyadenylation signal sequence from the pSG5 mammalian expression vector. Both Southern blotting and polymerase chain-reaction (PCR) analyses were performed, by means of a specific probe and a pair of oligodeoxynucleotides to OclntmAme(A)+, respectively. The animals which showed transgene-positive in both analyses were further used to establish F1 animals. Heterozygocity was confirmed with F1 animals by PCR analysis of DNA from the F0 x FVB/N pups. Three independent transgenic F1 heterozygous lines (640t, 706t, and 708t) have now been established. The generation of F2 homozygous lines is under way. The heterozygous transgenic animals are currently being analyzed for alterations in the morphology and structure of various bone tissues.
Alfred MJ, Crawford JM, Roberts E., Thomas NST (1992). Identification of a nonsense mutation in the amelogenin gene (AMELX) in a family with X-linked amelogenesis imperfecta (AIH1). Human Genet90:413-416.
2.
Aoba T., Moreno EC, Kresak M., Tanabe T. (1989). Possible roles of partial sequences at N- and C-termini of amelogenin in protein-enamel mineral interaction. J Dent Res68:1331-1336.
3.
Aoba T.Shimoda S., Shimokawa H., Inage T. (1992). Common epitopes of mammalian amelogenins at the C-terminus and possible function roles of the corresponding domain in enamel mineralization. Calcif Tissue Int51:85-91.
4.
Baker AR, Hollingshead PG, Pitts-Meek S., Hansen S., Taylor R., Stewart TA (1992). Osteoblast-specific expression of growth hormone stimulates bone growth in transgenic mice. Molec Cell Biol12:5541-5547.
5.
Catalano-Sherman J., Palmon A., Burstein Y., Deutsch D. (1993). Amino acid sequence of a major human amelogenin protein employing Edman degradation and cDNA sequencing. J Dent Res72:1566-1572.
6.
Couwenhoven RI, Snead ML (1994). Early determination and permissive expression of amelogenin transcription during mouse mandibular first molar development . Dev Biol164:290-299.
7.
DenBesten PK, Li RS (1992). Characterization of amelogenin mRNA from secretory- and maturation-stage rat incisor enamel. Arch Oral Biol37:1097-1100.
8.
Diekwisch TG, David S., Bringas P. Jr, Santos V., Slavkin HC (1993). Antisense inhibition of AMEL translation demonstrates supramolecular controls for enamel HAP crystal growth during embryonic mouse molar development. Development117:471-482.
9.
Diekwisch TGBerman BJ, Gentner S., Slavkin HC (1995). Initial enamel crystals are not spatially associated with mineralized dentine. Cell Tissue Res279:149-167.
10.
Fincham AG, Belcourt AB, Termine JD (1982). Changing patterns of enamel matrix proteins in the developing bovine tooth. Caries Res16:64-71.
11.
Fincham AG, Moradian-Oldak J., Simmer JP, Sarte PE, Lau EC, Diekwisch TG, et al. (1994). Self-assembly of a recombinant amelogenin protein generates supramolecular structures. J Struct Biol112:103-109.
12.
Gibson CW, Golub E., Ding W., Shimokawa H., Young MF, Termine JD, et al. (1991). Identification of the leucine-rich amelogenin peptide (LRAP) as the translation product of an alternatively spliced transcript . Biochem Biophys Res Commun174:1306-1312.
13.
Glimcher MJ, Brickley-Parsons D., Levine PT (1977). Studies of enamel proteins during maturation. Calcif Tissue Res24:259-270.
14.
Huang MH, Gorman CM (1990). Intervening sequences increase efficiency of RNA 3' processing and accumulation of cytoplasmic RNA. Nucleic Acids Res18:937-947.
15.
Krebsbach PH, Harrison JR, Lichtler AC, Woody CO, Rowe DW, Kream BE (1993). Transgenic expression of COL1A1-chloramphenicol acetyltransferase fusion genes in bone: differential utilization of promoter elements in vivo and cultured cells. Molec Cell Biol13:5168-5174.
16.
Lagerstrom M., Dahl N., Nakahori Y., Nakagome Y., Backman B., Landegren U., et al. (1991). A deletion in the amelogenin gene (AMG) causes X-linked amelogenesis imperfecta (AIH1). Genomics10:971-975.
17.
Lagerstrom-Fermer M., Pettersson U., Landegren U. (1993). Molecular basis and consequences of a deletion in the amelogenin gene, analyzed by capture PCR. Genomics17:89-92.
18.
Lau EC, Simmer JP, Bringas P. Jr, Hsu DD-J , Hu CC, Zeichner-David M., et al. (1992). Alternative splicing of the mouse amelogenin primary RNA transcript contributes to amelogenin heterogeneity. Biochem Biophys Res Commun188:1253-1260.
19.
Lench NJ, Winter GB (1995). Characterization of molecular defects in X-linked amelogenesis imperfecta (AIH1). Human Mutat5:251-259.
20.
Nakamura M., Bringas P. Jr, Nanci A., Zeichner-David M., Ashdown B., Slavkin HC (1994). Translocation of enamel proteins from inner enamel epithelia to odontoblasts during mouse tooth development. Anat Rec238:383-396.
21.
Nakase T., Takaoka K., Hirakawa K., Hirota S., Takemura T., Onoue H., et al. (1994). Alterations in the expression of osteonectin, osteopontin and osteocalcin mRNAs during the development of skeletal tissues in vivo. Bone Miner26:109-122.
22.
Owen TA, Arnow M., Shalhoub V., Barone LM, Wilming L., Tassinari MS, et al. (1990). Progressive development of the rat osteoblast phenotype in vitro: Reciprocal relationships in expression of genes associated with osteoblast proliferation and differentiation during formation of the bone extracellular matrix. J Cell Physiol143:420-430.
23.
Owen TA, Arnow MS, Barone LM, Bettencourt B., Stein GS, Lian JB ( 1991). Pleiotropic effects of vitamin D on osteoblast gene expression are related to the proliferative and differentiated state of the bone cell phenotype: Dependency upon basal levels of gene expression, duration of exposure, and bone matrix competency in normal rat osteoblast cultures. Endocrinology128:1496-1504.
24.
Robinson C., Kirkham J., Briggs HD, Atkinson PJ (1982). Enamel proteins: from secretion to maturation . J Dent Res61:1490-1495.
25.
Sambrook J., Fritsch EF, Maniatis T. (1989). In: Molecular cloning: A laboratory manual. 2nd ed. Cold Spring Harbor, NY: Cold Spring Harbor Press.
26.
Sasaki T. (1990). Amelogenesis disturbances. In: Cell biology of tooth enamel formation. Vol. 14. Monographs in oral science . Myers HM, editor. Basel: Karger, pp. 159-171.
27.
Shimokawa H., Ogata Y., Sasaki S., Sobel ME, McQuillan CI, Termine JD, et al. (1987). Molecular cloning of bovine amelogenin cDNA. Adv Dent Res1:293-297.
28.
Sicher H, Bhaskar SN, editors (1972). In: Orban's oral histology and embryology . St. Louis, MO: The C.V. Mosby Company.
29.
Slavkin HC, Zeichner-David M., Snead ML, Bringas P., Bessem C., Santos V. (1984). Epithelial-mesenchymal interactions and enamel gene induction-evolutionary implications. In: Tooth enamel IV. Fearnhead RW, Suga S, editors. Amsterdam: Elsevier , pp. 167-171.
30.
Snead ME, Lau EC, Zeichner-David M., Fincham AG, Woo SLC, Slavkin HC (1985). DNA sequence for cloned cDNA for murine amelogenin reveals the amino acid sequence for enamel-specific protein. Biochem Biophys Res Commun129:812-818.
31.
Stewart TA, Clifft S., Pitts-Meek S., Martin L., Terrell TG, Liggett D., et al. (1992). An evaluation of the functions of the 22 kD, the 20 kD, and the N-terminal polypeptide forms of human growth hormone using transgenic mice. Endocrinology130:405-414.
