Background: The enamel layerof kallikrein 4 (Klk4)-null mice has a normal thickness and a decussating pattern of enamel rods, but it contains residual enamel proteins, is less highly mineralized, and fractures in its deepest part just above the dentino-enamel junction (DEJ). The plane of fracture is puzzling because the deepest enamel is deposited earliest and, through the action of the secretory stage enamel protease (Mmp20), is the most mature part of the enamel layer at the time of the onset of Klk4 expression. Objectives: To characterize the planes of fracture in Mmp20- and Klk4-null mice and to localize Klk4 expression in developing teeth. Methods:Klk4- and Mmp20-null mice were sacrificed at 7 weeks and their mandibular incisors were characterized by scanning electron microscopy. Klk4+/lacZ mice were mated with Klk4+/lacZ mice. Offspring were genotyped by polymerase chain reaction. Klk4+/+, Klk4+/lacZ, and Klk4lacZ/lacZ (null) littermates on postnatal days 5, 8, 11, and 14 were processed for β-galactosidase histochemistry. Results: The enamel layer fractures at the DEJ in Mmp20-null mice, and fractures occur in enamel above the DEJ in Klk4-null mice. Klk4 is not expressed by secretory-stage ameloblasts, murine odontoblasts beneath the secretory stage, or maturation-stage ameloblasts. Klk4 is specifically expressed by transition and maturation-stage ameloblasts. Conclusions: The breakage of enamel near the DEJ in Klk4-null mice is not due to a failure of odontoblasts to express Klk4, but it relates to a progressive hypomineralization of enamel with depth.

Keywords:
Scanning electron microscopy, lacZ histostaining, Amelogenesis, Proteases, Teeth, Ameloblasts
1.
Bartlett, J.D., J.P. Simmer (1999) Proteinases in developing dental enamel. Crit Rev Oral Biol Med 10: 425–441.
2.
Begue-Kirn, C., P.H. Krebsbach, J.D. Bartlett, W.T. Butler (1998) Dentin sialoprotein, dentin phosphoprotein, enamelysin and ameloblastin: tooth-specific molecules that are distinctively expressed during murine dental differentiation. Eur J Oral Sci 106: 963–970.
3.
Beniash, E., Z. Skobe, J.D. Bartlett (2006) Formation of the dentino-enamel interface in enamelysin (MMP-20)-deficient mouse incisors. Eur J Oral Sci 114(suppl 1): 24–29, discussion 39–41, 379.
4.
Caterina, J.J., Z. Skobe, J. Shi, Y. Ding, J.P. Simmer, H. Birkedal-Hansen, J.D. Bartlett (2002) Enamelysin (matrix metalloproteinase 20)-deficient mice display an amelogenesis imperfecta phenotype. J Biol Chem 277: 49598–49604.
5.
Chun, Y.H., Y. Yamakoshi, F. Yamakoshi, M. Fukae, J.C. Hu, J.D. Bartlett, J.P. Simmer (2010) Cleavage site specificity of MMP-20 for secretory-stage ameloblastin. J Dent Res 89: 785–790.
6.
Dymecki, S. (1996) A modular set of Flp, FRT and lac Z fusion vectors for manipulating genes by site-specific recombination. Gene 171: 197–201.
7.
Fejerskov, O., A. Thylstrup (1986) Dental enamel; in Mjor, I., O. Fejerskov (eds): Human Oral Embryology and Histology. Copenhagen, Munksgaard, pp 50–89.
8.
Fukae, M., T. Tanabe, T. Nagano, H. Ando, Y. Yamakoshi, M. Yamada, J.P. Simmer, S. Oida (2002) Odontoblasts enhance the maturation of enamel crystals by secreting EMSP1 at the enamel-dentin junction. J Dent Res 81: 668–672.
9.
Hart, P.S., T.C. Hart, M.D. Michalec, O.H. Ryu, D. Simmons, S. Hong, J.T. Wright (2004) Mutation in kallikrein 4 causes autosomal recessive hypomaturation amelogenesis imperfecta. J Med Genet 41: 545–549.
10.
Hu, J.C., Y. Hu, C.E. Smith, M.D. McKee, J.T. Wright, Y. Yamakoshi, P. Papagerakis, G.K. Hunter, J.Q. Feng, F. Yamakoshi, J.P. Simmer (2008) Enamel defects and ameloblast- specific expression in Enam knock-out/lacz knock-in mice. J Biol Chem 283: 10858–10871.
11.
Hu, J.C., O.H. Ryu, J.J. Chen, T. Uchida, K. Wakida, C. Murakami, H. Jiang, Q. Qian, C. Zhang, V. Ottmers, J.D. Bartlett, J.P. Simmer (2000a) Localization of EMSP1 expression during tooth formation and cloning of mouse cDNA. J Dent Res 79: 70–76.
12.
Hu, J.C., X. Sun, C. Zhang, S. Liu, J.D. Bartlett, J.P. Simmer (2002) Enamelysin and kallikrein-4 mRNA expression in developing mouse molars. Eur J Oral Sci 110: 307–315.
13.
Hu, J.C., C. Zhang, X. Sun, Y. Yang, X. Cao, O. Ryu, J.P. Simmer (2000b) Characterization of the mouse and human PRSS17 genes, their relationship to other serine proteases, and the expression of PRSS17 in developing mouse incisors. Gene 251: 1–8.
14.
Iwata, T., Y. Yamakoshi, J.C. Hu, I. Ishikawa, J.D. Bartlett, P.H. Krebsbach, J.P. Simmer (2007) Processing of ameloblastin by MMP-20. J Dent Res 86: 153–157.
15.
Kim, J.W., J.P. Simmer, T.C. Hart, P.S. Hart, M.D. Ramaswami, J.D. Bartlett, J.C. Hu (2005) MMP-20 mutation in autosomal recessive pigmented hypomaturation amelogenesis imperfecta. J Med Genet 42: 271–275.
16.
Lee, S.K., F. Seymen, H.Y. Kang, K.E. Lee, K. Gencay, B. Tuna, J.W. Kim (2010) MMP20 hemopexin domain mutation in amelogenesis imperfecta. J Dent Res 89: 46–50.
17.
Meredith, R.W., J. Gatesy, J. Cheng, M.S. Springer (2010) Pseudogenization of the tooth gene enamelysin (MMP20) in the common ancestor of extant baleen whales. Proc Biol Sci, E-pub ahead of print.
18.
Nagano, T., A. Kakegawa, Y. Yamakoshi, S. Tsuchiya, J.C. Hu, K. Gomi, T. Arai, J.D. Bartlett, J.P. Simmer (2009) Mmp-20 and Klk4 cleavage site preferences for amelogenin sequences. J Dent Res 88: 823–828.
19.
Ozdemir, D., P.S. Hart, O.H. Ryu, S.J. Choi, M. Ozdemir-Karatas, E. Firatli, N. Piesco, T.C. Hart (2005) MMP20 active-site mutation in hypomaturation amelogenesis imperfecta. J Dent Res 84: 1031–1035.
20.
Papagerakis, P., H.K. Lin, K.Y. Lee, Y. Hu, J.P. Simmer, J.D. Bartlett, J.C. Hu (2008) Premature stop codon in MMP20 causing amelogenesis imperfecta. J Dent Res 87: 56–59.
21.
Ryu, O.H., A.G. Fincham, C.C. Hu, C. Zhang, Q. Qian, J.D. Bartlett, J.P. Simmer (1999) Characterization of recombinant pig enamelysin activity and cleavage of recombinant pig and mouse amelogenins. J Dent Res 78: 743–750.
22.
Ryu, O., J.C. Hu, Y. Yamakoshi, J.L. Villemain, X. Cao, C. Zhang, J.D. Bartlett, J.P. Simmer (2002) Porcine kallikrein-4 activation, glycosylation, activity, and expression in prokaryotic and eukaryotic hosts. Eur J Oral Sci 110: 358–365.
23.
Simmer, J.P., Y. Hu, R. Lertlam, Y. Yamakoshi, J.C. Hu (2009) Hypomaturation enamel defects in Klk4 knockout/LacZ knockin mice. J Biol Chem 284: 19110–19121.
24.
Simmer, J.P., X. Sun, Y. Yamada, C.H. Zhang, J.D. Bartlett, J.C.-C. Hu (2004) Enamelysin and kallikrein-4 expression in the mouse incisor; in Kobayashi, I., H. Ozawa (eds): Biomineralization: Formation, Diversity, Evolution and Application – Proceedings of the 8th International Symposium on Biomineralization, Niigata, Japan, September 25–28, 2001. Hadano, Tokai University Press, pp 348–352.
25.
Smith, C.E., A.S. Richardson, Y. Hu, J.D. Bartlett, J.C.-C. Hu, J.P. Simmer (2011): Effects of loss of function of kallikrein 4 on mineralization of enamel: comparison to mice lacking matrix metalloproteinase 20. J Biol Chem, E-pub ahead of print, PMID: 21454549.
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2011
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