SP Transcription Family Involve In Tooth Development ? Ivan Arie Wahyudi1 1 Department of Dental Biomedical Sciences Faculty of DentistryUniversitas Gadjah MadaYogyakarta-Indonesia Abstract Specificity Protein (SP) family members are tissue specific transcription factors. They regulate a wide range of cellular function including cells growth, apoptosis, differentiation and tumor formation. This family composed of more than 25 member proteins, which contain a DNA-binding domain very well conserve between all members with three tandem zinc fingers of the C2H2 type in their Cterminal region. Recently, it has reported in mammalian genome, there are nine Sp genes (Sp1-Sp9). Some previous study reported Sp3 knockout mice have enamel/dentin layer defect. Sp4 mRNA express in E13- E16 of WT mouse incisor. Sp7 mRNA express in at 15, 17 days of post coitum and P1 (postnatal day 1). Sp6 has another name epiprofin, I also worked with this genes. Sp6 mRNA expressed in early stage of tooth development to the secretory stage of ameloblast. It also reported Sp6 weakly expressed in mesenchymal odontoblast of the incisor. Sp6 deficient mice reported delay tooth development. SP family members play an important role in tooth development. Keyword: SP family, transcription factor, tooth development SP Transcription Family Terlibat dalam Perkembangan Gigi? Abstrak Specificity Protein (SP)merupakan faktor transkripsi pada jaringanspesifik.Protein ini mengaturberbagaifungsi seltermasuk pertumbuhansel,apoptosis,diferensiasi danpembentukan tumor.Keluarga protein initerdiri darilebih dari 25 anggota, yangmengandung domainpengikat DNA yang sangat miripdisetiap anggotapada tiga tandem zinc fingers dari tipeC2H2di bagian C-terminal.Saat initelahdilaporkan terdapat sembilangenSp(Sp1-Sp9) dalam genom mamalia. Beberapapenelitian sebelumnyamelaporkanSp3tikus KOmenyebabkan kerusakan pada lapisan email/dentin.mRNASp4terlihat pada E13-E16 WTgigi incisivus tikus. Sp7mRNAterlihat pada hari ke 15, 17pascacoitumdanP1(pascakelahiran haripertama).Sp6memiliki nama lain yaitu Ivan Arie Wahyudi epiprofin, saya juga bekerja dengan gen ini.Sp6mRNAterlihat padatahap awal perkembangan gigi ke tahap keluar nya Korespondensi : Ivan Ari Wahyudi, Department of Dental Biomedical Sciences, Faculty of DentistryUniversitas Gadjah MadaYogyakarta-Indonesia, e-mail: ivanocovic@yahoo.com IDJ, Volume 1,No. 1, Tahun 2012 83 : SP Transcription Family Involve In Tooth Development ? gigi dari ameloblast. Juga dilaporkan Sp6 terlihat agak lemah di odontoblast mesenchymal pada gigi incisivus. Defisiensi gen Sp6 pada tikus dilaporkan bahwa terjadi penundaan pertumbuhan gigi. SP memainkan peran penting dalam perkembangan gigi. Kata kunci: keluarga SP, factor trankripsi, perkembangan gigi Introduction Regulation of transcription is an important to explore the question of how DNA sequence information is used appropriately by mammalian cells. Using an array of biomolecular tools, we can identify all the genes that encode transcription factor belonging the certain class and also study their biological function 1. The SP/KLF transcription factor family contains over 25 members sharing a DNA- binding domain composed of three zinc fingers motif of the C2H2 type at the C-terminus and binds to GGGCGG motifs or related GC-rich sequence. This family comprises of nine members (Sp1-Sp9) in mammals 1-2 . Each members are located adjacent to a HOX gene cluster 3. A number of transcription factors control tooth development in order to form unique structures specialized for tooth function especially for shapes and sizes. The developing tooth is a good model for studying the aspects of molecular and genetic on mammalian tooth development 3. During tooth development epithelial and mesenchyme interaction is thought very important. Early signals for tooth development arise in the oral ectoderm, appearing as thickening of the dental lamina. The dental lamina invaginates into the underlying neural crest-derived mesenchyme to form the tooth bud. The dental epithelial cells proliferate to form a double layer cap that is called the enamel organ. After the cap stage, the tooth germ progresses to the bell and late bell stages before the tooth erupts into the oral cavity. All these stages are regulated not only by cytokines, such as bone morphogenetic proteins (BMPs), sonic hedgehogs (Shhs), fibroblast growth factors (FGFs), and wingless (Wnts), but also by extracellular matrices. The deletion of these gene functions results in the arrest of tooth development 4-7. The SP-Family Ivan Arie Wahyudi: SP Transcription Family Involve In Tooth Development ? IDJ, Volume 1,No. 1, Tahun 2012 The first identified member of this family is termed SP1, forSpecificity Protein, in the 21bprepeats of the simian virus ( SV40) early promoter. DNA binding domain of SP1 iscomposed of three zinc fingers of the classical Cys2– His2 type. The first four members of the Sp-family (Sp1-4) are more closely related to each other than to Sp5-8. Sp 1-4 contain an N-terminal activation domain and a C-terminal DNA binding domain. Sp5-8 proteins are shorter, lacking the N-terminal activation domain. This may decrease transcriptional activation potential. The overview of each Sp-family domain is performing in the figure 1. 87 IDJ, Volume 1,No. 1, Tahun 2012 Figure 1. The domain structure of Sp-family transcription factors (Modified from Bowman P. and Philipsen S., 2002)8 At C-terminal, three black boxes are C2H2 type zinc finger motifs for DNA binding. This domain is a common feature of Spfamily. At N-terminal is a transcriptional regulatory domain. The red box is buttonhead box, it may contribute to transactivation 3 Regulation of Sp transcription factors in tooth development Both of Sp1 and Sp3 have been reported to exhibit ubiquitous expression and dental epthelium. Sp2 expression has been observed in a number of cell lines, whereas Sp4 expression, currently reported expressed in CNS, liver, lung, kidney, Ivan Arie Wahyudi heart, gonads, intestine and also in dental papilla and dental sac 3,9 . Sp7 was observed play roles as a bone specific transcription factor required for osteoblast differentiation and bone formation (10). Sp5, Sp8, and Sp9 are expressed in specific tissue and developing stages 9. Sp6 that correspond to epiprofin. This gene has reported by some researchers play roles in teeth developing, caudal neuropore, limb bud, hair follicles, skin and dental epithelium 11. I also worked using this Sp6 gene, our laboratory has found that Sp6 promotes amelogenesis through inhibition of follistatin gene expression while follistatin is responsiblefor the formation of an enamel-free area in the mouse incisor and molar by inhibiting ameloblast differentiation 12 . For overview of the expression pattern of Sp transcription factors, : SP Transcription Family Involve In Tooth Development ? including the knockout mice of each members perform in Table 1. Table 1. Expression pattern of Sp transcription factors in vertebrate embryos 9 Factors Expression Chromosomal Location Major phenotypes in knockout mice Sp1 Ubiquitous, Dental epthelium Human: 12q13.1 Mouse : 15 Growth retardation, prenatal lethality Sp2 Ubiquitous Human: 17q21.32 Mouse : 11 Growth retardation, prenatal lethality Sp3 Ubiquitous, Dental epthelium Human: 2q31 Mouse : 2 Growth retardation, Defect in tooth, lung, bone and hematopoetic Sp4 CNS, liver,lung, kidney,heart, gonads, intestine, Dental papilla, and dental sac Human: 7q15.3 Mouse : 12 Postnatal mortality, smaller body size Sp5 Mesoderm precursors, derivates posterior neuroectoderm, Human: 2q31 Mouse : 2 No morphological changes enhanced frequency of taillessness Sp6/Epip rofin Developing teeth, caudal neuropore, limb bud, hair follicles, skin and dental epithelium Human: 17q21.32 Mouse : 11 Enamel defect, supernumerary teeth, defective cups and root formation Sp7/Oste rix Developing bone and teeth Odontoblast and dental follicle cells Human: 12q13.13 Mouse : 15 Death at birth, Failure in ossification Sp8 CNS, limb buds Human: 7q15.3 Mouse : 12 Neural tube closure failure, shorter limbs Ivan Arie Wahyudi: SP Transcription Family Involve In Tooth Development ? IDJ, Volume 1,No. 1, Tahun 2012 Sp9 In specific domain of CNS, limb Human: 2q31 Mouse : 2 unknown Table 1 above showed that most of Sp-family members are correlate with tooth development. Sp1 is expressed in dental epithelium. Sp1 and Sp2 knockout mice die in embryonic stage. Sp3 is also expressed in dental epithelium. Mice lacking Sp3 showed defect of enamel, lack amelogenin, and ameloblast, impaired ossification. Sp4 is expressed Dental papilla, and dental sac 8,9. Sp4 and Sp7 are expressed in dental mesenchyme 9. Since I was working with Sp6 gene that we believed might play role in tooth development. Sp6in tooth development Sp6mRNA is expressed tooth germ 13.Expression of Sp6 is detected at the initiation stage of tooth development. Sp6 is clearly expressed in dental epithelium of dental lamina but not expressed in dental mesenchyme (another report said that Sp6 weakly expressed in mesenchymal odontoblast of the incisor) at early stage of tooth development. During the bud stage, Sp6 is expressed widely in dental epithelial cells and tooth bud develops rapidly by dental epithelial cell proliferation. At the cap stage, 89 IDJ, Volume 1,No. 1, Tahun 2012 Ivan Arie Wahyudi dental epithelial cells determine their cell fate into several lineage such as stellate reticulum and inner and outer enamel epithelium. At the bell stage, Sp6 is expressed in pre ameloblast and ameloblast 9. Figure 2 is describing the expression of Sp6 in each stage of tooth development. Figure 2.the expression of Sp6 during tooth development. The meaning of the red symbol ( ) expression Sp6 in epithelium (http:bite- it.helsinki.fi/)14 As reported Nakamura et.al., 2011, in Table 1. Sp6 knockout mice showed enamel defect, supernumerary teeth, defective cups and root formation. This result is consistence with their previous report in 2008, are showed in Figure 3, Sp6 deficient mice. : SP Transcription Family Involve In Tooth Development ? IDJ, Volume 1,No. 1, Tahun 2012 Ivan Arie Wahyudi: SP Transcription Family Involve In Tooth Development ? Figure 3. Amazing teeth phenotype in Sp6 deficient mice11 The surprising/amazing phenotype of Sp6 deficient mice. On the left side are Wild type (WT), the right side are mutant mice. At 3 weeks age, incisor and molar mutant mice were not erupted. In contras at 12 month age mutant incisors showed multiple teeth. This result strongly suggested the involvement of Sp6 in tooth development. Conclusion Tooth development is regulated by inductive interactions between the epithelium and the mesenchyme via reciprocal signalings and some cytokines are involved, Some of Sp-family have reported appear have diverse play roles in tooth development. However, the regulatory mechanism of reciprocal epithelium and mesenchyme, Sp-family, the cytokines and other signaling require further characterization. References 1. Suske G, Bruford E, Philipsen S: Mammalian SP/KLF transcription factors: bring in the family. Genomics 85: 551-556, 2005 2. Jimenez-Rojo L, Ibarretxe G, Aurrekoetxea M, de-Vega S, Nakamura T, Yamada Y, Unda F: Epiprofin/ Sp6: A new player in the regulation of tooth development. Histol Histopathol 25: 1621-1630, 2010 3. Waby, J.S, Bingle, C.D and Corfe, B.M., Post-Translation control of Sp-family transcription factors. Current Genomics 9;301-311, 2008 4. Maas R, Bei M: The genetic control of early tooth development. Crit Rev Oral Biol Med Mass 8: 4-39, 1997 5. Jernvall J, Thesleff I: Reiterative signaling and patterning during mammalian tooth morphogenesis. Mech Dev 92: 19-29, 2000 6. Thesleff I: Epithelialmesenchymal signalling regulating tooth morphogenesis. J Cell Sci 116: 1647-1648, 2003 7. Wahyudi IA, Horiguchi T, Miyoshi K, Muto T, Utami TW, Hagita H,Noma T: Isolation and characterization of mouse Specificity Protein 6 promoter. Ind J Dent Res 1: 13-26, 2010 8. Bouwman, P., H. Gollner, H. P. Elsasser, G. Eckhoff, A. Karis, F. Grosveld, S. Philipsen, and G. Suske. Transcription factor Sp3 is essential for post-natal survival and late tooth development. EMBO J. 19:655- 661, 2000 9. Nakamura T, Fukumoto S, Yamada Y. Diverse function of epiprofin in tooth development. J. Oral Biosci. 53(1): 22-30, 2011 10. Zhang C. Transcriptional regulation of bone formation by the osteoblast-specific transcription factors Osx. Journal of Orthopaedic surgery and Research 5(37): 1-7, 2010 86 IDJ, Volume 1,No. 1, Tahun 2012 11. Nakamura T, de-Vega S, Fukumoto S, Jimenez L, Unda F, Yamada Y: Transcription factor epiprofin is essential for Ivan Arie Wahyudi tooth morphogenesis by regulating epithelial cell fate and 12. Ruspita I, Miyoshi K, Muto T, Abe K, Horiguchi T, Noma T: Sp6 downregulation of follistatin gene expression in ameloblasts. J Med Invest 55: 87-98, 2008 13. Nakamura T, Unda F, de-Vega S, Vilaxa A, Fukumoto S, Yamada KM, Yamada Y: The tooth number. J Biol Chem 283: 4825- 4833, 2008 Krüppel-like factor epiprofin is expressed by epithelium of developing teeth, hair follicles, and limb buds and promotes cell proliferation. J Biol Chem 279: 626-634, 2004 14. http://bite-it.helsinki.fi/ 87