Spatio-Temporal Expression and Functional Analysis of miR-206 in Developing Orofacial Tissue

Author(s): Partha Mukhopadhyay, Irina Smolenkova, Dennis Warner, Michele M. Pisano, Robert M. Greene*

Journal Name: MicroRNA

Volume 8 , Issue 1 , 2019

Become EABM
Become Reviewer
Call for Editor

Graphical Abstract:


Background: Development of the mammalian palate is dependent on precise, spatiotemporal expression of a panoply of genes. MicroRNAs (miRNAs), the largest family of noncoding RNAs, function as crucial modulators of cell and tissue differentiation, regulating expression of key downstream genes.

Observations: Our laboratory has previously identified several developmentally regulated miRNAs, including miR-206, during critical stages of palatal morphogenesis. The current study reports spatiotemporal distribution of miR-206 during development of the murine secondary palate (gestational days 12.5-14.5).

Result and Conclusion: Potential cellular functions and downstream gene targets of miR-206 were investigated using functional assays and expression profiling, respectively. Functional analyses highlighted potential roles of miR-206 in governing TGFß- and Wnt signaling in mesenchymal cells of the developing secondary palate. In addition, altered expression of miR-206 within developing palatal tissue of TGFß3-/- fetuses reinforced the premise that crosstalk between this miRNA and TGFß3 is crucial for secondary palate development.

Keywords: Embryo, fetus, microRNA, miR-206, mouse, palate.

Murray JC, Schutte BC. Cleft palate: players, pathways, and pursuits. J Clin Invest 2004; 113: 1676-8.
Greene RM, Pisano MM. Palate morphogenesis: current understanding and future directions. Birth Defects Res C Embryo Today 2010; 90: 133-54.
Bush JO, Jiang R. Palatogenesis: morphogenetic and molecular mechanisms of secondary palate development. Development 2012; 139: 231-43.
Smith TM, Lozanoff S, Iyyanar PP, Nazarali AJ. Molecular signaling along the anterior-posterior axis of early palate development. Front Physiol 2013; 3: 488.
Potter AS, Potter SS. Molecular anatomy of palate development. PLoS One 2015; 10: e0132662.
Cuervo R, Valencia C, Chandraratna RA, Covarrubias L. Programmed cell death is required for palate shelf fusion and is regulated by retinoic acid. Dev Biol 2002; 245: 145-56.
Sohn WJ, Ji YR, Kim HS, et al. Rgs19 regulates mouse palatal fusion by modulating cell proliferation and apoptosis in the MEE. Mech Dev 2012; 129: 244-54.
Nakajima A, Tanaka E, Ito Y, et al. The expression of TGF-beta3 for epithelial-mesenchyme transdifferentiated MEE in palatogenesis. J Mol Histol 2010; 41: 343-55.
Jalali A, Zhu X, Liu C, Nawshad A. Induction of palate epithelial mesenchymal transition by transforming growth factor beta3 signaling. Dev Growth Differ 2012; 54: 633-48.
Martinez-Alvarez C, Tudela C, Perez-Miguelsanz J, O’Kane S, Puerta J, Ferguson MW. Medial edge epithelial cell fate during palatal fusion. Dev Biol 2000; 220: 343-57.
Bartel DP. MicroRNAs: target recognition and regulatory functions. Cell 2009; 136: 215-33.
Ebert MS, Sharp PA. Roles for microRNAs in conferring robustness to biological processes. Cell 2012; 149: 515-24.
Brennecke J, Stark A, Russell R, Cohen S. Principles of microRNA-target recognition. PLoS Biol 2005; 3: e85.
Newman M, Hammond S. Emerging paradigms of regulated microRNA processing. Genes Dev 2010; 24: 1086-92.
Hausser J, Syed AP, Bilen B, Zavolan M. Analysis of CDS-located miRNA target sites suggests that they can effectively inhibit translation. Genome Res 2013; 23: 604-15.
Vasudevan S, Tong Y, Steitz JA. Switching from repression to activation: microRNAs can up-regulate translation. Science 2007; 318: 1931-4.
Du WW, Yang W, Xuan J, Gupta S, et al. Reciprocal regulation of miRNAs and piRNAs in embryonic development. Cell Death Differ 2016; 23: 1458-70.
Sheehy N, Cordes K, White M, Ivey K, Srivastava D. The neural crest-enriched microNA miR-452 regulates epithelial-mesenchymal signaling in the first pharyngeal arch. Development 2010; 137: 4307-16.
Nie X, Wang Q, Jiao K. Dicer activity in neural crest cells is essential for craniofacial organogenesis and pharyngeal arch artery morphogenesis. Mech Dev 2011; 128: 200-7.
Barritt L, Miller J, Scheetz L, et al. Conditional deletion of the human ortholog gene Dicer1 in Pax2-Cre expression domain impairs orofacial development. Indian J Hum Genet 2012; 18: 310-9.
Tavares AL, Artinger KB, Clouthier DE. Regulating craniofacial development at the 3′ end: microRNAs and their function in facial morphogenesis. Curr Top Dev Biol 2015; 115: 335-75.
Eberhart JK, He X, Swartz ME, et al. MicroRNA Mirn140 modulates Pdgf signaling during palatogenesis. Nat Genet 2008; 40: 290-8.
Mukhopadhyay P, Brock G, Pihur V, Pisano MM, Greene RM. Developmental microRNA expression profiling of murine embryonic orofacial tissue. Birth Defects Res A 2010; 88: 511-34.
Seelan R, Warner D, Mukhopadhyay P, et al. Epigenetic analysis of laser capture microdissected fetal epithelia. Anal Biochem 2013; 442: 68-74.
Warner DR, Ding J, Mukhopadhyay P, et al. Temporal expression of miRNAs in laser capture microdissected palate medial edge epithelium from TGFβ3-null mouse fetuses. miRNA 2015. 4: 64-71.
Ding H, Hooper J, Batzel P, et al. MicroRNA profiling during craniofacial development: potential roles for Mir23b and Mir133b. Front Physiol 2016; 7: 1-16.
Powder K, Ku Y, Brugmann S, et al. A cross-species analysis of microRNAs in the developing avian face. PLoS One 2012; 7: e35111.
Gao S, Moreno M, Eliason S, et al. TBX1 protein interactions and microRNA-96-5p regulation controls cell proliferation during craniofacial and dental development: implications for 22q11.2 deletion syndrome. Hum Mol Genet 2015; 24: 2330-48.
Abba M, Patil N, Leupold J, Allgayer H. MicroRNA regulation of epithelial to mesenchymal transition. J Clin Med 2016; 5: 1-20.
Li L, Meng T, Jia Z, Zhu G, Shi B. Single nucleotide polymorphism associated with nonsyndromic cleft palate influences the processing of miR-140. Am J Med Genet A 2010; 152A: 856-62.
Ma L, Xu M, Li D, et al. A miRNA-binding-site SNP of MSX1 is associated with NSOC susceptibility. J Dent Res 2014; 93: 559-64.
Kannu P, Campos-Xavier AB, Hull D, Martinet D, Ballhausen D, Bonafé L. Post-axial polydactyly type A2, overgrowth and autistic traits associated with a chromosome 13q31.3 microduplication encompassing miR-17-92 and GPC5. Eur J Med Genet 2013; 56: 452-7.
Tassano E, Di Rocco M, Signa S, Gimelli G. De novo 13q31.1-q32.1 interstitial deletion encompassing the miR-17-92 cluster in a patient with Feingold syndrome-2. Am J Med Genet 2013; 161A: 894-6.
Proetzel G, Pawlowski SA, Wiles MV, et al. Transforming growth factor-beta 3 is required for secondary palate fusion. Nat Genet 1995; 11: 409-14.
Taya Y, O’Kane S, Ferguson MW. Pathogenesis of cleft palate in TGF-beta3 knockout mice. Development 1999; 126: 869-79.
Mukhopadhyay P, Seelan RS, Rezzoug F, et al. Determinants of orofacial clefting I: effects of 5-Aza-2′-deoxycytidine on cellular processes and gene expression during development in the first branchial arch. Reprod Toxicol 2017; 67: 85-99.
Kloosterman WP, Wienholds E, De Bruijn E, Kauppinen S, Plasterk RH. In situ detection of miRNAs in animal embryos using LNA-modified oligonucleotide probes. Nat Methods 2006; 3: 27-9.
Warner DR, Webb CL, Greene RM, Pisano MM. Altered signal transduction in Folr1-/- mouse embryo fibroblasts. Cell Biol Int 2011; 35: 1253-9.
Warner DR, Pisano MM, Greene RM. Functional analysis of CBP/p300 in embryonic orofacial mesenchymal cells. J Cell Biochem 2006; 99: 1374-9.
Anderson C, Catoe H, Werner R. MIR-206 regulates connexin43 expression during skeletal muscle development. Nucleic Acids Res 2006; 34: 5863-71.
Wang R, Hu Y, Song G, et al. MiR-206 regulates neural cells proliferation and apoptosis via Otx2. Cell Physiol Biochem 2012; 29: 381-90.
Zhang Y, Lei W, Yan W, et al. microRNA-206 is involved in survival of hypoxia preconditioned mesenchymal stem cells through targeting Pim-1 kinase. Stem Cell Res Ther 2016; 7: 61.
Xiao H, Xiao W, Cao J, et al. miR-206 functions as a novel cell cycle regulator and tumor suppressor in clear-cell renal cell carcinoma. Cancer Lett 2016; 374: 107-16.
Hall JG. Importance of muscle movement for normal craniofacial development. J Craniofac Surg 2010; 21: 1336-8.
Hu X, Gao J, Liao Y, Tang S, Lu F. Retinoic acid alters the proliferation and survival of the epithelium and mesenchyme and suppresses Wnt/β-catenin signaling in developing cleft palate. Cell Death Dis 2013; 4: e898.
Morris-Wiman J, Brinkley L. An extracellular matrix infrastructure provides support for murine secondary palatal shelf remodeling. Anat Rec 1992; 234: 575-86.
D’Angelo M, Chen JM, Ugen K, Greene RM. TGF beta 1 regulation of collagen metabolism by embryonic palate mesenchymal cells. J Exp Zool 1994; 270: 189-201.
Vaziri Sani F, Kaartinen V, El Shahawy M, Linde A, Gritli-Linde A. Developmental changes in cellular and extracellular structural macromolecules in the secondary palate and in the nasal cavity of the mouse. Eur J Oral Sci 2010; 118: 221-36.
Gkantidis N, Blumer S, Katsaros C, Graf D, Chiquet M. Site-specific expression of gelatinolytic activity during morphogenesis of the secondary palate in the mouse embryo. PLoS One 2012; 7: e47762.
Jin JZ, Ding J. Analysis of cell migration, transdifferentiation and apoptosis during mouse secondary palate fusion. Development 2006; 133: 3341-7.
Yin K, Yin W, Wang Y, et al. MiR-206 suppresses epithelial mesenchymal transition by targeting TGF-β signaling in estrogen receptor positive breast cancer cells. Oncotarget 2016; 7: 24537-48.
Liu G, Luo G, Bo Z, Liang X, Huang J, Li D. Impaired osteogenic differentiation associated with connexin43/microRNA-206 in steroid-induced avascular necrosis of the femoral head. Exp Mol Pathol 2016; 101: 89-99.
Ni Z, Shang X, Tang G, Niu L. Expression of miR-206 in human knee articular chondrocytes and effects of miR-206 on proliferation and apoptosis of articular chondrocytes. Am J Med Sci 2018; 355: 240-6.
Murillo J, Maldonado E, Barrio MC, et al. Interactions between TGF-beta1 and TGF-beta3 and their role in medial edge epithelium cell death and palatal fusion in vitro. Differentiation 2009; 77: 209-20.
Zhu X, Ozturk F, Liu C, Oakley GG, Nawshad A. Transforming growth factor-β activates c-Myc to promote palatal growth. J Cell Biochem 2012; 113: 3069-85.
Jin JZ, Warner DR, Lu Q, Pisano MM, Greene RM, Ding J. Deciphering TGF-β function in medial edge epithelium specification and fusion during mouse secondary palate development. Dev Dyn 2014; 243: 1536-43.
Nie X, Luukko K, Kettunen P. BMP signaling in craniofacial development. Int J Dev Biol 2006; 50: 511-21.
Iwata J, Parada C, Chai Y. The mechanism of TGF-β signaling during palate development. Oral Dis 2011; 17: 733-44.
Parada C, Chai Y. Roles of BMP signaling pathway in lip and palate development. Front Oral Biol 2012; 16: 60-70.
Kaartinen V, Voncken JW, Shuler C, et al. Abnormal lung development and cleft palate in mice lacking TGF-beta 3 indicates defects of epithelial-mesenchymal interaction. Nat Genet 1995; 11: 415-21.
Loeys BL, Chen J, Neptune ER, et al. A syndrome of altered cardiovascular, craniofacial, neurocognitive and skeletal development caused by mutations in TGFBR1 or TGFBR2. Nat Genet 2005; 37: 275-81.
Dudas M, Sridurongrit S, Nagy A, Okazaki K, Kaartinen V. Craniofacial defects in mice lacking BMP type I receptor Alk2 in neural crest cells. Mech Dev 2004; 121: 173-82.
Liu W, Sun X, Braut A, et al. Distinct functions for Bmp signaling in lip and palate fusion in mice. Development 2005; 132: 1453-61.
Sahoo T, Theisen A, Sanchez-Lara PA, et al. Microdeletion 20p12.3 involving BMP2 contributes to syndromic forms of cleft palate. Am J Med Genet A 2011; 155A: 1646-53.
Chen Q, Wang H, Hetmanski JB, et al. BMP4 was associated with NSCL/P in an Asian population. PLoS One 2012; 7: e35347.
Wyatt AW, Osborne RJ, Stewart H, Ragge NK. Bone morphogenetic protein 7 (BMP7) mutations are associated with variable ocular, brain, ear, palate, and skeletal anomalies. Hum Mutat 2010; 31: 781-7.
Rot I, Kablar B. Role of skeletal muscle in palate development. Histol Histopathol 2013; 28: 1-13.
Kim KO, Sampson ER, Maynard RD, et al. Ski inhibits TGF-β/phospho-Smad3 signaling and accelerates hypertrophic differentiation in chondrocytes. J Cell Biochem 2012; 113: 2156-66.
Li J, Li P, Zhang Y, et al. c-Ski inhibits the proliferation of vascular smooth muscle cells via suppressing Smad3 signaling but stimulating p38 pathway. Cell Signal 2013; 25: 159-67.
Huang QK, Qiao HY, Fu MH, et al. MiR-206 attenuates denervation-induced skeletal muscle atrophy in rats through regulation of satellite cell differentiation via TGF-β1, Smad3, and HDAC4 signaling. Med Sci Monit 2016; 22: 1161-70.
Juriloff DM, Harris MJ, McMahon AP, Carroll TJ, Lidral AC. Wnt9b is the mutated gene involved in multifactorial nonsyndromic cleft lip with or without cleft palate in A/WySn mice, as confirmed by a genetic complementation test. Birth Defects Res A Clin Mol Teratol 2006; 76: 574-9.
Menezes R, Letra A, Kim AH, et al. Studies with Wnt genes and nonsyndromic cleft lip and palate. Birth Defects Res A Clin Mol Teratol 2010; 88: 995-1000.
Yu H, Ye X, Guo N, Nathans J. Frizzled 2 and frizzled 7 function redundantly in convergent extension and closure of the ventricular septum and palate: evidence for a network of interacting genes. Development 2012; 139: 4383-94.
Jin YR, Han XH, Taketo MM, Yoon JK. Wnt9b-dependent FGF signaling is crucial for outgrowth of the nasal and maxillary processes during upper jaw and lip development. Development 2012; 139: 1821-30.
Cvjetkovic N, Maili L, Weymouth KS, et al. Regulatory variant in FZD6 gene contributes to nonsyndromic cleft lip and palate in an African-American family. Mol Genet Genomic Med 2015; 3: 440-51.
Lu YP, Han WT, Liu Q, et al. Variations in WNT3 gene are associated with incidence of non-syndromic cleft lip with or without cleft palate in a northeast Chinese population. Genet Mol Res 2015; 14: 12646-53.
Warner DR, Greene RM, Pisano MM. Cross-talk between the TGFß and Wnt signaling pathways in murine embryonic maxillary mesenchymal cells. FEBS Lett 2005; 579: 3539-46.
Warner DR, Smith HS, Webb CL, Greene RM, Pisano MM. Expression of Wnts in the developing murine secondary palate. Int J Dev Biol 2009; 53: 1105-12.
He F, Chen Y. Wnt signaling in lip and palate development. Front Oral Biol 2012; 16: 81-90.
Liu Y, Wang M, Zhao W, et al. Gpr177-mediated Wnt signaling is required for secondary palate development. J Dent Res 2015; 94: 961-7.
Cobourne MT, Xavier GM, Depew M, et al. Sonic hedgehog signalling inhibits palatogenesis and arrests tooth development in a mouse model of the nevoid basal cell carcinoma syndrome. Dev Biol 2009; 331: 38-49.
Wannasilp N, Solomon BD, Warren-Mora N, et al. Holoprosencephaly in a family segregating novel variants in ZIC2 and GLI2. Am J Med Genet A 2011; 155A: 860-4.
Jin B, Tao Q, Peng J, et al. DNA methyltransferase 3B (DNMT3B) mutations in ICF syndrome lead to altered epigenetic modifications and aberrant expression of genes regulating development, neurogenesis and immune function. Hum Mol Genet 2008; 17: 690-709.
Demeer B, Andrieux J, Receveur A, et al. Duplication 16p13.3 and the CREBBP gene: confirmation of the phenotype. Eur J Med Genet 2013; 56: 26-31.
Kraft M, Cirstea IC, Voss AK, et al. Disruption of the histone acetyltransferase MYST4 leads to a Noonan syndrome-like phenotype and hyperactivated MAPK signaling in humans and mice. J Clin Invest 2011; 121: 3479-91.
DeLaurier A, Nakamura Y, Braasch I, et al. Histone deacetylase-4 is required during early cranial neural crest development for generation of the zebrafish palatal skeleton. BMC Dev Biol 2012; 12: 16.
Winbanks CE, Beyer C, Hagg A, Qian H, Sepulveda PV, Gregorevic P. miR-206 represses hypertrophy of myogenic cells but not muscle fibers via inhibition of HDAC4. PLoS One 2013; 8: e73589.

Rights & PermissionsPrintExport Cite as

Article Details

Year: 2019
Published on: 31 July, 2018
Page: [43 - 60]
Pages: 18
DOI: 10.2174/2211536607666180801094528
Price: $65

Article Metrics

PDF: 26