Nonsyndromic cleft lip with or without cleft palate (NSCL±P) is a common craniofacial anomaly of complex etiology in people. WNT pathway genes have important roles during craniofacial development, and an association of WNT genes with NSCL±P has been demonstrated in different populations. The aim of this study was to evaluate the association between polymorphisms in WNT3 and WNT9B genes and CL/P in Brazilian families.
Patients
Seventy nuclear families composed of an affected child and the child's unaffected parents were examined clinically. Saliva samples were collected for molecular analyses.
Design
Three single nucleotide polymorphisms (SNPs) in the WNT3 gene and two in WNT9B were investigated in real-time polymerase chain reaction using TaqMan chemistry. The Family-Based Association Test and the transmission disequilibrium test were used to verify the association between each marker allele and NSCL±P. The level of significance was established at P ≤ .01 after Bonferroni correction.
Results
A positive association was detected between NSCL±P and SNP rs1530364 in the WNT9B gene. Haplotype analysis showed an association of WNT3 and WNT9B haplotypes. No association was detected between NSCL±P and individual SNPs in WNT3.
Conclusion
Our study further supports the involvement of WNT9B as a cleft susceptibility gene in Brazilian families experiencing NSCL±P. Although additional studies are still necessary to unveil the exact mechanism by which WNT genes would contribute to NSCL±P, allelic polymorphisms in these genes and their interactions may partly explain the variance of individual susceptibility to NSCL±P.
BeatyT.H., TaubM.A., ScottA.F., MurrayJ.C., MarazitaM.L., SchwenderH., ParkerM.M., HetmanskiJ.B., BalakrishnanP., MansillaM.A.. Confirming genes influencing risk to cleft lip with/without cleft palate in a case-parent trio study. Hum Genet.2013; 132: 771–781.
2.
BirnbaumS., LudwigK.U., ReutterH., HermsS., SteffensM., RubiniM., BaluardoC., FerrianM., Almeida de AssisN., AlblasM.A.. Key susceptibility locus for nonsyndromic cleft lip with or without cleft palate on chromosome 8q24. Nat Genet.2009; 41: 473–477.
3.
BrugmannS.A., GoodnoughL.H., GregorieffA., LeuchtP., Ten BergeD., FuererC., CleversH., NusseR., HelmsJ.A.Wnt signaling mediates regional specification in the vertebrate face. Development.2007; 134: 3283–3295.
4.
CarinciF., ScapoliL., PalmieriA., ZollinoI., PezzettiF.Human genetic factors in nonsyndromic cleft lip and palate: an update. Int J Pediatr Otorhinolaryngol.2007; 71: 1509–1519.
5.
CarrollT.J., ParkJ.S., HayashiS., MajumdarA., McMahonA.P.Wnt9b plays a central role in the regulation of mesenchymal to epithelial transitions underlying organogenesis of the mammalian urogenital system. Dev Cell.2005; 9: 283–292.
6.
ChiquetB.T., BlantonS.H., BurtA., MaD., StalS., MullikenJ.B., HechtJ.T.Variation in WNT genes is associated with non-syndromic cleft lip with or without cleft palate. Hum Mol Genet.2008; 17(14): 2212–2218.
7.
FossatN., JonesV., KhooP.L., BoganiD., HardyA., SteinerK., MukhopadhyayM., WestphalH., NolanP.M., ArkellR.. Stringent requirement of a proper level of canonical WNT signaling activity for head formation in mouse embryo. Development.2011; 138: 667–676.
8.
GorlinR.J., CohenM.M., HennekamR.C.M.Syndromes of the Head and Neck.New York: Oxford University Press;2001.
9.
GrantS.F., WangK., ZhangH., GlabersonW., AnnaiahK., KimC.E., BradfieldJ.P., GlessnerJ.T., ThomasK.A., GarrisM.. A genome-wide association study identifies a locus for nonsyndromic cleft lip with or without cleft palate on 8q24. J Pediatr.2009; 155: 909–913.
10.
HeF., XiongW., YuX., Espinoza-LewisR., LiuC., GuS., NishitaM., SuzukiK., YamadaG., MinamiY., ChenY.Wnt5a regulates directional cell migration and cell proliferation via Ror2-mediated noncanonical pathway in mammalian palate development. Development.2008; 135: 3871–3879.
11.
HorvathS., XuX., LairdN.M.The family based association test method: strategies for studying general genotype-phenotype associations. Eur J Hum Genet.2001; 9: 301–306.
12.
JuriloffD.M., HarrisM.J., DewellS.L.A digenic cause of cleft lip in Astrain mice and definition of candidate genes for the two loci. Birth Defects Res A Clin Mol Teratol.2004; 70: 509–518.
13.
JuriloffD.M., HarrisM.J., DewellS.L., BrownC.J., MagerD.L., GagnierL., MahD.G.Investigations of the genomic region that contains the clf1 mutation, a causal gene in multifactorial cleft lip and palate in mice. Birth Defects Res A Clin Mol Teratol.2005; 73: 103–113.
14.
JuriloffD.M., HarrisM.J., McMahonA.P., CarrollT.J., LidralA.C.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: 574–579.
15.
LairdN.M., HorvathS., XuX.Implementing a unified approach to family-based tests of association. Genet Epidemiol.2000; 19(suppl 1): S36–S42.
16.
LanY., RyanR.C., ZhangZ., BullardS.A., BushJ.O., MaltbyK.M., LidralA.C., JiangR.Expression of Wnt9b and activation of canonical Wnt signaling during midfacial morphogenesis in mice. Dev Dyn.2006; 235: 1448–1454.
LetraA., SilvaR.A., MenezesR., GranjeiroJ.M.MMP gene polymorphisms as contributors for cleft lip/palate: association with MMP3 but not MMP1. Arch Oral Biol.2007; 86: 986–991.
19.
MarazitaM.L.Subclinical features in non-syndromic cleft lip with or without cleft palate (CL/P): review of the evidence that subepithelial orbicularis oris muscle defects are part of an expanded phenotype for CL/P. Orthod Craniofac Res.2007; 10: 82–87.
20.
MenezesR., LetraA., KimA.H., KüchlerE.C., DayA., TannureP.N., Gomes da MottaL., PaivaK.B., GranjeiroJ.M., VieiraA.R.Studies with Wnt genes and nonsyndromic cleft lip and palate. Birth Defects Res A Clin Mol Teratol.2010; 88: 995–1000.
21.
MostowskaA., HozyaszK.K., BiedziakB., WojcickaP., LianeriM., JagodzinskiP.P.Genotype and haplotype analysis of WNT genes in non-syndromic cleft lip with or without cleft palate. Eur J Oral Sci.2012; 120: 1–8.
NeiswangerK., WalkerK., KlotzC.M., CooperM.E., BardiK.M., BrandonC.A., WeinbergS.M., VieiraA.R., MartinR.A., CzeizelA.E.. Whorl patterns on the lower lip are associated with non-syndromic cleft lip with or without cleft palate. Am J Med Genet A.2009; 149A: 2673–2679.
24.
NiemannS., ZhaoC., PascuF., StahlU., AuleppU., NiswanderL., WeberJ.L., MullerU.Homozygous WNT3 mutation causes tetraamelia in a large consanguineous family. Am J Hum Genet.2004; 74: 558–563.
25.
NikopensiusT., KempaI., AmbrozaitytèL., JagomägiT., SaagM., MatulevičienėA., UtkusA., KrjutškovK., TammekiviV., PiekuseL.. Variation in FGF1, FOXE1, and TIMP2 genes is associated with nonsyndromic cleft lip with or without cleft palate. Birth Defects Res A Clin Mol Teratol.2011; 91: 218–225.
26.
PurcellS., ChernyS.S., ShamP.C.Genetic power calculator: design of linkage and association genetic mapping of complex traits. Bioinformatics.2003; 19: 149–150.
27.
RabinowitzD., LairdN.A unified approach to adjusting association tests for population admixture with arbitrary pedigree structure and arbitrary missing marker information. Hum Hered.2000; 50: 211–223.
28.
RanadeK., ChangM.S., TingC.T., PeiD., HsiaoC.F., OlivierM., OlivierM., PesichR., HebertJ., ChenY.D., DzauV.J.. High-throughput genotyping with single nucleotide polymorphisms. Genome Res.2001; 111262–111268.
29.
ReidB.S., YangH., MelvinV.S., TaketoM.M., WilliamsT.Ectodermal WNT/b-catenin signaling shapes the mouse face. Dev Biol.2011; 349: 261–269.
30.
SongL., LiY., WangK., WangY.Z., MolotkovA., GaoL., ZhaoT., YamagamiT., WangY., GanQ.. Lrp6- mediated canonical Wnt signaling is required for lip formation and fusion. Development.2009; 136: 3161–3171.
31.
WeinbergS.M., MaherB.S., MarazitaM.L. (2006). Parental craniofacial morphology in cleft lip with or without cleft palate as determined by cephalometry: a meta-analysis. Orthod Craniofacial Res9: 18–30.
32.
WodarzA., NusseR.Mechanisms of Wnt signaling in development. Annu Rev Cell Dev Biol.1998; 14: 59–88.
33.
YaoT., YangL., LiP.Q., WuH., XieH.B., ShenX., XieX.D.Association of Wnt3A gene variants with non-syndromic cleft lip with or without cleft palate in Chinese population. Arch Oral Biol.2011; 56: 73–78.
34.
YuanH.Y., ChiouJ.J., TsengW.H., LiuC.H., LiuC.K., LinY.J., WangH.H., YaoA., ChenY.T., HsuC.N.FASTSNP: an always up-to-date and extendable service for SNP function analysis and prioritization. Nucleic Acids Res.2006; 34(Web Server issue): W635–641.
35.
XuZ., TaylorJ.A.SNPinfo: integrating GWAS and candidate gene information into functional SNP selection for genetic association studies. Nucleic Acids Res2009; 37: W600–605. Available from: http://manticore.niehs.nih.gov/.
