Association of Novel Polymorphisms in Lymphoid Enhancer Binding Factor 1 (LEF-1) Gene with Number of Teats in Different Breeds of Pig

  • Xu, Ru-Xiang (Laboratory of Animal Fat Deposition and Muscle Development, College of Animal Science and Technology, Northwest A&F University) ;
  • Wei, Ning (Laboratory of Animal Fat Deposition and Muscle Development, College of Animal Science and Technology, Northwest A&F University) ;
  • Wang, Yu (Laboratory of Animal Fat Deposition and Muscle Development, College of Animal Science and Technology, Northwest A&F University) ;
  • Wang, Guo-Qiang (Laboratory of Animal Fat Deposition and Muscle Development, College of Animal Science and Technology, Northwest A&F University) ;
  • Yang, Gong-She (Laboratory of Animal Fat Deposition and Muscle Development, College of Animal Science and Technology, Northwest A&F University) ;
  • Pang, Wei-Jun (Laboratory of Animal Fat Deposition and Muscle Development, College of Animal Science and Technology, Northwest A&F University)
  • Received : 2013.11.29
  • Accepted : 2014.04.14
  • Published : 2014.09.01


Lymphoid enhancer binding factor 1 (LEF-1) is a member of the T-cell specific factor (TCF) family, which plays a key role in the development of breast endothelial cells. Moreover, LEF-1 gene has been identified as a candidate gene for teat number trait. In the present study, we detected two novel mutations (NC_010450.3:g. 99514A>G, 119846C>T) by DNA sequencing and polymerase chain reaction-restriction fragment length polymorphism in exon 4 and intron 9 of LEF-1 in Guanzhong Black, Hanjiang Black, Bamei and Large White pigs. Furthermore, we analyzed the association between the genetic variations with teat number trait in these breeds. The 99514A>G mutation showed an extremely significant statistical relevance between different genotypes and teat number trait in Guanzhong (p<0.001) and Large White (p = 0.002), and significant relevance in Hanjiang (p = 0.017); the 119846C>T mutation suggested significant association in Guanzhong Black pigs (p = 0.042) and Large White pigs (p = 0.003). The individuals with "AG" or "GG" genotype displayed more teat numbers than those with "AA"; the individuals with "TC" or "CC" genotype showed more teat numbers than those with "TT". Our findings suggested that the 99514A>G and 119846C>T mutations of LEF-1 affected porcine teat number trait and could be used in breeding strategies to accelerate porcine teat number trait improvement of indigenous pigs breeds through molecular marker assisted selection.


Pig;LEF-1 Gene;Expression Profile;Polymerase Chain Reaction-Restriction Fragment Length Polymorphism (PCR-RLFP);Teat Number;Haplotype


  1. Balinsky, B. I. 1950. On the prenatal growth of the mammary rudiment in the mouse. J. Anat. 84:227-235.
  2. Davenport, T. G., L. A. Jerome-Majewska, and V. E. Papaioannou. 2003. Mammary gland, limb and yolk sac defects in mice lacking Tbx3, the gene mutated in human ulnar-mammary syndrome. Development 130:2263-2273.
  3. Eblaghie, M. C., S. J. Song, J. Y. Kim, K. Akita, C. Tickle, and H. S. Jung. 2004. Interactions between FGF and Wnt signals and Tbx3 gene expression in mammary gland initiation in mouse embryos. J. Anat. 205:1-13.
  4. Holnthoner, W., M. Pillinger, M. Groger, K. Wolff, A. W. Ashton, C. Albanese, P. Neumeister, R. G. Pestell, and P. Petzelbauer. 2002. Fibroblast growth factor-2 Induces Lef/Tcf-dependent transcription in human endothelial cells. J. Biol. Chem. 277:45847-45853.
  5. Gebeshuber, C. A., S. Sladecek, and S. Grunert. 2007. Beta-catenin/LEF-1 signaling in breast cancer-central players activated by a plethora of inputs. Cells Tissues Organs 185:51-60.
  6. Jerome-Majewska, L. A., G. P. Jenkins, E. Ernstoff, F. Zindy, C. J. Sherr, and V. E. Papaioannou. 2005. Tbx3, the ulnar-mammary syndrome gene, and Tbx2 interact in mammary gland development through a p19Arf / p53-independent pathway. Dev. Dyn. 234:922-933.
  7. Jonas, E., H. J. Schreinemachers, T. Kleinwachter, C. Un, I. Oltmanns, S. Tetzlaff, D. Jennen, D. Tesfaye, S. Ponsuksili, E. Murani, H. Juengst, E. Tholen, K. Schellander, and K. Wimmers. 2008. QTL for the heritable inverted teat defect in pigs. Mamm. Genome 19:127-138.
  8. Bamshad, M., R. C. Lin, D. J. Law, W. S. Watkins, P. A. Krakowiak, M. E. Moore, P. Franceschini, R. Lala, L. B. Holmes, T. C. Gebuhr, B. G. Bruneau, A. Schinzel, J. G. Seidman, C. E. Seidman, and L. B. Jorde. 1997. Mutations in human TBX3 alter limb, apocrine and genital development in ulnar-mammary syndrome. Nat. Genet. 16:311-315.
  9. Boras-Granic, K., H. Chang, R. Grosschedl, and P. A. Hamel. 2006. Lef1 is required for the transition of Wnt signaling from mesenchymal to epithelial cells in the mouse embryonic mammary gland. Dev. Biol. 295:219-231.
  10. Bucan, V., K. Mandel, C. Bertram, A. Lazaridis, K. Reimers, T. W. Park-Simon, P. M. Vogt, and R. Hass. 2012. LEF-1 regulates proliferation and MMP-7 transcription in breast cancer cells. Genes Cells 17:559-567.
  11. Cho, K. W., J. Y. Kim, S. J. Song, E. Farrell, M. C. Eblaghie, H. J. Kim, C. Tickle, and H. S. Jung. 2006. Molecular interactions between Tbx3 and Bmp4 and a model for dorsoventral positioning of mammary gland development. Proc. Natl. Acad. Sci. 103:16788-16793.
  12. Chu, E. Y., J. Hens, T. Andl, A. Kairo, T. P. Yamaguchi, C. Brisken, A. Glick, J. J. Wysolmerski, and S. E. Millar. 2004. Canonical WNT signaling promotes mammary placode development and is essential for initiation of mammary gland morphogenesis. Development 131:4819-4829.
  13. Arce, L., N. N. Yokoyama, and M. L.Waterman. 2006. Diversity of LEF/TCF action in development and diseases. Oncogene 25:7492-7504.
  14. Wiesner, E. and S. Willer. 1978. Problems of occurrence of inverted nipples in swine. Monatshefte. Fur. Veterinarmedi-zin. 33:189-190.
  15. Xie, X., J. Lu, E. J. Kulbokas, T. R. Golub, V. Mootha, K. Lindblad-Toh, E. S. Lander, and M. Kellis. 2005. Systematic discovery of regulatory motifs in human promoters and 3'UTRs by comparison of several mammals. Nature 434:338-345.
  16. Sambrook, J. and D. W. Russell. 2001. Molecular Cloning: A Laboratory Manual. 3rd edn. Cold Spring Harbor Laboratory Press, New York, USA.
  17. Sham, P., J. S. Bader, I. Craig, M. O'Donovan, and M. Owen. 2002. DNA Pooling: a tool for large-scale association studies. Nat. Rev. Genet. 3:862-871.
  18. Tetzlaff, S., S. Chomdej, E. Jonas, S. Ponsuksili, E. Murani, C. Phatsara, K. Schellander and K. Wimmers. 2009. Association of parathyroid hormone-like hormone (PTHLH) and its receptor (PTHR1) with the number of functional and inverted teats in pigs. J. Anim. Breed. Genet. 126:237-241.
  19. Tetzlaff, S., E. Jonas, C. Phatsara, E. Murani, S. Ponsuksili, K. Schellander, and K. Wimmers. 2009. Evidence for association of lymphoid enhancer-binding factor-1 (LEF1) with the number of functional and inverted teats in pigs. Cytogenet. Genome Res. 124:139-146.
  20. van Genderen, C., R. M. Okamura, I. Farinas, R. G. Quo, T. G. Parslow, L. Bruhn, and R. Grosschedl. 1994. Development of several organs that require inductive epithelial-mesenchymal interactions is impaired in LEF-1-deficient mice. Genes Dev. 8:2691-2703.
  21. Veltmaat, J. M., F. Relaix, L. T. Le, K. Kratochwil, F. G. Sala, W. van Veelen, R. Rice, B. Spencer-Dene, A. A. Mailleux, D. P. Rice, J. P. Thiery, and S. Bellusci. 2006. Gli3-mediated somitic Fgf10 expression gradients are required for the induction and patterning of mammary epithelium along the embryonic axes. Development 133:2325-2335.
  22. Veltmaat, J. M., W. Van Veelen, J. P. Thiery, and S. Bellusci. 2004. Identification of the mammary line in mouse by Wnt10b expression. Dev. Dyn. 229:349-356.
  23. Wang, F., S. Reierstad, and D. A. Fishman. 2006. Matrilysin over-expression in MCF-7 cells enhances cellular invasiveness and pro-gelatinase activation. Cancer Let. 236:292-301.
  24. Livak, K. J. and T. D. Schmittgen. 2001. Analysis of relative gene expression data using real-time quantitative PCR and the $2^{-\Delta \Delta{CT}}$ method. Methods 25:402-408.
  25. Mailleux, A. A., B. Spencer-Dene, C. Dillon, D. Ndiaye, C. Savona-Baron, N. Itoh, S. Kato, C. Dickson, J. P. Thiery, and S. Bellusci. 2002. Role of FGF10/FGFR2b signaling during mammary gland development in the mouse embryo. Development 129:53-60.
  26. Mateescu, R. G., Z. Zhang, K. Tsai, J. Phavaphutanon, N. I. Burton-Wurster, G. Lust, R. Quaas, K. Murphy, G. M. Acland, and R. J. Todhunter. 2005. Analysis of allele fidelity, polymorphic information content, and density of microsatellites in a genome-wide screening for hip dysplasia in a crossbreed pedigree. J. Hered. 96:847-853.
  27. Nei, M. and W. H. Li. 1979. Mathematical model for studying genetic variation in terms of restriction endonucleases. Proc. Natl. Acad. Sci. USA. 76:5269-5273.
  28. Ravindranath, A., A. O'Connell, P. G. Johnston, and M. K. El-Tanani. 2008. The role of LEF/TCF factors in neoplastic transformation. Curr. Mol. Med. 8:38-50.