Case–Control Possible Association between Congenital Supernumerary Extra Functional Teats and IGF1, Leptin, Calpastatin and FBP30 Candidate Genes in Fat-Tailed Ghezel Ewes
Subject Areas :A. Javanmard 1 , A.H. Rajoni 2
1 - Department of Animal Science, Faculty of Agriculture, University of Tabriz, Tabriz, Iran
2 -
Keywords: candidate gene, IGF1, mammary gland, PCR-RFLP, sheep, supernumerary teat,
Abstract :
Supernumerary teats (SNT) are a prevalent and intricate polygenic trait that is frequently observed at birth in sheep. These additional teats have been linked to a heritable trait associated with maternal capability, especially in ewes with high fertility. Investigating the genomic basis of these characteristics is an ongoing endeavor and poses an intriguing research inquiry with implications for both biological understanding and practical applications. In order to delve into this scientific enigma, a specific cohort of 20 local fat-tailed Ghezel ewes, each possessing four functional teats, was carefully chosen to form the case group. In parallel, a control group comprising 25 healthy ewes with the standard two normal teats was established for com-parative analysis. Blood samples were meticulously collected from both groups of Ghezel ewes demonstrat-ing distinct phenotypic expressions, specifically those with either two or four functional teats. Following the sample collection, the genomic DNA extraction process was carried out using a commercial DNA purifica-tion kit provided by Qiagen company, known for its high-quality DNA extraction products. We conducted an examination of four PCR-RFLP markers on four candidate genes (IGF1, Leptin, Calpastatin, and FBP30). Through our research, we unveiled the allele distribution and genotype frequency of these potential genes in both the case and control groups of sheep. Our findings demonstrated notable disparities in the genotype and gene frequencies between the case and control groups for each candidate gene. Furthermore, we calculated the odds ratio (OR) and 95 percent confidence intervals (CIs) under Hardy-Weinberg disequi-librium using various genetic models. The χ2 test was also utilized to compare the genotype and allele dis-tribution of these candidate gene polymorphisms between the case and control groups. The results of our study are particularly intriguing as they indicate that individuals with an AA genotype for the leptin gene are significantly more likely to have four additional functional teats (OR=7.7143, 95 percent CI=1.6025, P-value=0.0108). Additionally, the results in a dominant model (AA vs. GA+GG) revealed an OR of 0.1296 and 95 percent CI of 0.02669–0.6240, with a p-value of 0.0108, further highlighting the association be-tween leptin genotypes and supernumerary teats (SNT). Moreover, the risk of having more than four func-tional teats is heightened by the GG genotype in the FBP3 gene (P<0.13). However, our analysis did not uncover any additional significant correlation between the polymorphism of another investigated candidate gene and supernumerary teats (SNT). Overall, this topic presents an intriguing area that warrants further research to gain a comprehensive understanding of the biological mechanisms governing the development of four functional teats in ewes with large litter sizes.
Aali M., Moradi-Shahrbabak H., Moradi-Shahrbabak M., Sadeghi M. and Yousefi A.R. (2017). Association of the calpastatin genotypes, haplotypes, and SNPs with meat quality and fatty acid composition in two Iranian fat-and thin-tailed sheep breeds. Small Rumin. Res. 149, 40-51.
Amao O.A., Osinowo O.A., Lakpini C.A.M., Dipeolu M.A., Abiola S.S. and Onwuka C.I. (2003). Types and frequency of udder shapes and abnormalities in West African Dwarf and Red Sokoto goats. Nigerian J. Anim. Prod. 30, 253-261.
Arnandis T., Ferrer-Vicens I., Garcia-Trevijano E.R., Miralles V.J., Garcia C., Torres L., Viña J.R. and Zaragozá R. (2012). Calpains mediate epithelial-cell death during mammary gland involution: Mitochondria and lysosomal destabilization. Cell Death Differ. 19, 1536-1548.
Bakhtiar R., Abdolmohammadi A., Hajarian H., Nikousefat Z. and Kalantar-Neyestanaki D. (2017). Investigation of the 5′ flank-ing region and exon 3 polymorphisms of IGF-1 gene showed moderate association with semen quality in Sanjabi breed rams. Theriogenology. 104, 186-191.
Bemji M.N. and Popoola S.A. (2011). A note on the incidence of udder abnormalities in West African Dwarf goat in South Western Nigeria. J. Livest. Res. Rural Dev. 23, 3-11.
Boucher D., Palin M.F., Castonguay F., Gariépy C. and Pothier F. (2006). Detection of polymorphisms in the ovine leptin (LEP) gene: Association of a single nucleotide polymorphism with muscle growth and meat quality traits. Canadian J. Anim. Sci. 86, 31-36.
Butty A.M., Frischknecht M., Gredler B., Baes C., Neuenschwander S. and Moll J. (2016). Genome-wide asso-ciation study for supernumerary teats in Swiss Brown Swiss Cattle reveals LGR5 as a major gene on chromosome 5. J. Anim. Sci. 94, 157-167.
Calvo J.H., Marcos S., Jurado J.J. and Serrano M. (2004). Asso-ciation of the heart fatty acid-binding protein (FABP3) gene with milk traits in Manchega breed sheep. Anim. Genet. 35, 347-356.
Chu E.Y., Hens J., Andl T., Kairo A., Yamaguchi T.P., Brisken C., Glick A., Wysolmerski J.J. and Millar S.E. (2004). Ca-nonical WNT signaling promotes mammary placode develop-ment and is essential for initiation of mammary gland morphogenesis. Development. 131(19), 4819-4829.
Darwish H.R., El-Shorbagy H.M., Abou-Eisha A.M., El-Din A.E. and Farag I.M. (2017). New polymorphism in the 5′ flanking region of IGF-1 gene and its association with wool traits in Egyptian Barki sheep. J. Genet. Eng. Biotechnol. 15(2), 437-441.
Davenport T.G., Jerome-Majewska L.A. and Papaioannou V.E. (2003). Mammary gland, limb and yolk sac defects in mice lacking Tbx3, the gene mutated in human ulnar mammary syndrome. Dev. Dis. 130, 2263-2273.
Dimitrova I., Bozhilova-Sakova M., Stancheva N. and Tzonev T. (2016). Molecular analysis of ovine myostatin gene (MSTN) in northeast Bulgarian Merino sheep breed using PCR-RFLP. Bulgarian J. Agric. Sci. 22, 10-18.
Famakinde S.A., Okwelum N. and Leigh O.O. (2019). Supernu-merary teats in Kalahari Red Goats in the humid tropics. Anim. Rev. 6, 17-23.
Foley J., Dann P., Hong J., Cosgrove J., Dreyer B., Rimm D., Dunbar M.E., Philbrick W.M. and Wysolmerski J. (2001). Parathyroid hormone-related protein maintains mammary epithelial fate and triggers nipple skin differentiation during embryonic breast development. Development. 128, 513-525.
Gobikrushanth M., Purfield D.C., Colazo M.G., Wang Z., Butler S.T. and Ambrose D.J. (2018). The relationship between se-rum insulin-like growth factor-1 (IGF-1) concentration and re-productive performance, and genome-wide associations for se-rum IGF-1 in Holstein cows. J. Dairy Sci. 101, 9154-9167.
Grochowska E., Borys B., Grześkowiak E. and Mroczkowski S. (2017). Effect of the calpain small subunit 1 gene (CAPNS1) polymorphism on meat quality traits in sheep. Small Rumin. Res. 150, 15-21.
Hardwick L.J.A., Phythian C.J., Fowden A.L. and Hughes K. (2020). Size of supernumerary teats in sheep correlates with complexity of the anatomy and microenvironment. J. Anat. 236, 954-962.
He J.N., Zhang B.Y., Chu M.X., Wang P.Q., Feng T., Cao G.L., Di R., Fang L., Huang D.W., Tang Q.Q. and Li N. (2012). Polymorphism of insulin-like growth factor 1 gene and its as-sociation with litter size in Small Tail Han sheep. Mol. Biol. Rep. 39, 9801-9807.
Huang D., Chen R., Yang Z., Mao Y., LI Y., Tian D., Liang C. and Yong Z.X. (2008). Analysis on associations of SNPs of leptin gene with growth traits in four sheep breeds. Chinese J. Anim. Vet. Sci. 12, 21-27.
Javanmard A., Mohammadabadi M.R., Zarrigabayi G.E., Ghara-hedaghi A.A., Nassiry M.R., Javadmansh A. and Asadzadeh N. (2008). Polymorphism within the intron region of the bo-vine leptin gene in Iranian Sarabi cattle (Iranian Bos taurus). Russian J. Genet. 44, 495-502.
Kenny S. (2014). Investigation of prevalence of supernumerary teat in livestock. PhD Thesis. Szent István Univ., Hungary. Lakhssassi K., Serrano M., Lahoz B., Sarto M.P., Iguácel L.P., Folch J., Alabart J.L. and Calvo J.H. (2020). The LEPR gene is associated with reproductive seasonality traits in Rasa Ara-gonesa sheep. Animals. 10, 2448-2456.
Liang M., Hou X., Qu B., Zhang N., Li N., Cui Y., Li Q.Z. and Gao X.J. (2014). Functional analysis of FABP3 in the milk fat synthesis signaling pathway of dairy cow mammary epithelial cells. In vitro Cell. Dev. Biol. Anim. 50, 865-873.
Liu W., Fang G., Fang Y., Tian K., Huang X., Yao X., Wang M., Yu H., Huang Y., Xin J. and Xin Y. (2010). The polymor-phism of a mutation of IGF-1 gene on two goat breeds in China. J. Anim. Vet. Adv. 9, 790-794.
Lundeheim N., Chalkias H. and Rydhmer L. (2013). Genetic analysis of teat number and litter traits in pigs. Acta Agric. Scandinavica A. Anim. Sci. 63, 121-126.
Martin P., Palhière I., Tosser-Klopp G. and Rupp R. (2016). Heritability and genome-wide association mapping for super-numerary teats in French Alpine and Saanen dairy goats. J. Dairy Sci. 99, 8891-900.
Meira A.N., Montenegro H., Coutinho L.L., Mourão G.B., Azevedo H.C., Muniz E.N., Machado A.L., Sousa Jr L.P., Pedrosa V.B. and Pinto L.F.B. (2019). Single nucleotide polymorphisms in the growth hormone and IGF type-1 (IGF1) genes associated with carcass traits in Santa Ines sheep. Ani-mal. 13, 460-468.
Ozoje M.O. (2002). Incidence and relative effects of qualitative traits in West African Dwarf goat. Small Rumin. Res. 43, 97-100.
Peng W.F., Xu S.S., Ren X., Lv F.H., Xie X.L., Zhao Y.X., Zhang M., Shen Z.Q., Ren Y.L., Gao L. and Shen M. (2017). A ge-nome‐wide association study reveals candidate genes for the supernumerary nipple phenotype in sheep (Ovis aries). Anim. Genet. 48, 570-579.
Pophiwa P., Webb E.C. and Frylinck L. (2020). A review of fac-tors affecting goat meat quality and mitigating strategies. Small Rumin. Res. 183, 106035-106041.
Raheem K.A. and Leigh O.O. (2014). Supernumenary teat in West Africa dwarf goat in Ibadan, Southwest Nigeria. IOSR J. Ag-ric. Vet. Sci. 7, 60-63.
Scatà M.C., Catillo G., Annicchiarico G., De Matteis G., Napoli-tano F. and Signorelli F. and Moioli B. (2010). Investigation on lactation persistency and IGF-I gene polymorphisms in dairy sheep. Small Rumin. Res. 89, 7-11.
Sharma A., Dutt G., Sivalingam J., Singh M.K., Pathodiya O.P., Khadda B.S. and Dixit S.P. (2013). Novel SNPs in IGF1, GHR and IGFBP-3 genes reveal significant association with growth traits in Indian goat breeds. Small Rumin. Res. 115, 7-14.
Utsunomiya Y.T., Pérez O’Brien A.M., Sonstegard T.S., Van Tassell C.P., do Carmo A.S., Meszaros G., Sölkner J. and Garcia J.F. (2013). Detecting loci under recent positive selec-tion in dairy and beef cattle by combining different genome-wide scan methods. PloS One. 8, e64280.
Van Genderen C., Okamura R.M., Farinas I., Quo R.G., Parslow T.G., Bruhn L. and Grosschedl R. (1994). Development of several organs that require inductive epithelial-mesenchymal interactions is impaired in LEF-1-deficient mice. Genes Dev. 8, 2691-2703.
Veltmaat J.M., Van Veelen W., Thiery J.P. and Bellusci S. (2004). Identification of the mammary line in mouse by Wnt10b ex-pression. Dev. Dyn. 229, 349-356.
Wang P., Ying Tan., Zhang B., Chu M., Deng L., Qi F.A.N. and Liu, C.X. (2011). DNA polymorphisms of 5′-flanking region of insulin-like growth factor 1 gene and their association with reproduction traits in goats. Agric. Sci. China. 10, 1609-1617.
Xu R.X., Wei N., Wang Y., Wang G.Q., Yang G.S. and Pang W.J. (2014). Association of novel polymorphisms in lymphoid en-hancer binding factor 1 (LEF-1) gene with number of teats in different breeds of pig. Asian-Australasian J. Anim. Sci. 27(9), 1254-1262.
Zhang L., Peng F., Yu F., Wan L. and Zhou Z. (2019). Expres-sion of ESR1, PRLR, GHR, and IGF1R in mammary glands of Hu sheep with four teats. Czech J. Anim. Sci. 64, 49-58.
Zhou H., Hickford J.G. and Gong H. (2009). Identification of allelic polymorphism in the ovine leptin gene. Mol. Biotech-nol. 41, 22-27.
