Ordovás Laura, Roy Rosa, Pampín Sandra, Zaragoza Pilar, Osta Rosario, Rodríguez-Rey Jose Carlos, Rodellar Clementina
Laboratorio de Genética Bioquímica (LAGENBIO), Facultad de Veterinaria, Universidad de Zaragoza, Zaragoza.
Physiol Genomics. 2008 Jul 15;34(2):144-8. doi: 10.1152/physiolgenomics.00043.2008. Epub 2008 May 13.
Fatty acid synthase (FASN) is an enzyme that catalyzes de novo synthesis of fatty acids in cells. The bovine FASN gene maps to BTA 19, where several quantitative trait loci for fat-related traits have been described. Our group recently reported the identification of a single nucleotide polymorphism (SNP), g.763G>C, in the bovine FASN 5' flanking region that was significantly associated with milk fat content in dairy cattle. The g.763G>C SNP was part of a GC-rich region that may constitute a cis element for members of the Sp transcription factor family. Thus the SNP could alter the transcription factor binding ability of the FASN promoter and consequently affect the promoter activity of the gene. However, the functional consequences of the SNP on FASN gene expression are unknown. The present study was therefore directed at elucidating the underlying molecular mechanism that could explain the association of the SNP with milk fat content. Three cellular lines (3T3L1, HepG2, and MCF-7) were used to test the promoter and the transcription factor binding activities by luciferase reporter assays and electrophoretic mobility shift assays, respectively. Band shift assays were also carried out with nuclear extracts from lactating mammary gland (LMG) to further investigate the role of the SNP in this tissue. Our results demonstrate that the SNP alters the bovine FASN promoter activity in vitro and the Sp1/Sp3 binding ability of the sequence. In bovine LMG, the specific binding of Sp3 may account for the association with milk fat content.
脂肪酸合酶（FASN）是一种催化细胞内脂肪酸从头合成的酶。牛FASN基因定位于BTA 19，在该位置已报道了几个与脂肪相关性状的数量性状位点。我们的研究小组最近报告，在牛FASN 5'侧翼区域鉴定出一个单核苷酸多态性（SNP），g.763G>C，它与奶牛的乳脂含量显著相关。g.763G>C SNP是富含GC区域的一部分，该区域可能构成Sp转录因子家族成员的顺式元件。因此，该SNP可能会改变FASN启动子的转录因子结合能力，进而影响该基因的启动子活性。然而，该SNP对FASN基因表达的功能影响尚不清楚。因此，本研究旨在阐明可能解释该SNP与乳脂含量关联的潜在分子机制。分别使用三种细胞系（3T3L1、HepG2和MCF-7）通过荧光素酶报告基因检测和电泳迁移率变动分析来检测启动子和转录因子结合活性。还使用泌乳乳腺（LMG）的核提取物进行条带迁移分析，以进一步研究该SNP在该组织中的作用。我们的结果表明，该SNP在体外改变了牛FASN启动子活性以及该序列的Sp1/Sp3结合能力。在牛LMG中，Sp3的特异性结合可能解释了其与乳脂含量的关联。
