Abstract

Intramuscular fat (IMF) content is a key determinant of beef quality, making it a key topic of research interest. ATP5B serves as the catalytic component of the mitochondrial ATP synthase complex and plays essential roles in controlling fat contents and oxidative metabolism in bovine skeletal muscle. In this study, we determined that bovine ATP5B was highly expressed in longissimus thoracis. To elucidate the molecular mechanisms involved in bovine ATP5B regulation, we cloned and characterized the promoter region of ATP5B. Applying 5'-rapid amplification of cDNA end analysis (RACE), we identified two transcriptional start sites (TSSs) in its promoter region. Using a series of 5'-deletion promoter plasmids in luciferase reporter assay, we found that the proximal minimal promoter of ATP5B was located within the region −539/220 relative to the TSS. Site-directed mutation in combination with chromatin immunoprecipitation (ChIP) assays demonstrated that MyoD and GATA1 binding to the promoter region drives bovine ATP5B transcription. Taken together, these results provide new insight into the regulatory mechanisms of ATP5B transcription in mediating the IMF content of beef.

摘要

肌内脂肪 (IMF) 含量是牛肉质量的关键决定因素，使其成为研究兴趣的关键课题。ATP5B 作为线粒体 ATP 合酶复合物的催化成分，在控制牛骨骼肌中的脂肪含量和氧化代谢中起重要作用。在这项研究中，我们确定牛 ATP5B 在胸最长肌中高度表达。为了阐明参与牛 ATP5B 调控的分子机制，我们克隆并表征了 ATP5B 的启动子区域。应用 cDNA 末端分析 (RACE) 的 5'-快速扩增，我们在其启动子区域确定了两个转录起始位点 (TSS)。在荧光素酶报告基因检测中使用一系列 5'-缺失启动子质粒，我们发现 ATP5B 的近端最小启动子相对于 TSS 位于 -539/220 区域内。定点突变与染色质免疫沉淀 (ChIP) 测定相结合，证明 MyoD 和 GATA1 与启动子区域的结合驱动牛 ATP5B 转录。总之，这些结果为 ATP5B 转录调节牛肉 IMF 含量的调节机制提供了新的见解。