Overexpression of PER2 Promotes De Novo Fatty Acid Synthesis, Fatty Acid Desaturation, and Triglyceride Accumulation in Bovine Mammary Epithelial Cells

The core clock gene <i>Period2 (PER2)</i> is associated with mammary gland development and lipid synthesis in rodents and has recently been found to have a diurnal variation in the process of lactation, but has not yet been demonstrated in bovine mammary epithelial cells (BMECs). To explore the regulatory function of <i>PER2</i> on milk fat synthesis in bovine mammary epithelial cells, we initially assessed the expression of clock genes and milk fat metabolism genes for 24 h using real-time quantitative PCR and fitted the data to a cosine function curve. Subsequently, we overexpressed the <i>PER2</i> in BMECs using plasmid vector (pcDNA3.1-PER2), with empty vector pcDNA3.1-myc as the control. After transfecting BMECs for 48 h, we assessed the protein abundance related to milk fat synthesis by Western blot, the expression of genes coding for these proteins using real time-quantitative PCR, the production of triacylglycerol, and the fatty acid profile. The findings indicated that a total of nine clock genes (<i>PER1/2</i>, <i>CRY1/2</i>, <i>REV-ERBα</i>, <i>BMAL1</i>, <i>NCOR1</i>, <i>NR2F2</i>, <i>FBXW11</i>), seven fatty acid metabolism genes (<i>CD36</i>, <i>ACSS2</i>, <i>ACACA</i>, <i>SCD</i>, <i>FADS1</i>, <i>DGAT1</i>, <i>ADFP</i>), and six nuclear receptor-related genes (<i>INSIG1</i>, <i>SCAP</i>, <i>SREBF1</i>, <i>C/EBP</i>, <i>PPARG</i>, <i>LXR</i>) exhibited oscillation with a period close to 24 h in non-transfected BMECs (R² ≥ 0.7). Compared to the control group (transfected with empty pcDNA3.1-myc), the triglyceride content significantly increased in the <i>PER2</i> overexpression group (<i>p</i> < 0.05). The lipogenic genes for fatty acid transport and triglyceride synthesis (<i>ACACA</i>, <i>SCD</i>, <i>LPIN1</i>, <i>DGAT1</i>, and <i>SREBF1</i>) were upregulated after <i>PER2</i> overexpression, along with the upregulation of related protein abundance (<i>p</i> < 0.05). The contents and ratios of palmitic acid (C16:0), oleic acid (C18:1n9c), and trans-oleic acid (C18:1n9t) were significantly increased in the overexpression group (<i>p</i> < 0.05). Overall, the data supported that <i>PER2</i> participated in the process of milk fat metabolism and is potentially involved in the de novo synthesis and desaturation of fatty acid in bovine mammary epithelial cells.