Proteomics Insights into the Gene Network of cis9, trans11-Conjugated Linoleic Acid Biosynthesis in Bovine Mammary Gland Epithelial Cells

Abstract

The objective of the study was to elucidate the stearoyl-coenzyme A desaturase (SCD1)-dependent gene network of c9, t11-CLA biosynthesis in MAC-T cells from an energy metabolism perspective. The cells were divided into the CAY group (firstly incubated with CAY10566, a chemical inhibitor of SCD1, then incubated with trans-11-octadecenoic acid, (TVA)), the TVA group (only TVA), and the control group (without CAY, TVA). The c9, t11-CLA, and TVA contents were determined by gas chromatography. The mRNA levels of SCD1 and candidate genes were analyzed via real-time PCR. Tandem mass tag (TMT)-based quantitative proteomics, bioinformatic analysis, parallel reaction monitoring (PRM), and small RNA interference were used to explore genes involved in the SCD1-dependent c9, t11-CLA biosynthesis. The results showed that the SCD1 deficiency led by CAY10566 blocked the biosynthesis of c9, t11-CLA. In total, 60 SCD1-related proteins mainly involved in energy metabolism pathways were primarily screened by TMT-based quantitative proteomics analysis. Moreover, 17 proteins were validated using PRM analysis. Then, 11 genes were verified to have negative relationships with SCD1 after the small RNA interference analysis. Based on the above results, we concluded that genes involved in energy metabolism pathways have an impact on the SCD1-dependent molecular mechanism of c9, t11-CLA biosynthesis.

Progress of Conjugated Linoleic Acid on Milk Fat Metabolism in Ruminants and Humans

“…Specifically, trans-10, cis-12 CLA inhibits the expression of SCAP and INSIG1, which are responsible for the abundance of nuclear SREBP1 (nSREBP1), thereby suppressing the activation of SREBP1. Subsequently, mRNA levels of SREBP1 downstream genes ACACA, FASN, and SCD1 are all downregulated [66,67]. In vitro studies have shown that trans-10, cis-12 CLA treatment does not alter the transcription and translation levels of nSREBP1, but directly suppresses and reduces its mRNA levels when 26S protease activity is inhibited.…”