Knockout of butyrophilin subfamily 1 member A1 (BTN1A1) alters lipid droplet formation and phospholipid composition in bovine mammary epithelial cells

Liqiang Han, Menglu Zhang, Zhiyang Xing, Danielle N. Coleman, Yusheng Liang, Juan J. Loor, Guoyu Yang

Research output: Contribution to journalArticlepeer-review

Abstract

Background: Milk lipids originate from cytoplasmic lipid droplets (LD) that are synthesized and secreted from mammary epithelial cells by a unique membrane-envelopment process. Butyrophilin 1A1 (BTN1A1) is one of the membrane proteins that surrounds LD, but its role in bovine mammary lipid droplet synthesis and secretion is not well known. Methods: The objective was to knockout BTN1A1 in bovine mammary epithelial cells (BMEC) via the CRISPR/Cas9 system and evaluate LD formation, abundance of lipogenic enzymes, and content of cell membrane phospholipid (PL) species. Average LD diameter was determined via Oil Red O staining, and profiling of cell membrane phospholipid species via liquid chromatography-tandem mass spectrometry (LC-MS/MS). Results: Lentivirus-mediated infection of the Cas9/sgRNA expression vector into BMEC resulted in production of a homozygous clone BTN1A1 (-/-) . The LD size and content decreased following BTN1A1 gene knockout. The mRNA abundance of fatty acid synthase (FASN) and peroxisome proliferator-activated receptor-gamma (PPARG) was downregulated in the BTN1A1 (-/-) clone. Subcellular analyses indicated that BTN1A1 and LD were co-localized in the cytoplasm. BTN1A1 gene knockout increased the percentage of phosphatidylethanolamine (PE) and decreased phosphatidylcholine (PC), which resulted in a lower PC/PE ratio. Conclusions: Results suggest that BTN1A1 plays an important role in regulating LD synthesis via a mechanism involving membrane phospholipid composition.

Original languageEnglish (US)
Article number72
JournalJournal of Animal Science and Biotechnology
Volume11
Issue number1
DOIs
StatePublished - Jul 3 2020

Keywords

  • Lipid droplet
  • Mammary epithelial cell
  • Milk fat globule
  • Phospholipid

ASJC Scopus subject areas

  • Biotechnology
  • Food Science
  • Biochemistry
  • Animal Science and Zoology

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