Hepatic patatin-like phospholipase domain-containing protein 3 sequence, single nucleotide polymorphism presence, protein confirmation, and responsiveness to energy balance in dairy cows

Abstract

Patatin-like phospholipase domain-containing protein 3 (PNPLA3), commonly known as adiponutrin, is part of a novel subfamily of triglyceride lipase enzymes with potential effects on triglyceride metabolism in adipose and hepatic tissues. The predicted bovine PNPLA3 sequence has been identified, but expression of the gene had not been examined. The objectives of this study were to confirm the predicted bovine PNPLA3 gene sequence, determine expression of the bovine PNPLA3 gene in response to whole-animal energy balance, identify single nucleotide polymorphisms present in dairy cows, and verify the presence of the protein in the liver. Using liver biopsy samples collected from cows at +28d relative to calving (DRTC), RNA was isolated and used to generate a cDNA template for amplification of the entire predicted coding sequence of PNPLA3 via PCR. To determine if energy balance alters the expression of PNPLA3, RNA was isolated and mRNA expression quantified in liver samples from mid-lactation cows after a 5-d ad libitum period (n=5) and after a subsequent 5-d 50% feed restriction period (n=5), and in samples collected from cows at -14, +1, +14, and +28 DRTC (n=16). The presence of PNPLA3 protein was detected by Western blot in liver protein samples collected at +28 DRTC. Expression of hepatic PNPLA3 was decreased after a period of feed restriction (8.14 vs. 1.08±2.17 arbitrary units, ad libitum vs. fasted). Expression of PNPLA3 mRNA was decreased at +1 and +14 DRTC compared with -14 DRTC (23.35, 7.28, 10.17, and 14.5±4.9 arbitrary units, -14, +1, +14, and +28 DRTC, respectively). The presence of PNPLA3 protein was detected as a 55-kDa band in hepatic protein isolations from liver tissue collected at +28 DRTC. These data confirm the presence and sequence of the bovine hepatic PNPLA3 gene and single nucleotide polymorphisms. Furthermore, these data indicate responsiveness of bovine hepatic PNPLA3 to energy balance.

Responsiveness of PNPLA3 and lipid-related transcription factors is dependent upon fatty acid profile in primary bovine hepatocytes

“…It may be possible that the differences seen in PNPLA3 expression and resulting protein both in vitro and in vivo are due to regulation at both the transcriptional and translational levels. Energy status is known to affect PNPLA3 protein in rodents and humans 20 , and a growing body of evidence is in agreement that bovine PNPLA3 mRNA and its resulting protein are also affected by energy status 12 , 21 . Although cell culture models cannot fully recapitulate physiological energy status, a recent in vivo study demonstrated the relationships between energy status, liver TG, and PNPLA3 protein abundance 12 .…”

ADSA Foundation Scholar Award: Influencing hepatic metabolism: Can nutrient partitioning be modulated to optimize metabolic health in the transition dairy cow?

“…Studies in human hepatocytes focused on understanding nonalcoholic liver disease have demonstrated an inactivated isoform of PNPLA3 present in the cells cause a significant increase in TG accumulation (Huang et al, 2010). In dairy cows, it has been shown that PNPLA3 mRNA is decreased during feed restriction and at calving (McCann et al, 2014). Protein abundance is inversely related to hepatic lipid White: ADSA FOUNDATION SCHOLAR AWARD accumulation across the transition to lactation period (Pralle et al, 2015;Pralle et al, 2019).…”