Premium
Palmitate Increases Ubiquitination in H4IIE and Primary Rat Hepatocytes
Author(s) -
Stewart Claire M,
Estrada Andrea L,
Kim Paul Y,
Wang Dong,
Wei Yuren,
Pagliassotti Michael
Publication year - 2017
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.31.1_supplement.470.4
Subject(s) - mg132 , ubiquitin , proteasome , autophagy , endoplasmic reticulum , proteasome inhibitor , western blot , chemistry , biochemistry , biology , microbiology and biotechnology , apoptosis , gene
Saturated fatty acids induce endoplasmic reticulum (ER) stress in liver cells. The presence of ER stress implies that protein processing is impaired. We hypothesized that saturated fatty acid‐induced impairment in protein processing would result in increased ubiquitination and reduced autophagy and proteasome activity. Ubiquitination was assessed by western blot in H4IIE liver cells (n=3) or primary rat hepatocytes (n=3) in the presence of MG132 (5μM), a proteasome inhibitor, palmitate (250μM), a saturated fatty acid, or oleate (250μM), an unsaturated fatty acid, for 6 hours. Proteasome activity was measured in H4IIE liver cells (n=3) using the ZsProsensor‐1, a fluorescent reporter that contains a proteasome degradation domain, in the presence of MG132, palmitate, or oleate for 6 hours. Autophagy was assessed H4IIE liver cells (n=3) by western blot in the presence of palmitate or oleate for 6 hours. MG132 increased ubiquitination by 49% in H4IIE liver cells and primary rat hepatocytes. Palmitate increased ubiquitination by 26% in H4IIE hepatocytes and primary hepatocytes while oleate did not increase ubiquitination. MG132 reduced proteasome activity in H4IIE liver cells by 50% while palmitate or oleate had no significant effect on proteasome activity. Lastly, palmitate or oleate had no effect on autophagy. Overall, these data suggest that the palmitate‐mediated increase in ubiquitination occurred independently of changes in autophagy and the proteasome. Support or Funding Information NIH: DK072017