Limits...
Lpcat3-dependent production of arachidonoyl phospholipids is a key determinant of triglyceride secretion.

Rong X, Wang B, Dunham MM, Hedde PN, Wong JS, Gratton E, Young SG, Ford DA, Tontonoz P - Elife (2015)

Bottom Line: Mice lacking Lpcat3 in the intestine fail to thrive during weaning and exhibit enterocyte lipid accumulation and reduced plasma TGs.Mice lacking Lpcat3 in the liver show reduced plasma TGs, hepatosteatosis, and secrete lipid-poor very low-density lipoprotein (VLDL) lacking arachidonoyl PLs.Mechanistic studies indicate that Lpcat3 activity impacts membrane lipid mobility in living cells, suggesting a biophysical basis for the requirement of arachidonoyl PLs in lipidating lipoprotein particles.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology and Laboratory Medicine, Howard Hughes Medical Institute, University of California, Los Angeles, Los Angeles, United States.

ABSTRACT
The role of specific phospholipids (PLs) in lipid transport has been difficult to assess due to an inability to selectively manipulate membrane composition in vivo. Here we show that the phospholipid remodeling enzyme lysophosphatidylcholine acyltransferase 3 (Lpcat3) is a critical determinant of triglyceride (TG) secretion due to its unique ability to catalyze the incorporation of arachidonate into membranes. Mice lacking Lpcat3 in the intestine fail to thrive during weaning and exhibit enterocyte lipid accumulation and reduced plasma TGs. Mice lacking Lpcat3 in the liver show reduced plasma TGs, hepatosteatosis, and secrete lipid-poor very low-density lipoprotein (VLDL) lacking arachidonoyl PLs. Mechanistic studies indicate that Lpcat3 activity impacts membrane lipid mobility in living cells, suggesting a biophysical basis for the requirement of arachidonoyl PLs in lipidating lipoprotein particles. These data identify Lpcat3 as a key factor in lipoprotein production and illustrate how manipulation of membrane composition can be used as a regulatory mechanism to control metabolic pathways.

Show MeSH
Lpcat3 activity regulates membrane lipid mobility and apoB lipidation in hepatocytes.(A) The distribution of apoB-containing lipoproteins in the Golgi fractions isolated from livers of Lpcat3fl/fl (F/F) and Lpcat3fl/flAlbumin-Cre (LKO) mice. Golgi luminal contents were subject to a density gradient ultracentrifugation. 12 fractions from the gradient were harvested from top to bottom and analyzed by western blot. (B) Triglyceride contents of the Golgi fractions isolated from livers of Lpcat3fl/fl (F/F) and Lpcat3fl/flAlbumin-Cre (LKO) mice. Results from two representative experiments were shown. (C) Live primary hepatocytes from Lpcat3fl/fl (Flox/Flox) and Lpcat3fl/flAlbumin-Cre (L-Lpcat3 KO) mice were stained with laurdan. The laurdan emission spectrum was captured by a 2-photon laser-scan microscope. Generalized polarization (GP) was calculated from the emission intensities obtained from images. Higher GP value indicates that membranes are more ordered and less dynamic. The GP value of each pixel was used to generate a pseudocolor GP image. (D) The binary histograms of the GP distribution of the GP images (n = 4). The size of the GP binary is 0.05.DOI:http://dx.doi.org/10.7554/eLife.06557.021
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4400582&req=5

fig10: Lpcat3 activity regulates membrane lipid mobility and apoB lipidation in hepatocytes.(A) The distribution of apoB-containing lipoproteins in the Golgi fractions isolated from livers of Lpcat3fl/fl (F/F) and Lpcat3fl/flAlbumin-Cre (LKO) mice. Golgi luminal contents were subject to a density gradient ultracentrifugation. 12 fractions from the gradient were harvested from top to bottom and analyzed by western blot. (B) Triglyceride contents of the Golgi fractions isolated from livers of Lpcat3fl/fl (F/F) and Lpcat3fl/flAlbumin-Cre (LKO) mice. Results from two representative experiments were shown. (C) Live primary hepatocytes from Lpcat3fl/fl (Flox/Flox) and Lpcat3fl/flAlbumin-Cre (L-Lpcat3 KO) mice were stained with laurdan. The laurdan emission spectrum was captured by a 2-photon laser-scan microscope. Generalized polarization (GP) was calculated from the emission intensities obtained from images. Higher GP value indicates that membranes are more ordered and less dynamic. The GP value of each pixel was used to generate a pseudocolor GP image. (D) The binary histograms of the GP distribution of the GP images (n = 4). The size of the GP binary is 0.05.DOI:http://dx.doi.org/10.7554/eLife.06557.021

Mentions: To further investigate the hypothesis that defective apoB lipidation was responsible for the phenotype of L-Lpcat3 KO mice, we isolated the Golgi apparatus from livers, fractionated the luminal contents by density gradient centrifugation, and analyzed the distribution of ApoB in the different fractions. There was reduced apoB in the most buoyant lipoprotein fractions, consistent with reduced lipidation of apoB particles in the absence of Lpcat3 (Figure 10A). In line with this finding, we found reduced TG levels in Golgi membrane fractions isolated by density gradient centrifugation from L-Lpcat3 KO mice compared to controls in two independent purifications (Figure 10B).10.7554/eLife.06557.021Figure 10.Lpcat3 activity regulates membrane lipid mobility and apoB lipidation in hepatocytes.


Lpcat3-dependent production of arachidonoyl phospholipids is a key determinant of triglyceride secretion.

Rong X, Wang B, Dunham MM, Hedde PN, Wong JS, Gratton E, Young SG, Ford DA, Tontonoz P - Elife (2015)

Lpcat3 activity regulates membrane lipid mobility and apoB lipidation in hepatocytes.(A) The distribution of apoB-containing lipoproteins in the Golgi fractions isolated from livers of Lpcat3fl/fl (F/F) and Lpcat3fl/flAlbumin-Cre (LKO) mice. Golgi luminal contents were subject to a density gradient ultracentrifugation. 12 fractions from the gradient were harvested from top to bottom and analyzed by western blot. (B) Triglyceride contents of the Golgi fractions isolated from livers of Lpcat3fl/fl (F/F) and Lpcat3fl/flAlbumin-Cre (LKO) mice. Results from two representative experiments were shown. (C) Live primary hepatocytes from Lpcat3fl/fl (Flox/Flox) and Lpcat3fl/flAlbumin-Cre (L-Lpcat3 KO) mice were stained with laurdan. The laurdan emission spectrum was captured by a 2-photon laser-scan microscope. Generalized polarization (GP) was calculated from the emission intensities obtained from images. Higher GP value indicates that membranes are more ordered and less dynamic. The GP value of each pixel was used to generate a pseudocolor GP image. (D) The binary histograms of the GP distribution of the GP images (n = 4). The size of the GP binary is 0.05.DOI:http://dx.doi.org/10.7554/eLife.06557.021
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4400582&req=5

fig10: Lpcat3 activity regulates membrane lipid mobility and apoB lipidation in hepatocytes.(A) The distribution of apoB-containing lipoproteins in the Golgi fractions isolated from livers of Lpcat3fl/fl (F/F) and Lpcat3fl/flAlbumin-Cre (LKO) mice. Golgi luminal contents were subject to a density gradient ultracentrifugation. 12 fractions from the gradient were harvested from top to bottom and analyzed by western blot. (B) Triglyceride contents of the Golgi fractions isolated from livers of Lpcat3fl/fl (F/F) and Lpcat3fl/flAlbumin-Cre (LKO) mice. Results from two representative experiments were shown. (C) Live primary hepatocytes from Lpcat3fl/fl (Flox/Flox) and Lpcat3fl/flAlbumin-Cre (L-Lpcat3 KO) mice were stained with laurdan. The laurdan emission spectrum was captured by a 2-photon laser-scan microscope. Generalized polarization (GP) was calculated from the emission intensities obtained from images. Higher GP value indicates that membranes are more ordered and less dynamic. The GP value of each pixel was used to generate a pseudocolor GP image. (D) The binary histograms of the GP distribution of the GP images (n = 4). The size of the GP binary is 0.05.DOI:http://dx.doi.org/10.7554/eLife.06557.021
Mentions: To further investigate the hypothesis that defective apoB lipidation was responsible for the phenotype of L-Lpcat3 KO mice, we isolated the Golgi apparatus from livers, fractionated the luminal contents by density gradient centrifugation, and analyzed the distribution of ApoB in the different fractions. There was reduced apoB in the most buoyant lipoprotein fractions, consistent with reduced lipidation of apoB particles in the absence of Lpcat3 (Figure 10A). In line with this finding, we found reduced TG levels in Golgi membrane fractions isolated by density gradient centrifugation from L-Lpcat3 KO mice compared to controls in two independent purifications (Figure 10B).10.7554/eLife.06557.021Figure 10.Lpcat3 activity regulates membrane lipid mobility and apoB lipidation in hepatocytes.

Bottom Line: Mice lacking Lpcat3 in the intestine fail to thrive during weaning and exhibit enterocyte lipid accumulation and reduced plasma TGs.Mice lacking Lpcat3 in the liver show reduced plasma TGs, hepatosteatosis, and secrete lipid-poor very low-density lipoprotein (VLDL) lacking arachidonoyl PLs.Mechanistic studies indicate that Lpcat3 activity impacts membrane lipid mobility in living cells, suggesting a biophysical basis for the requirement of arachidonoyl PLs in lipidating lipoprotein particles.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology and Laboratory Medicine, Howard Hughes Medical Institute, University of California, Los Angeles, Los Angeles, United States.

ABSTRACT
The role of specific phospholipids (PLs) in lipid transport has been difficult to assess due to an inability to selectively manipulate membrane composition in vivo. Here we show that the phospholipid remodeling enzyme lysophosphatidylcholine acyltransferase 3 (Lpcat3) is a critical determinant of triglyceride (TG) secretion due to its unique ability to catalyze the incorporation of arachidonate into membranes. Mice lacking Lpcat3 in the intestine fail to thrive during weaning and exhibit enterocyte lipid accumulation and reduced plasma TGs. Mice lacking Lpcat3 in the liver show reduced plasma TGs, hepatosteatosis, and secrete lipid-poor very low-density lipoprotein (VLDL) lacking arachidonoyl PLs. Mechanistic studies indicate that Lpcat3 activity impacts membrane lipid mobility in living cells, suggesting a biophysical basis for the requirement of arachidonoyl PLs in lipidating lipoprotein particles. These data identify Lpcat3 as a key factor in lipoprotein production and illustrate how manipulation of membrane composition can be used as a regulatory mechanism to control metabolic pathways.

Show MeSH