Modulation of hormone-sensitive lipase and protein kinase A-mediated lipolysis by perilipin A in an adenoviral reconstituted system.
Perilipin (Peri) A is a phosphoprotein located at the surface of intracellular lipid droplets in adipocytes. Activation of cyclic AMP-dependent protein kinase (PKA) results in the phosphorylation of Peri A and hormone-sensitive lipase (HSL), the predominant lipase in adipocytes, with concurrent stimulation of adipocyte lipolysis. To investigate the relative contributions of Peri A and HSL in basal and PKA-mediated lipolysis, we utilized NIH 3T3 fibroblasts lacking Peri A and HSL but stably overexpressing acyl-CoA synthetase 1 (ACS1) and fatty acid transport protein 1 (FATP1). When incubated with exogenous fatty acids, ACS1/FATP1 cells accumulated 5 times more triacylglycerol (TG) as compared with NIH 3T3 fibroblasts. Adenoviral-mediated expression of Peri A in ACS1/FATP1 cells enhanced TG accumulation and inhibited lipolysis, whereas expression of HSL fused to green fluorescent protein (GFPHSL) reduced TG accumulation and enhanced lipolysis. Forskolin treatment induced Peri A hyperphosphorylation and abrogated the inhibitory effect of Peri A on lipolysis. Expression of a mutated Peri A Delta 3 (Ser to Ala substitutions at PKA consensus sites Ser-81, Ser-222, and Ser-276) reduced Peri A hyperphosphorylation and blocked constitutive and forskolin-stimulated lipolysis. Thus, perilipin expression and phosphorylation state are critical regulators of lipid storage and hydrolysis in ACS1/FATP1 cells.
Souza
Sandra C
SC
Jean Meyer United States Department of Agriculture Human Nutrition Research Center on Aging at Tufts University, Boston, Massachusetts 02111, USA.
Muliro
Kizito V
KV
Liscum
Laura
L
Lien
Ping
P
Yamamoto
Mia T
MT
Schaffer
Jean E
JE
Dallal
Gerard E
GE
Wang
Xinzhong
X
Kraemer
Fredric B
FB
Obin
Martin
M
Greenberg
Andrew S
AS
D016475:3T3 Cells; D000256:Adenoviridae / Q000235:genetics*; D000409:Alanine / Q000737:chemistry; D000818:Animals; D001425:Bacterial Outer Membrane Proteins / Q000378:metabolism; D015153:Blotting, Western; D002352:Carrier Proteins; D002460:Cell Line; D005576:Colforsin / Q000494:pharmacology; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism*; D006868:Hydrolysis; D050356:Lipid Metabolism; D051379:Mice; D008856:Microscopy, Fluorescence; D016297:Mutagenesis, Site-Directed; D000070778:Perilipin-1; D010750:Phosphoproteins / Q000378:metabolism*; D010766:Phosphorylation; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D011487:Protein Conformation; D012694:Serine / Q000737:chemistry; D002787:Sterol Esterase / Q000378:metabolism*; D014280:Triglycerides / Q000378:metabolism
C118372:ACS1 protein, Haemophilus influenzae; D001425:Bacterial Outer Membrane Proteins; D002352:Carrier Proteins; D000070778:Perilipin-1; D010750:Phosphoproteins; D014280:Triglycerides; D005576:Colforsin; D012694:Serine; D017868:Cyclic AMP-Dependent Protein Kinases; D002787:Sterol Esterase; D000409:Alanine