Knockdown of ATP citrate lyase in pancreatic beta cells does not inhibit insulin secretion or glucose flux and implicates the acetoacetate pathway in insulin secretion.
OBJECTIVE
Glucose-stimulated insulin secretion in pancreatic beta cells requires metabolic signals including the generation of glucose-derived short chain acyl-CoAs in the cytosol from mitochondrially-derived metabolites. One concept of insulin secretion is that ATP citrate lyase generates short chain acyl-CoAs in the cytosol from mitochondrially-derived citrate. Of these, malonyl-CoA, is believed to be an important signal in insulin secretion. Malonyl-CoA is also a precursor for lipids. Our recent evidence suggested that, in the mitochondria of beta cells, glucose-derived pyruvate can be metabolized to acetoacetate that is exported to the cytosol and metabolized to the same short chain acyl-CoAs and fatty acids that can be derived from citrate. We tested for redundancy of the citrate pathway.


METHODS
We inhibited ATP citrate lyase activity using hydroxycitrate as well as studying a stable cell line generated with shRNA knockdown of ATP citrate lyase in the pancreatic beta cell line INS-1 832/13.


RESULTS
In both instances glucose-stimulated insulin release was not inhibited. Mass spectrometry analysis showed that the flux of carbon from [U-(13)C]glucose and/or [U-(13)C]α-ketoisocaproic acid (KIC) into short chain acyl-CoAs in cells with hydroxycitrate-inhibited ATP citrate lyase or in the cell line with stable severe (>90%) shRNA knockdown of ATP citrate lyase was similar to the controls. Both (13)C-glucose and (13)C-KIC introduced substantial (13)C labeling into acetyl-CoA, malonyl-CoA, and HMG-CoA under both conditions. Glucose flux into fatty acids was not affected by ATP citrate lyase knockdown.


CONCLUSIONS
The results establish the involvement of the acetoacetate pathway in insulin secretion in pancreatic beta cells.
Metabolism, Endocrinology and Diabetes, University of Michigan Medical School, Ann Arbor, MI 48109, United States.
Mahmoud
M
El Azzouny
Childrens Diabetes Center, University of Wisconsin School of Medicine and Public Health, Madison, WI 53706, United States.
Melissa J
MJ
Longacre
Childrens Diabetes Center, University of Wisconsin School of Medicine and Public Health, Madison, WI 53706, United States.
Israr-Ul H
IH
Ansari
Chemistry, University of Michigan Medical School, Ann Arbor, MI 48109, United States.
Robert T
RT
Kennedy
Metabolism, Endocrinology and Diabetes, University of Michigan Medical School, Ann Arbor, MI 48109, United States.
Charles F
CF
Burant
Childrens Diabetes Center, University of Wisconsin School of Medicine and Public Health, Madison, WI 53706, United States. Electronic address: mjmacdon@wisc.edu.
Michael J
MJ
MacDonald
ACC, acetyl-CoA carboxylase; ACLY, ATP citrate lyase; Acetoacetate pathway; Acetyl-CoA; Citrate; KIC, α-ketoisocaproic acid; Malonyl-CoA; Mass spectrometry; Mitochondrial biosynthesis; Palmitate; SCOT, succinyl-CoA:3-ketoacid-CoA transferase; ZMP, 5-aminoimidazole-4-carboxamide ribonucleotide
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