Overexpression of a modified human malonyl-CoA decarboxylase blocks the glucose-induced increase in malonyl-CoA level but has no impact on insulin secretion in INS-1-derived (832/13) beta-cells.
The long-chain acyl-CoA (LC-CoA) model of glucose-stimulated insulin secretion (GSIS) holds that secretion is linked to a glucose-induced increase in malonyl-CoA level and accumulation of LC-CoA in the cytosol. We have previously tested the validity of this proposal by overexpressing goose malonyl-CoA decarboxylase (MCD) in INS-1 cells, but these studies have been criticized due to: 1) the small insulin secretion response (2-4-fold) of the INS-1 cells used; 2) unknown contribution of the ATP-sensitive K(+) (K(ATP)) channel-independent pathway of GSIS in INS-1 cells, which has been implicated as the site at which lipids regulate insulin granule exocytosis; and 3) deletion of the N-terminal mitochondrial targeting sequence, but not the C-terminal peroxisomal targeting sequence in the goose MCD construct, raising the possibility that a significant fraction of the overexpressed enzyme was localized to peroxisomes. To address these outstanding concerns, INS-1-derived 832/13 cells, which exhibit robust K(ATP) channel-dependent and -independent pathways of GSIS, were treated with a new adenovirus encoding human MCD lacking both its mitochondrial and peroxisomal targeting sequences (AdCMV-MCD Delta 5), resulting in large increases in cytosolic MCD activity. Treatment of 832/13 cells with AdCMV-MCD Delta 5 completely blocked the glucose-induced rise in malonyl-CoA and attenuated the inhibitory effect of glucose on fatty acid oxidation. However, MCD overexpression had no effect on K(ATP) channel-dependent or -independent GSIS in 832/13 cells. Furthermore, combined treatment of 832/13 cells with AdCMV-MCD Delta 5 and triacsin C, an inhibitor of long chain acyl-CoA synthetase that reduces LC-CoA levels, did not impair GSIS. These findings extend our previous observations and are not consistent with the LC-CoA hypothesis as originally set forth.
Mulder
H
H
Touchstone Center for Diabetes Research and Department of Biochemistry, University of Texas Southwestern Medical Center, Dallas, Texas 75390, USA.
Lu
D
D
Finley
J
J
An
J
J
Cohen
J
J
Antinozzi
P A
PA
McGarry
J D
JD
Newgard
C B
CB
D000256:Adenoviridae / Q000235:genetics; D002262:Carboxy-Lyases / Q000502:physiology*; D002460:Cell Line; D003981:Diazoxide / Q000494:pharmacology; D005227:Fatty Acids / Q000378:metabolism; D005947:Glucose / Q000494:pharmacology*; D006801:Humans; D007328:Insulin / Q000378:metabolism*; D000078790:Insulin Secretion; D007515:Islets of Langerhans / Q000502:physiology*; D008316:Malonyl Coenzyme A / Q000096:biosynthesis*; D010084:Oxidation-Reduction; D014226:Triazenes / Q000494:pharmacology
D005227:Fatty Acids; D007328:Insulin; D014226:Triazenes; D008316:Malonyl Coenzyme A; C034613:triacsin C; D002262:Carboxy-Lyases; C021532:malonyl-CoA decarboxylase; D005947:Glucose; D003981:Diazoxide