Compensatory responses to pyruvate carboxylase suppression in islet beta-cells. Preservation of glucose-stimulated insulin secretion.
We have previously reported that glucose-stimulated insulin secretion (GSIS) is tightly correlated with pyruvate carboxylase (PC)-catalyzed anaplerotic flux into the tricarboxylic acid cycle and stimulation of pyruvate cycling activity. To further evaluate the role of PC in beta-cell function, we constructed a recombinant adenovirus containing a small interfering RNA (siRNA) specific to PC (Ad-siPC). Ad-siPC reduced PC mRNA levels by 83 and 64% and PC protein by 56 and 35% in INS-1-derived 832/13 cells and primary rat islets, respectively. Surprisingly, this manipulation did not impair GSIS in rat islets. In Ad-siPC-treated 832/13 cells, GSIS was slightly increased, whereas glycolytic rate and glucose oxidation were unaffected. Flux through PC at high glucose was decreased by only 20%, suggesting an increase in PC-specific activity. Acetyl carnitine, a surrogate for acetyl-CoA, an allosteric activator of PC, was increased by 36% in Ad-siPC-treated cells, suggesting a mechanism by which PC enzymatic activity is maintained with suppressed PC protein levels. In addition, the NADPH:NADP ratio, a proposed coupling factor for GSIS, was unaffected in Ad-siPC-treated cells. We conclude that beta-cells activate compensatory mechanisms in response to suppression of PC expression that prevent impairment of anaplerosis, pyruvate cycling, NAPDH production, and GSIS.
Jensen
Mette V
MV
Sarah W. Stedman Nutrition and Metabolism Center and Department of Pharmacology and Cancer Biology, Duke University Medical Center, Durham, North Carolina 27704.
Joseph
Jamie W
JW
Sarah W. Stedman Nutrition and Metabolism Center and Department of Pharmacology and Cancer Biology, Duke University Medical Center, Durham, North Carolina 27704.
Ilkayeva
Olga
O
Sarah W. Stedman Nutrition and Metabolism Center and Department of Pharmacology and Cancer Biology, Duke University Medical Center, Durham, North Carolina 27704.
Burgess
Shawn
S
Advanced Imaging Research Center and the Department of Radiology, University of Texas Southwestern Medical Center at Dallas, Dallas, Texas 75235.
Lu
Danhong
D
Sarah W. Stedman Nutrition and Metabolism Center and Department of Pharmacology and Cancer Biology, Duke University Medical Center, Durham, North Carolina 27704.
Ronnebaum
Sarah M
SM
Sarah W. Stedman Nutrition and Metabolism Center and Department of Pharmacology and Cancer Biology, Duke University Medical Center, Durham, North Carolina 27704.
Odegaard
Matthew
M
Sarah W. Stedman Nutrition and Metabolism Center and Department of Pharmacology and Cancer Biology, Duke University Medical Center, Durham, North Carolina 27704.
Becker
Thomas C
TC
Sarah W. Stedman Nutrition and Metabolism Center and Department of Pharmacology and Cancer Biology, Duke University Medical Center, Durham, North Carolina 27704; Department of Medicine, Duke University Medical Center, Durham, North Carolina 27704.
Sherry
A Dean
AD
Advanced Imaging Research Center and the Department of Radiology, University of Texas Southwestern Medical Center at Dallas, Dallas, Texas 75235.
Newgard
Christopher B
CB
Sarah W. Stedman Nutrition and Metabolism Center and Department of Pharmacology and Cancer Biology, Duke University Medical Center, Durham, North Carolina 27704; Department of Medicine, Duke University Medical Center, Durham, North Carolina 27704. Electronic address: Newga002@mc.duke.edu.
D000108:Acetylcarnitine / Q000032:analysis; D000494:Allosteric Regulation; D000818:Animals; D002460:Cell Line; D005947:Glucose / Q000494:pharmacology*; D007328:Insulin / Q000378:metabolism*; D000078790:Insulin Secretion; D050417:Insulin-Secreting Cells / Q000378:metabolism*; D007515:Islets of Langerhans; D009249:NADP / Q000096:biosynthesis; D011766:Pyruvate Carboxylase / Q000037:antagonists & inhibitors / Q000502:physiology*; D034741:RNA, Small Interfering / Q000494:pharmacology; D051381:Rats
D007328:Insulin; D034741:RNA, Small Interfering; D009249:NADP; D000108:Acetylcarnitine; D011766:Pyruvate Carboxylase; D005947:Glucose