Adenovirus-mediated expression of the catalytic subunit of glucose-6- phosphatase in INS-1 cells. Effects on glucose cycling, glucose usage, and insulin secretion

Khiet Trinh, Carol Minassian, Alex J Lange, Robert M. O'Doherty, Christopher B. Newgard

Research output: Contribution to journalArticle

39 Citations (Scopus)

Abstract

Glucose-6-phosphatase (Glu-6-Pase) catalyzes the terminal step of gluconeogenesis, the conversion of glucose 6-phosphate (Glu-6-P) to free glucose. This enzyme activity is thought to be conferred by a complex of proteins residing in the endoplasmic reticulum (ER), including a Glu-6-P translocase that transports Glu-6-P into the lumen of the ER, a phosphohydrolase catalytic subunit residing in the lumen, and putative glucose and inorganic phosphate transporters that allow exit of the products of the reaction. In this study, we have investigated the effect of adenovirus-mediated overexpression of the Glu-6-Pase catalytic subunit on glucose metabolism and insulin secretion, using a well differentiated insulinoma cell line, INS-1. We found that the overexpressed Glu-6-Pase catalytic subunit was normally glycosylated, correctly sorted to the ER, and caused a 10-fold increase in Glu-6-Pase enzymatic activity in in vitro assays. Consistent with these findings, a 4.2-fold increase in 3H2O incorporation into glucose was observed in INS-1 cells treated with the recombinant adenovirus containing the Glu-6-Pase catalytic subunit cDNA (AdCMV-Glu-6-Pase). 3-[3H]Glucose usage was decreased by 32% in AdCMV-Glu- 6-Pase-treated cells relative to controls, resulting in a proportional 30% decrease in glucose-stimulated insulin secretion. Our findings indicate that overexpression of the Glu-6-Pase catalytic subunit significantly impacts glucose metabolism and insulin secretion in islet β-cells. However, INS-1 cells treated with AdCMV-Glu-6-Pase do not exhibit the severe alterations of β-cell function and metabolism associated with islets from rodent models of obesity and non-insulin-dependent diabetes mellitus, suggesting the involvement of genes in addition to the catalytic subunit of Glu-6-Pase in the etiology of such β-cell dysfunction.

Original languageEnglish (US)
Pages (from-to)24837-24842
Number of pages6
JournalJournal of Biological Chemistry
Volume272
Issue number40
DOIs
StatePublished - Oct 3 1997

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Glucose-6-Phosphatase
Adenoviridae
Catalytic Domain
Insulin
Glucose
Glucose-6-Phosphate
Metabolism
Endoplasmic Reticulum
Cells
Phosphate Transport Proteins
Insulinoma
Gluconeogenesis
Enzyme activity
Medical problems
Islets of Langerhans
Phosphoric Monoester Hydrolases
Type 2 Diabetes Mellitus
Rodentia
Assays
Complementary DNA

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Adenovirus-mediated expression of the catalytic subunit of glucose-6- phosphatase in INS-1 cells. Effects on glucose cycling, glucose usage, and insulin secretion. / Trinh, Khiet; Minassian, Carol; Lange, Alex J; O'Doherty, Robert M.; Newgard, Christopher B.

In: Journal of Biological Chemistry, Vol. 272, No. 40, 03.10.1997, p. 24837-24842.

Research output: Contribution to journalArticle

Trinh, Khiet ; Minassian, Carol ; Lange, Alex J ; O'Doherty, Robert M. ; Newgard, Christopher B. / Adenovirus-mediated expression of the catalytic subunit of glucose-6- phosphatase in INS-1 cells. Effects on glucose cycling, glucose usage, and insulin secretion. In: Journal of Biological Chemistry. 1997 ; Vol. 272, No. 40. pp. 24837-24842.
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abstract = "Glucose-6-phosphatase (Glu-6-Pase) catalyzes the terminal step of gluconeogenesis, the conversion of glucose 6-phosphate (Glu-6-P) to free glucose. This enzyme activity is thought to be conferred by a complex of proteins residing in the endoplasmic reticulum (ER), including a Glu-6-P translocase that transports Glu-6-P into the lumen of the ER, a phosphohydrolase catalytic subunit residing in the lumen, and putative glucose and inorganic phosphate transporters that allow exit of the products of the reaction. In this study, we have investigated the effect of adenovirus-mediated overexpression of the Glu-6-Pase catalytic subunit on glucose metabolism and insulin secretion, using a well differentiated insulinoma cell line, INS-1. We found that the overexpressed Glu-6-Pase catalytic subunit was normally glycosylated, correctly sorted to the ER, and caused a 10-fold increase in Glu-6-Pase enzymatic activity in in vitro assays. Consistent with these findings, a 4.2-fold increase in 3H2O incorporation into glucose was observed in INS-1 cells treated with the recombinant adenovirus containing the Glu-6-Pase catalytic subunit cDNA (AdCMV-Glu-6-Pase). 3-[3H]Glucose usage was decreased by 32{\%} in AdCMV-Glu- 6-Pase-treated cells relative to controls, resulting in a proportional 30{\%} decrease in glucose-stimulated insulin secretion. Our findings indicate that overexpression of the Glu-6-Pase catalytic subunit significantly impacts glucose metabolism and insulin secretion in islet β-cells. However, INS-1 cells treated with AdCMV-Glu-6-Pase do not exhibit the severe alterations of β-cell function and metabolism associated with islets from rodent models of obesity and non-insulin-dependent diabetes mellitus, suggesting the involvement of genes in addition to the catalytic subunit of Glu-6-Pase in the etiology of such β-cell dysfunction.",
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