Expression of GLUT2 in insulin-secreting AtT20 pituitary cells

E L Davies, K I Shennan, K Docherty, C J Bailey

Research output: Contribution to journalArticle

3 Citations (Scopus)

Abstract

The importance of the glucose transporter isoform, GLUT2, in the construction of glucose-sensitive surrogate insulin-secreting cells was evaluated using murine pituitary AtT20 cells. The cells were double transfected with cDNAs for human preproinsulin (hppI-1) driven by the cytomegalovirus promoter, and human GLUT2 driven by the beta-actin promoter. The stably transfected clone, AtTinsGLUT2.36, which strongly expressed both the hppI-1 and GLUT2 genes, constitutively released 7.5 ng/10(6) cells/24 h of immunoreactive insulin-like material, 75% of which was fully processed mature human insulin. Increasing glucose concentrations in the subphysiological range up to 50 microM increased insulin release, but greater glucose concentrations did not further increase insulin release. Suppression of the low-K(m) glucose-phosphorylating enzyme, hexokinase, with 2-deoxy-D-glucose increased glucose-stimulated insulin release by two- to threefold in the presence of subphysiological and physiological glucose concentrations up to 10 mM. Physiological glucose concentrations increased the amount of GLUT2 mRNA, indicating that the beta-actin promoter responds in a glucose-dependent manner. Implantation of 2 x 10(7) AtTinsGLUT2.36 cells intraperitoneally into streptozotocin-diabetic nude mice slowed the progression of hyperglycaemia. The implanted cells formed vascularised tumour-like cell aggregates attached to the peritoneum. The results demonstrate that the beta-actin promoter is partially regulated by glucose. Expression of GLUT2 enables glucose to enter the cell at high K(m), but high-K(m) glucose phosphorylation is also required to signal glucose-stimulated genes affecting insulin release.
Original languageEnglish
Pages (from-to)75-82
Number of pages8
JournalJournal of Molecular Endocrinology
Volume20
Issue number1
Publication statusPublished - Feb 1998

Fingerprint

Insulin
Glucose
Actins
Hexokinase
Facilitative Glucose Transport Proteins
Peritoneum
Deoxyglucose
Insulin-Secreting Cells
Streptozocin
Cytomegalovirus
Nude Mice
Hyperglycemia
Genes
Protein Isoforms
Complementary DNA
Clone Cells
Phosphorylation
Messenger RNA
Enzymes

Keywords

  • Animals
  • Cell Line
  • Cell Transplantation
  • Clone Cells
  • Gene Expression Regulation
  • Glucose
  • Glucose Transporter Type 2
  • Humans
  • Insulin
  • Mice
  • Monosaccharide Transport Proteins
  • Pituitary Gland
  • Proinsulin
  • Protein Precursors
  • Transfection

Cite this

Expression of GLUT2 in insulin-secreting AtT20 pituitary cells. / Davies, E L; Shennan, K I; Docherty, K; Bailey, C J.

In: Journal of Molecular Endocrinology, Vol. 20, No. 1, 02.1998, p. 75-82.

Research output: Contribution to journalArticle

Davies, EL, Shennan, KI, Docherty, K & Bailey, CJ 1998, 'Expression of GLUT2 in insulin-secreting AtT20 pituitary cells' Journal of Molecular Endocrinology, vol. 20, no. 1, pp. 75-82.
Davies, E L ; Shennan, K I ; Docherty, K ; Bailey, C J. / Expression of GLUT2 in insulin-secreting AtT20 pituitary cells. In: Journal of Molecular Endocrinology. 1998 ; Vol. 20, No. 1. pp. 75-82.
@article{f6ed2886d6c74209832d94fd11a225c3,
title = "Expression of GLUT2 in insulin-secreting AtT20 pituitary cells",
abstract = "The importance of the glucose transporter isoform, GLUT2, in the construction of glucose-sensitive surrogate insulin-secreting cells was evaluated using murine pituitary AtT20 cells. The cells were double transfected with cDNAs for human preproinsulin (hppI-1) driven by the cytomegalovirus promoter, and human GLUT2 driven by the beta-actin promoter. The stably transfected clone, AtTinsGLUT2.36, which strongly expressed both the hppI-1 and GLUT2 genes, constitutively released 7.5 ng/10(6) cells/24 h of immunoreactive insulin-like material, 75{\%} of which was fully processed mature human insulin. Increasing glucose concentrations in the subphysiological range up to 50 microM increased insulin release, but greater glucose concentrations did not further increase insulin release. Suppression of the low-K(m) glucose-phosphorylating enzyme, hexokinase, with 2-deoxy-D-glucose increased glucose-stimulated insulin release by two- to threefold in the presence of subphysiological and physiological glucose concentrations up to 10 mM. Physiological glucose concentrations increased the amount of GLUT2 mRNA, indicating that the beta-actin promoter responds in a glucose-dependent manner. Implantation of 2 x 10(7) AtTinsGLUT2.36 cells intraperitoneally into streptozotocin-diabetic nude mice slowed the progression of hyperglycaemia. The implanted cells formed vascularised tumour-like cell aggregates attached to the peritoneum. The results demonstrate that the beta-actin promoter is partially regulated by glucose. Expression of GLUT2 enables glucose to enter the cell at high K(m), but high-K(m) glucose phosphorylation is also required to signal glucose-stimulated genes affecting insulin release.",
keywords = "Animals, Cell Line, Cell Transplantation, Clone Cells, Gene Expression Regulation, Glucose, Glucose Transporter Type 2, Humans, Insulin, Mice, Monosaccharide Transport Proteins, Pituitary Gland, Proinsulin, Protein Precursors, Transfection",
author = "Davies, {E L} and Shennan, {K I} and K Docherty and Bailey, {C J}",
year = "1998",
month = "2",
language = "English",
volume = "20",
pages = "75--82",
journal = "Journal of Molecular Endocrinology",
issn = "0952-5041",
publisher = "Society for Endocrinology",
number = "1",

}

TY - JOUR

T1 - Expression of GLUT2 in insulin-secreting AtT20 pituitary cells

AU - Davies, E L

AU - Shennan, K I

AU - Docherty, K

AU - Bailey, C J

PY - 1998/2

Y1 - 1998/2

N2 - The importance of the glucose transporter isoform, GLUT2, in the construction of glucose-sensitive surrogate insulin-secreting cells was evaluated using murine pituitary AtT20 cells. The cells were double transfected with cDNAs for human preproinsulin (hppI-1) driven by the cytomegalovirus promoter, and human GLUT2 driven by the beta-actin promoter. The stably transfected clone, AtTinsGLUT2.36, which strongly expressed both the hppI-1 and GLUT2 genes, constitutively released 7.5 ng/10(6) cells/24 h of immunoreactive insulin-like material, 75% of which was fully processed mature human insulin. Increasing glucose concentrations in the subphysiological range up to 50 microM increased insulin release, but greater glucose concentrations did not further increase insulin release. Suppression of the low-K(m) glucose-phosphorylating enzyme, hexokinase, with 2-deoxy-D-glucose increased glucose-stimulated insulin release by two- to threefold in the presence of subphysiological and physiological glucose concentrations up to 10 mM. Physiological glucose concentrations increased the amount of GLUT2 mRNA, indicating that the beta-actin promoter responds in a glucose-dependent manner. Implantation of 2 x 10(7) AtTinsGLUT2.36 cells intraperitoneally into streptozotocin-diabetic nude mice slowed the progression of hyperglycaemia. The implanted cells formed vascularised tumour-like cell aggregates attached to the peritoneum. The results demonstrate that the beta-actin promoter is partially regulated by glucose. Expression of GLUT2 enables glucose to enter the cell at high K(m), but high-K(m) glucose phosphorylation is also required to signal glucose-stimulated genes affecting insulin release.

AB - The importance of the glucose transporter isoform, GLUT2, in the construction of glucose-sensitive surrogate insulin-secreting cells was evaluated using murine pituitary AtT20 cells. The cells were double transfected with cDNAs for human preproinsulin (hppI-1) driven by the cytomegalovirus promoter, and human GLUT2 driven by the beta-actin promoter. The stably transfected clone, AtTinsGLUT2.36, which strongly expressed both the hppI-1 and GLUT2 genes, constitutively released 7.5 ng/10(6) cells/24 h of immunoreactive insulin-like material, 75% of which was fully processed mature human insulin. Increasing glucose concentrations in the subphysiological range up to 50 microM increased insulin release, but greater glucose concentrations did not further increase insulin release. Suppression of the low-K(m) glucose-phosphorylating enzyme, hexokinase, with 2-deoxy-D-glucose increased glucose-stimulated insulin release by two- to threefold in the presence of subphysiological and physiological glucose concentrations up to 10 mM. Physiological glucose concentrations increased the amount of GLUT2 mRNA, indicating that the beta-actin promoter responds in a glucose-dependent manner. Implantation of 2 x 10(7) AtTinsGLUT2.36 cells intraperitoneally into streptozotocin-diabetic nude mice slowed the progression of hyperglycaemia. The implanted cells formed vascularised tumour-like cell aggregates attached to the peritoneum. The results demonstrate that the beta-actin promoter is partially regulated by glucose. Expression of GLUT2 enables glucose to enter the cell at high K(m), but high-K(m) glucose phosphorylation is also required to signal glucose-stimulated genes affecting insulin release.

KW - Animals

KW - Cell Line

KW - Cell Transplantation

KW - Clone Cells

KW - Gene Expression Regulation

KW - Glucose

KW - Glucose Transporter Type 2

KW - Humans

KW - Insulin

KW - Mice

KW - Monosaccharide Transport Proteins

KW - Pituitary Gland

KW - Proinsulin

KW - Protein Precursors

KW - Transfection

M3 - Article

VL - 20

SP - 75

EP - 82

JO - Journal of Molecular Endocrinology

JF - Journal of Molecular Endocrinology

SN - 0952-5041

IS - 1

ER -