Expression and Regulation by Insulin of Glut 3 in UMR 106-01, a Clonal Rat Osteosarcoma Cell Line D. M. Thomas,* ,1 F. Maher,† S. D. Rogers,* and J. D. Best* *University of Melbourne Department of Medicine, St Vincents Hospital, 41 Victoria Parade, Fitzroy 3065, Victoria, Australia; and †Department of Pathology, University of Melbourne, Grattan Street, Parkville, Victoria, Australia Received December 20, 1995 Expression of the glucose transporter GLUT 3 is mainly restricted to neuronal tissues, with lower levels in testis and placenta. In addition, GLUT 3 has recently been reported in neonatal rat calvaria by in situ hybridi- sation. We report the co-expression of GLUT 1 and GLUT 3 mRNA and protein in UMR 106-01, a clonal osteosarcoma cell line. By semi-quantitative analysis we show that GLUT 3 protein is expressed at levels comparable to GLUT 1. Insulin stimulates glucose uptake in UMR 106-01 cells. GLUT 3 mRNA and protein are increased by chronic (16 h) treatment with insulin, and the increase in GLUT 3 mRNA is not inhibited by cycloheximide. Regulation of GLUT 3 mRNA by insulin has not been previously shown. UMR 106-01 repre- sents a useful model for investigating regulation of GLUT 3 expression. © 1996 Academic Press, Inc. Physiological expression of the glucose transporter GLUT 3 is restricted principally to neuronal tissues, although low levels have been detected in testis, spermatazoa and placenta (1). The role of GLUT 3 in these tissues is unclear. Both GLUT 1 and GLUT 3 have high affinities for glucose which might provide a growth advantage by ensuring a supply of glucose under limiting conditions. Indeed, hypoglycemia induced GLUT 3 mRNA expression in mouse embryo neuronal cultures (2), and expression of GLUT 3 mRNA correlated with cell growth in CaCo-2 cell lines (3). Further- more, human tumors have increased levels of GLUT 1 and GLUT 3 mRNAs (4). Bone is a highly metabolically active tissue, but little is known of in vivo glucose transporter expression in bone. Evidence that GLUT 3 may be expressed in bone in vivo comes from in situ studies by Bondy et al. (5) in which membranous calvarial bone in neonatal rat brain is found to express high levels of GLUT 3 mRNA. Very little is known regarding the regulation of GLUT 3 mRNA by insulin. Longo et al. (6) found that insulin had no effect on GLUT 1 or GLUT 3 mRNA expression in cultured human fibroblasts, although phorbol esters induced a protein-synthesis dependent increase in both mRNAs. It was noted that this result might be explained by the low level of insulin receptor expression in these cells (1000 receptors/cell). However, it is known that treatment with 10nM IGF-I for 8 h increases GLUT 3 protein in L6 myotubes (7), and that both insulin and IGF-I act via the IGF-I receptor in this cell line (8). IGF-I and insulin bear considerable structural and functional homology, and exert their effects by common pathways involving activation of tyrosine kinase second messenger systems (9). Insulin is an important anabolic hormone in bone, with insulin deficiency in type I diabetic patients resulting in osteoporosis (10). In order to study the expression and regulation by insulin of GLUT 3 in bone cells in vitro we used UMR 106-01, a rat osteogenic sarcoma cell line which possesses phenotypic markers of the mature osteoblast phenotype, and which expresses abundant insulin receptors (80,000 receptors/cell) (11). Glucose transport is stimulated by prolonged insulin treatment in UMR 106-01 cells and this effect is due in part in part to increased GLUT 1 mRNA expression, accompanied by increased expression of GLUT 1 protein (12). We also studied UMR 201 and UMR 201-10B, clonal rat calvaria-derived cell lines with a preosteoblastic pheno- 1 To whom correspondence should be addressed. BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS 218, 789–793 (1996) ARTICLE NO. 0140 789 0006-291X/96 $12.00 Copyright © 1996 by Academic Press, Inc. All rights of reproduction in any form reserved.