IGF-1 controls GLUT3 expression in muscle via the transcriptional factor Sp1.
Glucose transporter 3 (GLUT3), while first found in human fetal muscle, is predominantly expressed in brain and neural tissue. By several independent techniques we have previously shown that GLUT3 is expressed in human skeletal muscle cells. The structure of the human GLUT3 gene has not been previously reported nor has there been any evaluation of the 5'-untranslated region (UTR). To this end, we have cloned and sequenced the human GLUT3 gene. Insulin-like growth factor-1 (IGF-1) increased endogenous Glut3 protein in cultured L6 myotubes, and similarly stimulated luciferase activity in a construct of the human GLUT3 5'-UTR linked to a luciferase reporter gene. Actinomycin D, an inhibitor of mRNA synthesis, prevented IGF-1 stimulation of Glut3 protein. Transfection of L6 cells with Sp1 increased Glut3 and augmented IGF-1 stimulation of Glut3 expression. Knockdown of Glut3 expression in cultured L6 muscle cells using small interference RNA (siRNA) specific for Glut3 significantly reduced myocyte glucose uptake. DNAse footprinting and gel shift assays showed Sp1 specifically bound to the human GLUT3 5'-UTR. Substitution mutants of the human GLUT3 5'-UTR luciferase construct indicated that only one of three Sp1 site clusters was involved in IGF-1 action. These data, using both a human GLUT3 5'-UTR construct and L6 cells' endogenous promoter, suggest that IGF-1 plays a role in maintaining muscle GLUT3 expression and basal glucose uptake via the transcriptional factor Sp1.
Copland
John A
JA
The Mayo Clinic Cancer Center, Jacksonville, Florida, USA.
Pardini
Aaron W
AW
Wood
Thomas G
TG
Yin
Deling
D
Green
Allan
A
Bodenburg
Yvonne H
YH
Urban
Randall J
RJ
Stuart
Charles A
CA
D020121:5' Untranslated Regions / Q000378:metabolism; D001483:Base Sequence; D002478:Cells, Cultured; D005786:Gene Expression Regulation; D005947:Glucose / Q000378:metabolism; D051274:Glucose Transporter Type 3 / Q000235:genetics*; D006801:Humans; D007334:Insulin-Like Growth Factor I / Q000502:physiology*; D008969:Molecular Sequence Data; D009132:Muscles / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000494:pharmacology; D016329:Sp1 Transcription Factor / Q000502:physiology*; D014158:Transcription, Genetic; D015854:Up-Regulation
D020121:5' Untranslated Regions; D051274:Glucose Transporter Type 3; D034741:RNA, Small Interfering; D016329:Sp1 Transcription Factor; D007334:Insulin-Like Growth Factor I; D005947:Glucose