Dexamethasone stimulates very low density lipoprotein (VLDL) receptor gene expression in differentiating 3T3-L1 cells.
To characterize endocrine mechanisms of very low density lipoprotein (VLDL) receptor regulation we studied mouse adipocytic 3T3-L1 cells. Lipid filled adipocyte-like cells are formed during a 5-7 day time course in the presence of insulin, dexamethasone and isobutylmethylxanthine (IBMX). The VLDL receptor protein, in the form of its approximately 120 and approximately 100 kDa type I and type II isoforms, as well as binding of (125)I-beta-VLDL, was induced several-fold during differentiation. Among the three different constituents added to the culture medium only dexamethasone (1 microM), but not insulin or IBMX, induced a time- and dose-dependent increase of VLDL receptor expression. Inclusion of RU-486 (10 microM) blocked the stimulatory effect of dexamethasone on VLDL receptor mRNA and protein levels. 3.6 kb of the 5'-untranslated region representing the VLDL receptor promoter were cloned and sequenced, and the transcriptional start site was determined by primer extension to be located 574 bases upstream from the initiating methionine. To investigate the functionality of the promoter, luciferase reporter gene constructs for the region -181 to -3726 bases were assembled and transfected into 3T3-L1 cells. An increased reporter gene activity was recorded when comparing preconfluent cells to fully differentiated cells. Between day 0 and day 2 (48 h after transfection) reporter gene activity was induced by dexamethasone, but not by insulin or IBMX. RU-486 inhibited this stimulatory effect for all constructs tested. No classical glucocorticoid receptor (GR) response element was found in the sequenced region of the VLDL receptor promoter. Thus, an indirect stimulatory effect mediated via GR on VLDL receptor gene transcription is the most likely mechanism of VLDL receptor gene activation in differentiating 3T3-L1 cells.
Ensler
Katharina
K
Department of Medicine, Center for Metabolism and Endocrinology, Molecular Nutrition Unit, Center for Nutrition and Toxicology, NOVUM, S-141 86 Huddinge, Sweden.
Mohammadieh
Majid
M
Bröijersén
Anders
A
Angelin
Bo
B
Gåfvels
Mats
M
D016475:3T3 Cells; D017667:Adipocytes / Q000378:metabolism*; D000818:Animals; D001483:Base Sequence; D002454:Cell Differentiation; D003907:Dexamethasone / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D017930:Genes, Reporter; D005938:Glucocorticoids / Q000494:pharmacology*; D051379:Mice; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000096:biosynthesis; D011973:Receptors, LDL / Q000096:biosynthesis / Q000235:genetics*; D020218:Response Elements; D024363:Transcription Initiation Site; D015533:Transcriptional Activation
D005938:Glucocorticoids; D012333:RNA, Messenger; D011973:Receptors, LDL; C036053:VLDL receptor; D003907:Dexamethasone