DIABETES, VOL. 49, APRIL 2000 539 Differential Activation of Peroxisome P roliferator-Activated Receptor- by Trog litazon e and Rosiglitazone Heidi S. Camp, Ou Li, Scott C. Wise, Yu H. Hong, Christy L. Frankowski, Xi-qiang Shen, Ruth Vanbogelen, and Todd Leff The antidiabetic thiazolidinediones, which include troglitazone and rosiglitazone, are ligands for the nuclear receptor peroxisome proliferator- act ivat e d receptor (PPA R)- and exert their antihyperglycemic effects by regulation of PPA R- –responsive genes. We report here that PPA R- activation by troglitazone depends on the experimental setting. Troglitazone acts as a partial agonist for PPA R- in transfected muscle ( C2C12) and kidney ( HEK 293T) cells, producing a sub- maximal transcriptional response (1.8- to 2.5-fold acti- vation) compared with rosiglitazone (7.4- to 13-fold activation). Additionally, troglitazone antagonizes rosiglitazone-stimulated PPAR- transcriptional activity. Limited protease digestion of PPA R- suggests confor- mational differences in the receptor bound to troglita- zone versus rosiglitazone. Consistent with this finding, an in vitro coactivator association assay demonstrated that troglitazone-bound PPA R- recruited the transcrip- tional coactivators p300 and steroid receptor coactiva- tor 1 less efficiently than rosiglitazone-bound receptor. In contrast to these observations, troglitazone behaves as a full agonist of PPA R- in 3T3L1 adipocytes. Two- dimensional protein gel electrophoresis demonstrated that troglitazone and rosiglitazone regulated distinct but overlapping sets of genes in several cell types. Thus, troglitazone may behave as a partial agonist under cer- tain physiological circumstances and as a full agonist in others. These differences could be caused by variations in the amount of specific cofactors, differences in PPA R response elements, or the presence of different iso- forms of PPA R- . Diabetes 4 9 :5 3 9–547, 2000 T ype 2 diabetes is characterized by decreased insulin sensitivity of peripheral tissues. Glucose homeostasis is maintained under these circum- stances by increased insulin secretion from pan- creatic -cells. In some cases, the -c ell is unable to maintain increased output. The antidiabetic thiazolidinediones (TZDs), suc h as troglitazone, improve peripheral insulin sensitivity, leading to reduced blood glucose and insulin levels and the preservation of pancreatic function (1–4). Improvement of insulin sensitivity by TZDs is mo st likely due to the activation of the peroxisome proliferator-activated receptor (PPAR)- (5). The TZDs are high-affinity ligands for PPAR- in vitro, and the rank order of receptor affinity co rrelates with their in vivo hypoglycemic activity (6), with one reported exception (7). Although many of the molecular details are not clearly under- stood, a model has emerged in which activated PPAR- mo d- ulates the transcriptional activity of a set of genes encoding proteins that are impo rtant in gluco se and lipid metabo lism. H o wever, the identity of these genes and the precise pathways leading to the normalization of insulin sensitivity remain largely unknown. Rec ent ly, the X-ray crystal structure of the ligand-binding domain of PPAR- has been elucidated (3,8), revealing that ligand binding causes a conformational change within P PA R- such that the receptor is converted to an “activated mode” that promotes the recruitment of coactivators such as steroid receptor coactivator 1 (SRC-1) and p300. It has been postulated that these coactivators act as bridges to transmit the nuclear receptor regulatory signals to the cellular transcriptional machinery. Studies on estrogen and proges- terone receptors have suggested that a range of distinct recep- tor conformations can be induced by the binding of different ligands (9–13). Although less is known about PPAR- in this regard, it is possible that different PPAR- ligands induce unique conformational changes in the receptor, eliciting dis- tinct downstream biological effects. We have compared trogli- tazone to other TZDs in terms of its ability to activate P PA R- 1 in several different experimental paradigms, includ- ing transcriptional activatio n, c oactivato r binding, and recep- tor conformation. We report here that in certain situations, troglitazone behaves as a partial agonist of the receptor. RESEARCH DESIGN AND METHODS Chemicals. Troglitazone and BRL49653 (rosiglitazone) were synthesized at Parke-Davis Pharmaceutical Research Divisio n o f Warner-Lambert (Ann Arbor, From the Department of Cell Biology (H.S.C., O.L., S.C.W., Y.H.H., C.L.F., X.S., R.V., T.L.), Parke-Davis Pharmac eutical Research Division, Warner-L am- bert Company; and the Department of Biological Chemistry (T.L.), Uni- versity of Michigan Medical School, Ann Arbor, Michigan. Address correspondence and reprint requests to Dr. Todd Leff, Depart- ment of Cell Biology, Parke-Davis Research, 2800 Plymouth Rd., Ann Arbor, MI 48105. E-mail: todd.leff@wl.com. Received for publication 20 September 1999 and accepted in revised form 27 December 1999. All of the authors are employees of the Parke-Davis Research Division of Warne r-Lambert Co. Warne r-Lambert markets Rezulin (troglitazone), one of the drugs described in this article. 2d, two -dimensional; CAP, c-Cbl–associated protein; CMV, cytomegalo- virus; DMEM, Dulbecco’s modified Eagle’s medium; DTT, dithiothreitol; EC 50 , concentration at half-maximal stimulation; FBS, fetal bovine serum; GST, glutathione S-transferase; IC 50 , co ncentration at half-maximal inhibi- tion; LBD, ligand binding domain; PPAR, peroxisome proliferator-ac tivate d receptor; PPRE, PPAR response element; SRC-1, steroid receptor coacti- vato r 1; TK, thymidine kinase; TZD, thiazolidinedione.