PPAR-gamma

Introduction

Peroxisome proliferator-activated receptor gamma (γ) is a protein in the nuclear receptors subfamily. It is one of three isotypes (-α, -β/ δ, and -γ) [1] of PPAR receptors and has two protein isoforms governed by splice variations, which result in differences in the length of the amino (N)-terminal region (PPARγ1 and PPARγ2) [2]. PPARγ is involved in transcriptional regulation of glucose and lipid homeostasis [1], and helps regulate adipocyte differentiation [3]. It has a , which allows it to interact with a wide array of ligands. typically triggers a conformational change of PPARγ, notably in the activation function-2 , which aids in the recruitment of co-regulatory factors to regulate gene transcription. PPARγ can form a with retinoic X receptor alpha (RXRα), a process necessary for most PPARγ-DNA interactions [4]. PPARγ is a molecular target for antidiabetic drugs such as thiazolidinediones (TZDs), which makes the protein a target for Type II Diabetes (T2D) drug research. Due to its involvement in metabolic and inflammatory processes, PPARγ also holds potential for treatments of many metabolic and chronic-inflammatory diseases, such as metabolic syndrome and inflammatory bowel disease, respectively. Errors in PPARγ-related regulation have also been implicated in atherosclerosis and various cancers, like colorectal, breast, and prostate cancers.

Overall Structure and Ligand Binding

PPARγ is composed of the ligand-independent activation domain (AF-1 region and A/B-domain), a DNA-binding domain (DBD) (C-domain), a hinge region (D-domain), and a ligand-dependent ligand-binding domain (LBD) (E/F-domain and AF-2 region) [5]. The two PPARγ isoforms, PPARγ1 and PPARγ2, differ by only 30 amino acids at the N-terminal end. These added amino acids on PPARγ2 result in increased potency and adipose-selectivity, which makes this protein a key player of adipocyte differentiation [3]. The is composed of 13 α helices and 4 short β strands [1]. It has a T-shaped binding pocket with a volume of ~1440 Å3 [1, 6], which is larger than that of most nuclear receptors [7], allowing for interactions with a variety of ligands [8]. The PPARγ LBD is folded into a helical sandwich to provide a binding site for ligands. It is located at the C-terminal end of PPARγ and is composed of about 250 amino acids [5]. Activation by full agonists occurs through hydrogen bond interactions between the S289, H323, Y473, and H449 residues of the PPARγ-LBD [7] and polar functional groups on the ligand which are typically carbonyl or carboxyl oxygen atoms. Agonist binding results in a conformational change of the LBD AF-2 region, which is necessary for coactivator recruitment. This change can either be dramatic or subtle [1], which leads to stabilization of a charge clamp between helices H3 and H12 [9] to aid in associations with the LXXLL (L, leucine; X, any amino acid) motif of the coactivator [1, 10]. Ligand binding of PPARγ is regulated by communication between the N-terminal A/B domain, which is adjacent to the DBD, and the carboxyl-terminal LBD [11].

Ligand Activity

PPARγ ligands, fall into one of three categories: full agonist, partial agonist, or antagonist [5]. Full agonists have higher efficacy for activating PPARγ and higher potency [7], and their binding leads to the more dramatic conformational change [1]. Binding of partial agonists leads to the more subtle change [1] and results in lower efficacy and potency [7]. Antagonists do not activate PPARγ, so there is either no conformational change to exclude coactivators or a minor conformational change to accommodate corepressors [9,5]. Natural ligands of PPARγ include fatty acids, eicosanoids, and prostaglandins [4-8,11-13].

Coactivators/Corepressors

The of PPARγ is a groove created by hydrophobic residues of the H3, H3’, H4, and H12 helices [1]. Stabilization of the AF-2 domain is important for coactivator interactions, and is achieved through ligand binding [1]. Upon agonist binding, coactivators and other chromatin-remodeling cofactors, like histone deacetylases, are recruited and transcription is activated [14]. Coactivators can be regulated at the transcriptional and post-transcriptional levels, as well as by protein-kinase cascades [3]. PPARγ can actively silence genes it is bound to by recruiting a corepressor in the absence of a ligand. Once this occurs, an antagonist binds to stabilize the AF-2 region, preventing interactions with coactivators and activation of transcription [9]. Corepressor binding creates a three-turn α-helix corepressor motif important for preventing the AF-2 domain from assuming an active conformation [9]. Common coactivators of PPARγ include CBP/p300, the SRC family, and TRAP220 [3]. Common corepressors include SMART, NCoR, and RIP140 [3].

PPARγ/RXRα

PPARγ shows preferential heterodimerization with RXRα [3]. The asymmetry of PPARγ/RXRα packs positively and negatively charged regions together, and is needed for PPARγ binding to DNA [1]. The LBD and DBD of PPARγ are located close together, whereas the RXRα LBD and DBD are positioned farther apart. This difference in region proximity plays a role in heterodimerization [5]. The PPARγ/RXRα complex associates with PPAR response elements (PPREs) in promoter regions of targeted genes [8]. Each ligand-bound PPAR/RXRα complex will bind to a specific PPRE based on the recruited cofactor [8].

Functions

PPARγ is a key regulator of glucose and lipid homeostasis [1]. PPARγ mediates adipocyte differentiation and alters insulin sensitivity, inflammatory processes, and cell proliferation [11]. Ligand-dependent mechanisms include inhibition of inflammatory cytokine production and macrophage activation [8]. In addition to adipocytes, PPARγ is also found in the retina, cells of the immune system, and colonic epithelial cells [5]. PPARγ controls the expression of various genes, particularly those involved in fatty acid metabolism [5]. Regulated genes include adipocyte fatty acid binding protein (aP2), lipoprotein lipase (LPL), and acyl-CoA oxidase [4]. Tissue-specific deletions of PPARγ lead to insulin resistance, low viability of mature adipocytes, and lipodystrophy [3].

Potential of PPARγ in Disease Treatment

Synthetic TZDs were the first class of PPARγ ligands identified [8]. The most potent and selective agonist of PPARγ from this class is the insulin sensitizer rosiglitazone [8]. T2D is linked to higher levels of free fatty acids (FFAs) and triglycerides in the blood [8], which is a contributing factor for insulin resistance. Treatment with rosiglitazone and other TZDs reduces levels of FFAs and triglycerides, helping to restore insulin sensitivity [8]. TZDs also work by increasing levels of GLUT-4 and decreasing levels of pro-inflammatory cytokines [5]. PPARγ is found in high levels in colonic epithelial cells. The role of PPARγ in these cells may be related to regulation of immune response and colon inflammation [12]. The onset of Inflammatory Bowel Disease is thought to be caused by inflammatory cytokines present in the colon [12]. In patients with ulcerative colitis, colonic epithelial cells displayed impaired expression of PPARγ, an important mediator of aminosalicylate activities in Inflammatory Bowel Diseases [13]. TZD ligands could be implemented to reduce colonic inflammation [12]. Agonists have also been used in the treatment of colitis and psoriasis by inhibiting the inflammatory response of the epithelium and reducing cytokine production [8]. PPARγ inhibits activity of nuclear factor NFκB, which is higher in active ulcerative colitis patients [15]. PPARγ could also be implemented in the treatment of other chronic inflammation-related diseases. Immunomodulatory effects have been found with PPARγ agonists [16]. Rosiglitazone alongside adiponectin reduces renal disease, atherosclerosis, and production of autoantibodies, all of which are characteristic of the inflammatory autoimmune disease Systemic Lupus Erythematosus (SLE) [16]. PPARγ ligands hold potential as cancer treatments [11] due to their ability to inhibit angiogenesis, the process required for the growth and metastasis of solid tumors [8]. PPARγ activators have pro-differentiation and anti-proliferation effects [3]. TZDs have also been shown to inhibit proliferation of human breast, prostate, and colon cancer cells [8].