NUCLEAR HORMONE RECEPTORS

Transcription

1 Transcription Regulation And Gene Expression in Eukaryotes Cycle G2 (lecture 13709) RG Clerc NUCLEAR HORMONE RECEPTORS Classification, type I, II, III DNA and ligand binding Co-factor recruitment NHR in health and disease

2 Transcription Regulation and Gene Expression in Eukaryotes NUCLEAR HORMONE RECEPTORS Brief history of steroid signaling (100 years on) Starling refers to bioactive chemicals extracted from glands as hormones Kendall crystallizes thyroid hormone Kendall and Reichstein complete structural analysis of cortisol from adrenal cortex 1946 Selye coins the term glucocorticoid, needed for survival and response to stress 1949 Hench administers cortisone to arthritic patients with dramatic effects 1950 Kendall, Hench and Reichstein get the Nobel Prize Classical model of steroid hormone action 1986 today - Receptor identification : reverse endocrinology GR, ER, TR, RAR, RXR RG. Clerc, May

3 Nuclear Receptors and Small Lipophilic Ligands The Nuclear Hormone Receptor Family. P. Chambon Academic Press 1991 R.G. Clerc 3

4 The family of nuclear hormone receptors steroid receptors adopted nuclear receptors orphan receptors Members Type I Type II Type III/orphan Glucocorticoid receptor Estrogen receptor Androgen receptor Mineralocorticoid receptor. Progesteron receptor Retinoic acid receptor (RAR) Retinoid X receptor (RXR) Vitamin D3 receptor (VD3R) Thyroid hormone receptor (T3R) Peroxisome proliferator activated receptor (PPAR) Liver X receptor (LXR) Farnesol X receptor (FXR) Pregnane activated X receptor (PXR) Steroid+xenobiotic X receptor (SXR) Benzoate X receptor (BXR) NGFI-B ELP Nurr77 SHP Androstane receptor(car) Localisation Cytoplasma nuclear nuclear nuclear (HSP90 complexation) GR Dimerisation homo hetero (RXR) hetero (RXR) Binding IR 3 DR Single Repeat + extension Mode of action Transactivation systemic Transactivation AP-1 antagonism? DAX LRH1 ERR (48 in human; 230 in C. elegans; 21 in D. melanogaster)

5 Nuclear Hormone Receptors are Modular in Nature (operationally defined from A-F) Ligand independent trx AD AF-1 function DNA binding dimerization Hinge NLS Hsp90 Ligand dependent trx AD dimerization AF-2 function co-regulator recruitment NLS Hsp90

6 C domain (DBD): 2 cys-cys Zinc Fingers eg. ERa Cooperative Binding (homodimer/heterodimer)

7 Homodimer/heterodimer (NR/RXR) formation involves both the C and E domains

8 Nuclear Hormone Receptors are Transcriptonal Regulators P. Chambon and co-workers. IGBMC. Illkirch France

9 The family of nuclear hormone receptor: unified nomenclature (48 in human) (based on the two well conserved domains C and E) Laudet V..J. Mol. Endo. 19:207 (1997) NHR Nomenclature Committee. Cell 97:161 (1999)

10 Steroid Signaling Pathway hormone Membrane receptor? steroid GR GR aporeceptor complex hsp90 hsp90 molecular chaperones dissociation dimerization GR GR active receptor Transfer into nucleus coregulator GR GR GRE POL2 G TF s eg. glucocorticoid responsive genes nucleus cytoplasm Induction of steroid responsive genes involves hormone dependent dissociation of the receptor from hsp90 (type I nuclear receptors)

11 LBD of GR Mediates Translocation to the Nucleus in presence of Hormone

12 Steroids and the Adrenal Cortex GLUCOCORTICOID The adrenal cortex is responsible for production of 3 major classes of steroid hormones: glucocorticoids, which regulate carbohydrate metabolism; mineralocorticoids, which regulate the body levels of sodium and potassium; and androgens, whose actions are similar to that of steroids produced by the male gonads MINERALOCORTICOID ESTRADIOL/TESTOSTERONE

13 PPAR/RXR-dependent nuclear signaling (type II nuclear receptors) PPAR - Peroxisome Proliferator Activated Receptor RXR - Rexinoid Receptor Fibrates TZDs Prostaglandins PUFAs RXR Rexinoids PPAR coregulator PGC-1 POL2 RXR apo A-I, II apo C-III aco P450 LPL PPRE NHR are the final effectors of a complex cytoplasmic/nuclear transduction cascade

14 E domain: canonical structure of the LBD Characteristical sandwich architecture with 3 layers built in by 12 alpha antiparallel helices and 1 antiparallel beta sheet Structure-AA sequence relationship for NHR LBD s Ligand binding dependent pocket remodeling Receptor dimerization surface Co-regulator proteins interface Control of agonist vs. antagonist modes of action

15 Nuclear Hormone Receptor Superfamily: Well Conserved DBD, Poorly Conserved LBD DNA-Binding Domain Ligand Binding Domain Progesterone Receptor N 100% 100% C NGF1B N 40% 18% C Retinoid X Receptor N 35% 18% C Thyroid Hormone Receptor N 28% 16% C well conserved at sequence level well conserved at structural level poorly conserved at sequence level well conserved at structural level

16 Corepressor vs. Coactivator Interfaces Structure Remodeling mouse trap unliganded co-repressor binding liganded co-activator binding

17 Protein:protein interaction in vivo screen: two hybrid-screen (b) S. Fields. Nature 340:245 (1989)

18 Coactivator CBP/p300, Corepresssor Ncor, etc Combinatorial roles of multiple cofactor complexes are required to switch between transcriptional repression and activation functions

19 Coactivator Family PGC1a, 1b and PRC Structure and function of the PGC-1 family coregulators: binding to the HAT and TRAP/DRIP/Mediator complexes at the amino and carboxyl termini, respectively. SirT1 and p160 bind to the repression domain, which also contains three p38 MAP kinase phosphorylation sites J. Lin, C. Handschin, BM. Spiegelmann. Cell Met 1:361 (2005)

20 NR PGC-1 Coactivator Functions in Maintenance of Glucose, Lipid and Energy Homeostasis Hormones PGC-1 other NRs FOXO1 PGC-1 GR /HNF4 txn PPAR TR/ PPAR ERR/ gluconeogenesis fatty acid oxidation mitochondrial biogenesis respiration

21 Coactivator and Corepressor Complexes with the Basal Machinery are Involved in the Regulation of NHR Transcriptional Activity Remodelling Complexes Acetylase (HAT) Complexes Mediator Complexes ACTIVATION REPRESSION Deacetylase (HDAC) Corepressor Complexes

22 Nuclear Receptor Coregulator Interaction Motifs

23 Nuclear receptors and coregulators manifest in reproduction, development, central and basal metabolism and energy homeostasis Bookout AL, Evans RM, Mangelsdorf DJ. Cell NURSA - Nuclear Receptor Signaling Atlas (

24 Molecular Bases for Agonism vs partial Agonist vs Antagonism: Selective Modulator Concept eg. ER Co-regulator box LXXLL

25 Estrogen : Hormone with Ambivalent Functions

26 Molecular Action of Estradiol and of SERM Tamoxifen Adapted from Howell A, Osborne CK, Morris C, Wakeling AE. ICI 182, 780 (Faslodex ), development of a novel, "pure" antiestrogen. Cancer 2000; 89: 819.

27 Selective Estrogen Receptor Modulators Estrogens Phytoestrogens SERMs SERMs- designed to act in specific ways at each of the estrogen receptor sites in different tissues Anti Estrogens ERDR

28 Integrated Physiology by PPAR Isoforms -b

29 Peroxisome proliferator-activated receptors PPAR Isoforms Inducing the proliferation of peroxisomes in rodents Intimately connected to the cellular metabolism and cell differentiation

30 Ligands and Functions of the PPAR and - Isoforms PPAR PPAR POLYUNSATURATED FATTY ACIDS Docohexaenoic acid, eicosapentaenoic acid, linoleic acid, linolenic acid and arachidonic acid FIBRATES Gemfibrozil, Benzafibrate, Fenofibrate THIAZOLIDINEDIONES Rosiglitazone, Pioglitazone Ciglitazone HDL RAISE, LIPID CATABOLISM PEROXISOME PROLIFERATION CONTROL OF INFLAMMATION GLUCOSE HOMEOSTASIS LIPID STORAGE ADIPOCYTE DIFFERENTIATION CONTROL OF INFLAMMATION

31 Ligands and Functions of PPARb PPARb/ POLYUNSATURED FATTY ACIDS PROSTAGLANDINS GW DIFFERENTIATION of oligodendrocytes, epithelial cells, keratinocytes and adipocytes LIPID METABOLISM IN THE BRAIN EMBRYO IMPLANTATION AND DECIDUALIZATION TUMORIGENESIS IN THE COLON REVERSE CHOLESTEROL TRANSPORT WOUND HEALING

32 Synthetic PPAR s Ligands thiazolidinediones (TZDs) treatment of Type II Diabetes PPAR alpha specific GW 7647 PPAR beta specific GW EC 50 (mm) O O N O S S N O with nm affinity

33 Correlations between PPAR -activity, insulin sensitivity and adipogenesis: The relationship between PPAR -activation and adipogenesis is linear, while bell-shaped with insulin sensitivity Thus, neither PPAR antagonism (lipodystrophic state) nor full agonism (obese state) results in optimal insulin sensitization (mouse/rat/human genetic models)

34 Selective modulator concept: SPPARMs agonists < partial > antagonists SRC1, PGC1 PPAR 1 RXR ligand-specific genomic & non-genomic effects TIF2, PGC1, DRIP/ TRAP PPAR 2 RXR Ligand-dependent differential coactivator/corepressor recruitment ligand-specific expression of LEAN genes ligand-specific expression of FAT genes

35 Selective Modulator Concept Gave boost to the continued research for SERMs. Concept of profiling selective modulators has been established since then eg. (SFXRM s, SPPARM s, SLXRM s) Ligand dependent selective co-regulator recruitment likely to elicit specific target gene repertoire linked to desirable pharmacological effects, (eg: LXR ligands which promote reverse cholesterol transport but do not induce hypertriglyceridaemia (SREBP1c gene pathway activation)

36 PPAR subtypes and expression patterns PPAR - cardiac muscle/glucogenesis tissues, liver, intestine, renal cortex PPARb - ubiquitous/skeleton muscle PPAR - adipose tissue, large intestine, immune cells 1 - predominant form in above tissues 2 - adipose tissue only 3 - macrophage and large intestine note: 2 has a distinct N-terminal extension

37 Association of PPAR polymorphisms with the metabolic syndrome

38 PPAR Gene Three mrnas via two promoters and alternative splicing 1and 3 encode the same protein, 2 is distinct identical ligand binding domains 28 aa N-terminal addition in PPAR 2 increases ligand-independent activation 5X PPAR 1 PPAR 2 PPAR 3 Tissue Distribution adipose, intestine, kidney, liver, muscle adipose - elevated in obese subjects very low in muscle & liver macrophage, adipose, colon epithelium

39 PPAR - critical role in adipogenesis adipogenic signals Dominant negative mutants and chemical inhibitors block agonist-mediated adipocyte differentiation Ectopic expression of PPAR converts fibroblasts to adipocytes PPAR KO mouse: complete absence of adipose tissue

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41 Association of PPAR polymorphisms with the metabolic syndrome? The Pro12Ala substitution in PPAR 2 associated with decreased receptor activity, lower body mass index and improved insulin sensitivity. Proline to Alanine substitution at codon 12 of PPAR 2 The codon Ala confers reduced activity compared to the Pro isoform Auwerx J. and co-workers Nat Genet.3:284

42 Association of PPAR polymorphisms with the metabolic syndrome? Haplotype analysis of the PPARgamma Pro12Ala and C1431T variants reveals opposing associations with BMI Doney et al BMC Genetics 3:1-8.