Nuclear Receptors & Endocrinology Lecture Notes PDF

Course HSS 2305 A Molecular Mechanism of Disease Lecture-17 Nuclear Receptors and Endocrinology Ajoy Basak, Ph. D. Adjunct and Part-time Professor, Pathology and Laboratory Medicine, Faculty of Medicine, U Ottawa, Ro...

Course HSS 2305 A Molecular Mechanism of Disease Lecture-17 Nuclear Receptors and Endocrinology Ajoy Basak, Ph. D. Adjunct and Part-time Professor, Pathology and Laboratory Medicine, Faculty of Medicine, U Ottawa, Roger Guindon Building 451 Smyth Road Ottawa, ON K1H 8M5 Tel 613-878-7043 (Cell) E-mail: [email protected] Alternate: [email protected] Affiliate Investigator, Chronic Disease Program, Ottawa Hospital Research Institute Web: https://med.uottawa.ca/pathology/people/basak-ajoy 1 2 Intracellular Receptors And Endocrine Systems 3  Intracellular Receptors (IR)  Two Types of “IR”  Cytoplasmic Receptors (Present in the Cytoplasm) (also called Nuclear Receptor Type I)  Nuclear Receptors (Present inside the nucleus) (also called Nuclear Receptor Type II)  Both bind to Extracellular Signaling Molecules that can migrate or penetrate through cell membrane or are transported across the cell membrane by other molecules  Signaling molecules for “IR” are lipid soluble 4  Intracellular Receptors  These are used specifically by the steroid hormones (Cortisol, Estrogens and Androgen derivatives like Testosterone, Progestogens etc). Steroid hormones are lipid soluble & diffuse through the cell membrane and bind to “Steroid Receptors”.  This hormone-receptor complex then binds to parts of DNA in the nucleus of the cell called “Hormone Responsive Elements” leading to alteration in gene expression. 5  Nuclear Receptors (NR)  NR’s are different from “Cell Surface Receptors” which are membrane bound via a transmembrane domain.  NRs are special class of soluble cytosolic proteins that function as receptors and bind specifically to  Steroid hormones  Thyroid hormones  Certain other molecules [Vitamins eg Retinoic acid (Vitamin-A), Vitamin-D, etc]  The above binding molecules known as “NR-Ligands” are found in animals but not in Plants, Algae or Fungi.  Their (NR) number in human 48 of which 24 have been described with their ligands. Others are “Orphan Receptors” meaning no known ligands. C-elegans have 270 NRs. 6  Nuclear Receptors (NR)  Unlike regular “Cell Surface Receptors”, NR bind to DNA and regulate the expression of adjacent genes hence are also classified as “Transcription Factors”.  NR-mediated gene expression happens when it binds to a specific “Ligand” (Steroid, Thyroid Hormones or Vitamins) resulting in a conformational change and activation of the receptor. This leads to “Up- (largely) or Down- Regulation” of gene expression.  Unlike regular “Transcription Factors”, NRs possess the ability to directly interact with and control the expression of genomic DNA.  Also known as “Nuclear Hormone Receptors” or “Ligand- Activated Transcription Factors” 7  NR-Ligands  Molecules that bind to Nuclear Receptor (NR)  NR-Ligands are mostly Steroid, Thyroid Hormones and Vitamins  These are lipophilic/highly hydrophobic and readily diffuse across cell membranes via hydrophobic interaction 8  Steroid Hormones as NR-Ligands 5 Major Types: Sex Steroid hormones:  Estrogens (Female hormones)  Androgens (Male hormones)  Progestogens (Female hormones) Non-sex Steroid Hormones  Glucocorticoids  Mineralocorticoids 9  Steroid Hormones as NR Ligands Female Steroid Hormones: Estrogens (Major: Estrone, Estradiol, Estriol) Progestogens (Only naturally occurring is Progesterone) Male Steroid Hormones: Androgens (Major: Testosterone, Androstenedione) All are present in some levels in both male and female Female Steroid Hormones Male Steroid Hormones Nonsex Steroid Hormones (Sex hormones) (Sex hormones) (from cortex of brain) Glucocorticoids Cortisol Corticosterone Mineralocorticoids 10 Non-steroid Hormones as NR Ligands) 11  Types of Nuclear Receptors  Classified according to either “Mechanism” or “Sequence Homology”  Monomer type (Mostly Steroid Hormone Receptors)  Homo or Hetero-Dimer type (Mostly nonsteroid Receptors)  Homodimer: Two identical “NR” homo-dimerizes  Heterodimer: Two different Types of NR hetero-dimerizes; Example: PPAR [Peroxisome Proliferator-Activated Receptor] heterodimerizes with Retenoid X Receptor (RXR); Ligand: Thiazolidinediones (a diabetes medication) 12 Nuclear Receptor Family With Their Two Most Important Domains (DBD & LBD) COUP-TF1: Chicken Ovalbumin Upstream Promoter Transcription Factor 1 13 Estrogens are Female Hormones - Receptors Androgens are Male Hormone -Receptors Comparison of amino-acid sequence for each domain of vertebrates (A) Estrogen receptor α (ESR1) & (B) Androgen receptor (AR). The DNA-binding (DBD) & Ligand-Binding (LBD) domains of both ESR1 & AR are well conserved among vertebrate species. Numbers indicate percent sequence identity compared to the human sequence. Bar indicates 100 amino acids. Note high degree of homology of DBD (shown by arrow) 14 A Typical Steroid Hormone Receptor AD: Auto-activation Domain NLS: Nuclear Localisation Signal. Retinoic Acid (Vitamin A product) Receptor (RAR) Retinoic Acid Receptor (RAR): 5 major functional domains: the N-terminal domain (A/B), which contains a transcriptional Activation Function (AF) (A/B), the DNA-binding domain (DBD) (C), a hinge (flexible) region (D), the ligand-binding domain (LBD) and the C- Terminal domain (F). Regions C & E are most conserved within RAR family, but differences in the ligand binding domain can be exploited for design of receptor-specific ligands. 15  Zn-Finger Domains in DBD AF1 AF2 (LBD) (A-B): Regulatory Domain (Variable): Regulates Transcriptional Activity via interaction of its “AF-1 (Activation Function 1)” region with “AF-2” region of E-domain to produce a robust gene up-regulation. (C): DNA-binding domain: Highly conserved domain containing two zinc fingers that binds to specific sequences of DNA called “Hormone Response Elements (HRE)”. (D): Hinge region: Thought to be a flexible domain that connects the DBD with the LBD. Influences intracellular trafficking and subcellular distribution. (E): Ligand binding domain: Moderately conserved in sequence but highly conserved in structure. Contains three anti parallel alpha helices (“Sandwich Filling") flanked by two alpha helices on one side and three on the other (like “Bread"). The ligand binding cavity is within the interior of the LBD. It contains the Activation Function 2 (AF-2) whose action is dependent on the presence of bound ligand. (F): C-terminal domain: Highly variable in sequence between various nuclear receptors 16 Heat Shock Proteins (HSP) as Co-Receptors to Steroid Receptors  HSP are “Chaperones” that assist proper folding of other proteins  Present in low level but increase in response to stress like sudden temperature increase or other conditions. They stabilize other proteins against “Heat Stress”. Also play roles in protein maturation, activation, translocation & degradation. Other HSP proteins act as co-chaperones  Stabilize proteins required for tumor growth, which is why Hsp90 inhibitors are investigated as anti-cancer drugs  Steroid receptor (“Glucocorticoid Receptor” or “GR”) activity depend on their binding with Hsp90 which maintains in a state capable of binding with hormone  Binding with HSP90, Immunophilin FKBP-51/52 & Dynein proteins allows “GR” to translocate into the nucleus  Once inside the nucleus, GR dimerizes and binds with DNA thereby upregulating gene expression  Other steroid hormones (Estrogen, Androgen, Progesterone, etc) also act in a similar manner 17 Translocation of GR into the Nucleus 90: Hsp90 51: Immunophilin FKBP51/52 Dyn: Dynein proteins Immunophilins are Peptidyl-Pro Isomerase enzymes which are involved in immunosuppresive events. They are targeted by immunosuppressive drugs to prevent Cis-Trans isomerisation of X-Pro bond. 18  Releasing Hormones: Steroid or Peptide Hormones Peptide Hormones Steroid Hormones LH Progesterone, Testosterone ACTH Cortisol FSH Estradiol Releasing Hormones: Produced by Hypothalamus which are capable of accelerating the secretion of a given hormone by Anterior Pituitary Gland - Lutenizing Hormone (LH) - Follicle Stimulating Hormone (FSH) - Adrenocorticotropic Hormone (ACTH) Steroid hormones have receptors that are nuclear or cytoplasmic 19  Biological Roles of NRs & Disease Implications  Work with other proteins to regulate specific gene expression thereby controlling cellular “Development”, “Metabolism” and most importantly “Homeostasis”  Homeostatic State (Maintain pH, Salt concentration, Temperature, Oxygen level). Many illnesses are caused by the disturbances in homeostatic state (also caused by Aging).  NRs are important mediators of many diseases  Play key roles in “Embryonic Development” & “Liver Disease”.  NR-Mutations are responsible for a range of diseases. Mutations in the Androgen Receptor can cause infertility & prostate cancer; Mutations in PPAR [Peroxisome Proliferator-Activated Receptors] can lead to colon cancer & diabetes mellitus and Mutations in Estrogen Receptor can cause breast cancer. 20 21 ENDOCRINE SYSTEM AND DISORDERS 22  Endocrinology  It is defined as the branch of biology and medicine which deal with the endocrine rather than Autocrine or Paracrine systems.  It is particularly associated with the study of hormones (the major message carriers in endocrine system), their specific secretions and the associated disorders and diseases.  The diseases associated with hormones and endocrine systems are Cell proliferation, Growth, Differentiation and Coordination of metabolism, Respiration, Excretion, Movement, Reproduction, and Sensory perception.  These events depend on chemicals and substances synthesized and secreted by specialized cells via endocrine systems and signal molecules called hormones.  Specific glands secrete specific hormones. 23 24  Protein Hormones  Bind cell surface receptors  Triggering an intracellular signaling cascade Examples: GPCR and its hormone ligands TSH, FSH, LH, oxytocin (G-Protein coupled receptor, Thyroid Stimulating Hormone. Follicle Stimulating Hormone, Luteinizing Hormone) 25  Endocrine System Endocrine glands  Regulate body's growth and development  Control function of various tissues  Support pregnancy and other reproductive functions  Regulate metabolism Endocrinologists are medical doctors who specialize in research and treatment of disorders and diseases of the endocrine system. 26  Endocrine System Group of specialized organs and body tissues that produce, store, and secrete chemical substances known as hormones. The major organs of the endocrine system  Hypothalamus  Pituitary gland  Thyroid gland  Parathyroid glands  The islets of the pancreas  Adrenal glands  Testes  Ovaries  Mammary gland  Adipose tissue (involved in mammary gland development) During pregnancy, the Placenta also acts as an endocrine gland in addition to its other functions 27 Endocrine System 28  HYPOTHALAMUS-PITUITARY-ADRENAL (“HPA”) AXIS CRH: Corticotropin Releasing Hormone (Released by Hypothalamus) ACTH: AdrenoCorticoTropic Hormone (Released by Pituitary) Cortisol: A steroid (Released by Adrenal Cortex) (CRH) “HPA” Play a major part in Neuroendocrine system that controls Stress response, Digestion, Immune system, Mood, Emotions, Sexuality, Energy Storage and Expenditure 29  Hormones Endocrine System  Steroids  Proteins  Peptides  Modified Amino acids Hormones are chemical substances in the body that control and regulate the activity of specific cells or organs; Normally secreted by special glands, such as thyroid hormone produced by Thyroid Gland, etc. Hormones are essential for every activity of life, including Digestion, Metabolism, Growth, Reproduction, and Mood control etc. 30  Protein/Peptide Hormones - Derived from inactive precursor molecules - Act as ligands / Signaling molecules - 31  Hypothalamic-Pituitary Hormone-Gland Loops Hypothalamus Releasing Hormone Anterior Pituitary Feedback Feedback Inhibition Inhibition Trophic Hormone Pituitary Hormones Endocrine 1. Growth Hormone Gland 2. Prolactin 3. Thyroid Stimulating Hormone (TSH) Circulating Hormone 4. Adrenocorticotrophic Hormone (ACTH) Trophic Hormone 5. Follicle stimulating Hormone (FSH) Targets other 6. Luteinizing Hormone (LH) Metabolic glands Effects 32  Hormonal Balance hormone hormone 1. Too much hormone Disease or Dysfunction 2. Too little hormone Disease or Dysfunction 33