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MD Consult: Books: Goldman: Cecil Medicine: HOW HORMONES WORK

Goldman: Cecil Medicine, 23rd ed.

Copyright © 2007 Saunders, An Imprint of Elsevier

HOW HORMONES WORK

Two classes of hormones operate via two distinct types of receptors ( Fig. 240-1 ). Peptide hormones are synthesized as parts of larger protein molecules and are processed to smaller proteins that are secreted. They act through receptors located in the cell membrane with the recognition and binding site exposed on the cell surface and the activity domain facing the inside of the cell. Activated cell surface receptors use a variety of strategies to transduce signal information, and they often activate second messengers that amplify and distribute the molecular information. Many peptide hormones ultimately signal by regulation of protein phosphorylation, a process through which proteins are covalently modified when a phosphate group is donated to the protein by adenosine triphosphate (ATP). This allows peptide hormones to change the conformation and thus the function of existing intracellular signaling intermediates rapidly. It also induces somewhat slower changes in gene transcription, which regulate the concentration of regulatory cellular proteins (e.g., enzymes). Biogenic amines function like peptide hormones and bind to cell surface receptors.

FIGURE 240-1  Mechanisms by which peptide and steroid hormones signal.

Steroid hormones are synthesized from precursor cholesterol. Thyroid hormone, retinoic acid (vitamin A), and vitamin D are synthesized through separate pathways but act through the same family of receptors and mechanisms as do steroid hormones. This group of hormones acts through structurally related receptors that bind to DNA recognition sites to regulate transcription of target genes. This results in changes in the concentration of cellular proteins, primarily enzymes, and thus in the metabolic activity underlying the physiologic response.

Peptide Hormone Binding and Signal Transduction

Peptide hormone receptors have one of three general structures ( Fig. 240-2 ): (1) a seven-membrane-spanning structure in which the recognition site is formed by exterior sequences between membrane-spanning helices and the activity site is formed by interhelical regions inside the cell; (2) a single membrane-spanning helical structure separating the extracellular recognition domain from the cytoplasmic domain, which contains an intrinsic enzyme activity; and (3) a single membrane-spanning helix that separates the recognition domain from an intracellular domain that couples to second messenger systems, as do the seven-membrane-spanning receptors. The protein coupled may be an intracellular tyrosine kinase or another enzyme.

FIGURE 240-2  Structures of peptide hormone receptors.

Hormone ligands and receptors bind with high affinities (equilibrium dissociation constants of nanomolar to picomolar), thus providing the specificity necessary for cells to decode the information provided by the low concentration of hormone present among the many other circulating and extracellular proteins. The conformational change resulting from peptide hormone binding activates receptors to signal from the cell surface. Removal of receptors from the cell surface results in downregulation and attenuation of the response. Binding affinities and dose-response curves for the initial event in cell signaling are the same. Biologic responses consequent to these initial events occur through a series of amplifications. The result is a dose-response curve for a biologic response that is more sensitive than that for binding and initial activation of the receptor. Full biologic responses may thus occur at a low concentration of hormone, with a resulting occupancy of only 10% of receptors or less. This provides high sensitivity to small changes in hormone concentration. It also provides significant reserve. Hormone-induced downregulation may remove 90% of receptors from the cell surface. This process renders the cell refractory to the initial hormone concentration, but if the need is great enough, hormone concentrations can increase and can activate the residual 10% of receptors, thereby increasing the biologic response. Such a response system provides high initial sensitivity, buffering through downregulation against excessive hormone responses, but it also leaves a reserve that can operate when the signal strength is strong enough.

Receptors are mobile in the plane of the membrane. Following ligand binding, the mobility is reduced, and they become relatively fixed while transducing signals. Ligand binding may induce sequestration of receptors and may result in their retention inside the cell through interactions with cell proteins, as occurs with rhodopsin and adrenergic receptors. Ligand binding may induce endocytosis through clathrin-coated pits with ultimate degradation of receptors by lysosomal enzymes, as occurs with the insulin and epidermal growth factor receptors. The concentration of cell surface receptors is regulated by interaction with hormone ligand and by other signals that regulate their synthesis and affinity. The concentration of receptors is an important determinant of the responsiveness of cells. Antagonists compete with hormones for binding to receptors, but in general they do not induce desensitization. When antagonists are removed, receptor concentrations are high, and cells are very responsive to hormone exposure. The changes can result in clinically significant changes, such as excessive adrenergic responses when β-blockers are rapidly withdrawn, or downregulation of insulin receptors resulting in insulin resistance in type 2 diabetes. Regulation of receptor synthesis is an important mechanism by which one hormone regulates responsiveness to another to coordinate their biologic effects.

A class of cell surface receptors serves to modulate nutrient delivery rather than perform an informational function. These molecules include the low-density lipoprotein (LDL) receptor and the transferrin receptor. LDL and transferrin receptors, which are clustered in coated pits, internalize, deliver LDL (cholesterol) and iron to the cell interior, and then recycle to the cell surface. Such receptors are not downregulated by ligand but undergo repeated rounds of recycling to provide the cell with essential nutrients. The concentration of these receptors is regulated in response to changes in the metabolic state.

Intracellular Second Messengers

Cyclic Adenosine Monophosphate and Cyclic Guanosine Monophosphate

The concept of second messengers was established by Earl Sutherland, who discovered cyclic adenosine monophosphate (AMP), an intracellular allosteric effector that mediates the action of many peptide hormones. Hormone receptors are coupled to catalytic adenylate cyclase through guanosine nucleotide binding (G) proteins, the β-adrenergic receptor being a paradigm for this signaling pathway ( Fig. 240-3 ). More than 600 human genes encode receptors of this class and mediate responses to a variety of ligands and physical stimuli (hormones, biogenic amines, light, sound, touch, pain, taste, smell). These receptors belong to the seven-membrane-spanning class. On ligand binding, the receptor interacts with a G protein trimer consisting of α, β, and γ subunits. Because G proteins bind guanosine diphosphate (GDP) with higher affinity than guanosine triphosphate (GTP), guanine nucleotide exchange is triggered by proteins that facilitate exchange of GTP for GDP; activity is reversed by hydrolysis of GTP to GDP. Binding of hormones to receptors that operate through the cyclic AMP second messenger system results in a conformational change and causes receptors to bind to G proteins. Ligand-activated receptors facilitate exchange of GTP for GDP, so the activated Gαs (the stimulating GTP-binding subunit) dissociates from the β and γ subunits. The [ligand·hormone receptor]·[Gαs·GTP] complex activates adenylate cyclase to catalyze formation of cyclic AMP from ATP. Each hormone ligand induces formation of multiple cyclic AMP molecules through this mechanism. Inhibitory G proteins operate in a similar manner to decrease cyclic AMP formation. In both cases, ligand-activated receptors act to exchange GTP for GDP, analogous to proteins that catalyze this process to regulate protein synthesis.

FIGURE 240-3  G protein–coupled signal transduction. Various hormones and other stimuli bind to seven-membrane-spanning receptors that activate G protein switches by exchanging guanosine diphosphate for guanosine triphosphate (GTP). GTP-bound G proteins then couple to a variety of signaling machines that transmit information. GTP hydrolysis turns off the switch.  (Adapted from Bockaert J, Pin JP: Molecular tinkering of G protein–coupled receptors: An evolutionary process. EMBO J 1999;18:1723-1729, by permission of Oxford University Press.)

Adenylate cyclase is a large complex molecule with a 12-membrane-spanning structure. The two large cytoplasmic domains have internal sequence similarities and are related to sequences in guanylate cyclase. Activation of adenylate cyclase is buffered and terminated by several mechanisms. First, hormone dissociates from receptor. Binding of Gα·GTP to the receptor decreases affinity for hormone approximately one order of magnitude to facilitate this dissociation. Second, receptors desensitize and are removed from the cell surface by a process involving phosphorylation and interaction with cell proteins termed arrestins. If hormone exposure is short, receptors are dephosphorylated and reappear on the cell surface; if exposure is prolonged, receptors are degraded, and resensitization requires synthesis of receptors. Third and most importantly, Gα proteins possess intrinsic GTPase activity so GTP is hydrolyzed to GDP, and, on GDP binding, Gα is inactivated and reassociates with the β/γ subunits. A family of proteins, the regulators of G protein signaling (RGS), enhances this GTPase activity to facilitate the “off” switch. Some RGS proteins are specific for the various G proteins (Gαs, Gαi, Gαq, Gα12).

There are many consequences when this mechanism of signal transduction is perturbed. Mutations in seven-membrane-spanning receptors may inactivate so signaling is defective; some mutations, such as those observed in thyroid-stimulating hormone (TSH) receptors in hyperfunctioning thyroid nodules, may activate so that receptors signal in the absence of hormone. Continuous exposure to hormone results in desensitization or tachyphylaxis. Deficiency of a G protein, which occurs in certain forms of pseudohypoparathyroidism, results in insensitivity to hormone. Cholera toxin, which activates adenosine diphosphate (ADP) ribosylation of Gαs, inhibits GTPase activity and interferes with reversibility so profound and prolonged elevations in cyclic AMP occur. Mutations in Gα proteins that are predicted to impair GTPase activity have been described in tumors that occur in endocrine glands.

Cyclic AMP, an intracellular allosteric effector, binds to the regulatory subunit of cyclic AMP–dependent protein kinase A. This kinase is a tetrameric protein consisting of two regulatory and two catalytic subunits. Binding of cyclic AMP dissociates the inhibitory regulatory subunits as a dimer from the two catalytic subunits. The latter then catalyzes the transfer of the γ phosphate of ATP to serine and threonine residues in proteins. This covalent modification by phosphorylation causes an allosteric conformational change in the substrate protein that results in a change in its activity. The hormonal signal is transduced into an alteration in enzyme activity and thus in cellular function. Phosphorylation of cytoplasmic proteins results in alterations such as glycolysis; the activated catalytic kinase subunit also migrates to the nucleus to phosphorylate and activate transcription factors such as the cyclic AMP response element–binding protein (CREB).

Cyclic AMP actions are reversed by hydrolysis of cyclic AMP by phosphodiesterase to 5′ AMP, and protein phosphorylation is reversed by the action of phosphatases. Phosphodiesterases are regulated and are a frequent target of inhibitor drugs such as methylxanthines, which prolong cyclic AMP action by blocking its degradation. Phosphatases are regulated by phosphatase-inhibitor proteins, which are refined by phosphorylation of these molecules.

A conceptually similar but structurally distinct system provides signal transduction through the second messenger cyclic guanosine monophosphate (GMP). Two forms of guanylate cyclase catalyze formation of cyclic GMP from GTP. The best characterized mammalian enzyme is the receptor for atrial natriuretic hormone. The binding site for atrial natriuretic hormone is located on the extracellular portion of its receptor and is separated by a single membrane-spanning domain from the cytoplasmic guanylate cyclase (see Fig. 240-2 ). In contrast to adenylate cyclase, receptor and catalytic activities reside in the same molecule. Activity is regulated primarily by ligand binding but also depends on phosphorylation of the enzyme, with dephosphorylation causing desensitization. Cyclic GMP acts by binding to the regulatory domain of cyclic GMP–dependent protein kinase. G-kinase, a dimeric enzyme that is evolutionarily related to A-kinase, is allosterically activated on cyclic GMP binding. Like A-kinase, it catalyzes protein phosphorylation to alter enzyme function and physiologic responses. Reactions are terminated by cyclic GMP phosphodiesterase and protein phosphatases. Cyclic GMP phosphodiesterase is activated by binding of calcium and calmodulin. This is an example of a mechanism that provides biochemical communication between two signaling systems.

Calcium and Diacylglycerol

Hormone receptors that activate the phosphatidylinositol (PI) cycle transmit information to the interior of the cell by two second messengers: calcium (Ca2+) and diacylglycerol (DAG). The cycle of PI me-tabolism consists of synthesis of this phospholipid, its breakdown, and its resynthesis. PI is composed of a three-carbon glycerol backbone with long-chain fatty acids esterified at carbons 1 and 2 and an inositol ring esterified by a phosphoester bond at carbon 3. Distinct kinase enzymes catalyze phosphorylation of the inositol ring at positions 3, 4, and 5. Quantitatively, the principal phosphorylations occur sequentially at positions 4 and then 5. The principal function of activated hormone receptors is to stimulate phosphoinositidase (phospholipase C), which releases the phosphorylated inositol to generate inositol trisphosphate (IP3, inositol 1,4,5 P3) and DAG (the glycerol backbone with fatty acids attached at carbons 1 and 2). IP3 increases the concentration of cytoplasmic Ca2+. It mobilizes stored intracellular Ca2+ by binding to specific receptors on intracellular membranes and by facilitating opening of calcium channels. The concentration of basal cytoplasmic Ca2+ is at least 1000-fold less than that in storage sites and outside the cell. The release from intracellular stores or entry of Ca2+ into the cell rapidly increases cytoplasmic Ca2+.

Ca2+ plays a regulatory role in muscle contraction, in neuromuscular transmission, and in hormone signaling. Ca2+ binds to calmodulin and alters its conformation, causing the Ca2+·calmodulin complex to bind to a variety of enzymes to regulate their activities. Ca2+·calmodulin regulates protein kinases, including myosin light chain kinase involved in smooth muscle contraction, phosphorylase kinase involved in breakdown of glycogen, and calmodulin-dependent protein kinase important in synaptic transmission. Ca2+·calmodulin regulates cyclic nucleotide phosphodiesterase and adenylate and guanylate cyclases to influence cyclic AMP and cyclic GMP concentrations, and it is involved in microtubule assembly and disassembly. Ca2+·calmodulin is thus able to bind to a variety of other proteins and to alter their activity in response to information provided by the cytoplasmic Ca2+ concentration.

DAG acts as a second messenger by binding to protein kinase C to activate this important regulatory enzyme. Protein kinase C also requires Ca2+ for activation, so both second messengers of this pathway coope-rate to increase the activity of this enzyme. Tumor promoters, such as active phorbol esters, are DAG analogues and act through protein kinase C.

The components of this second messenger system are diverse and complex. There are several isoenzyme forms of protein kinase C and of phosphoinositidase. Although one isoenzyme form of phosphoinositidase is activated by receptor-coupled G proteins, another is activated by binding to receptor tyrosine kinases and undergoing tyrosine phosphorylation. Additional kinases phosphorylate alternate positions on the inositol ring; PI3-kinase is activated by certain tyrosine kinases to yield unique PI metabolites with functions distinct from Ca2+ mobilization. Phosphatidylinositol (PtdIns), which is phosphorylated at the 4,5 position, functions in recognizing PH domains to localize and activate kinases and other proteins, and phosphatidylinositol (3) phosphate (PtdIns [3] P) binds to FYVE in proteins. PX domains also bind to phosphorylated PtdIns. PtdIns kinases thus create membrane docking sites for signal transduction complexes that assemble through proteins that contain PH, FYVE, and PX domains. This process also assembles protein complexes that direct trafficking through the membrane compartments of the cell. Sphingosine, a component of glycosphingolipid metabolism, inhibits protein kinase C, which provides dual regulation of this protein. A major mechanism to regulate inositol phosphate function is provided by specific phosphatases that remove the phosphate groups from the inositol ring to terminate its activity. PtdIns 3 phosphatase is one such phosphatase, and deletion of the gene, which occurs frequently in cancer, removes an essential “off” switch, thus leaving growth and survival signals in the “on” position. Lithium has the opposite effect because it blocks the activity of one of these phosphatases to enhance accumulation of the biologically active inositol phosphates. Like other information pathways, this one generates coordinated cellular responses and is buffered and ultimately turned off when the signal strength decreases.

Protein Tyrosine Kinases

A group of peptide hormone receptors (e.g., this insulin receptor) contains intrinsic protein tyrosine kinase activity. Ligand binding to the extracellular domain results in an allosteric change that is transmitted across the single membrane-spanning segment to activate the cytoplasmic kinase domain (see Fig. 240-2 ). In a second structural motif, a transmembrane receptor is coupled to a separate and distinct protein localized in the cytoplasm that contains tyrosine kinase activity. The growth hormone receptor and JAK2 kinase belong to this second class.

Within the cell, the majority of protein-bound phosphate is attached to serine and threonine residues, with only a small fraction attached to tyrosine. Numerous kinases, however, covalently modify tyrosine residues in proteins as a central regulatory function in cell proliferation, developmental processes, and differentiated function. The extracellular ligand binding domains of receptors of this class contain cysteine-rich regions that create the binding sites either as monomers (epidermal growth factor receptor) or as dimers (insulin receptor) or contain immunoglobulin-like structures (platelet-derived growth factor and fibroblast growth factor receptors). The cytoplasmic protein tyrosine kinase domains are highly homologous and contain ATP and substrate binding sites, but different receptors recognize distinct substrates to give specific biologic responses. For example, insulin stimulates glucose uptake, whereas epidermal growth factor stimulates cell proliferation. The tyrosine kinases contain variable domains on both sides of the tyrosine kinase core as well as inserts within the kinase domain, which provide regulatory sites that modulate ligand-activated tyrosine kinase activity.

Information received by a cell surface tyrosine kinase receptor is transmitted through a signal transduction pathway that begins with direct physical coupling of the receptor ligand and proceeds through the GTP-binding protein ras ( Fig. 240-4 ). In response to ligand binding, receptor tyrosine kinases either self-phosphorylate or phosphorylate a linker substrate. Proteins that contain a 100-amino acid domain homologous to a region in src, SH2, bind tightly to these sites of tyrosine phosphorylation. The growth factor receptor binding protein 2 (Grb2) is a molecular coupler containing an SH2 domain that plugs into a tyrosine phosphorylation site. Shc is another molecular coupler that is frequently used. Grb2 also contains two SH3 domains that act as a receptacle for proline-rich domains of the guanine nucleotide exchange protein SOS. These high-affinity protein-to-protein interactions bring SOS to the cell membrane where ras is present in its inactive GDP-bound form. Activated GTP-bound ras then couples to a serine/threonine protein kinase cascade involving first raf-1, then MEK and MAP (mitogen-activated protein) kinases. Information is thus relayed, expanded, and is fused ultimately to control gene expression and cell division. Operative mechanisms for this, as for other hormone-signaling pathways, include ligand or protein-protein interactions, activated GTP-bound G proteins, and protein phosphorylation. Receptor tyrosine kinases also couple to additional signaling pathways through SH2 domains in other proteins and through tyrosine phosphorylation of these proteins including phospholipase C-γ, transcription control proteins termed signal transducers and activators of transcription (STAT), and PI3-kinase. Different hormone receptors utilize different combinations of the signaling elements to activate their target cell responses. For example, the growth hormone receptor uses the JAK-STAT pathway, whereas the insulin receptor uses insulin receptor substrate 1 (IRS-1)/Grb-2 RAS and MAP kinase.

FIGURE 240-4  Information transfer through a receptor tyrosine kinase pathway. Sites of receptor tyrosine self-phosphorylation, Y-P, are recognized by the SH2 domain of the linker Grb2, which brings the guanine nucleotide exchange factor SOS to the membrane where ras is located. Activated guanosine triphosphate (GTP)–bound ras initiates signaling by contacting raf, a serine threonine kinase, to initiate a cascade of kinase activations.

Increased tyrosine kinase activity is reversed by four principal mechanisms: (1) ligand-induced endocytosis and downregulation of surface receptors; (2) tyrosine phosphatases, which specifically remove phosphate from tyrosine residues; (3) reversal of the kinase reaction to transfer the phosphate from tyrosine residues in protein to ADP; and (4) hydrolysis of ras-bound GTP to GDP.

Regulation and reversibility of ligand-activated tyrosine kinases are important. Mutations involving these proteins occur frequently in cells transformed from normal to cancerous patterns of growth. These mutations can bypass the normal regulatory mechanisms and result in constitutive kinase activation. The kinases may be overexpressed, most frequently owing to gene amplification but also to enhanced transcription, or the ligand may be constitutively expressed, resulting in continuous receptor activation. Mutant ras proteins may be constitutively active owing to decreased GTPase activity or to a defect in a protein that stimulates the GTPase activity of ras. Any of these changes converts a normal regulatory protein into an oncoprotein, one capable of causing neoplastic transformation.

Steroid Hormones Act through Nuclear Receptors

Superfamily of Steroid Hormone Receptors

All steroid hormone receptors share structural similarities indicative of a common ancestral molecule. Some bind their steroid ligand in the cell cytoplasm and move into the nucleus, whereas others encounter their ligand in the nucleus (see Fig. 240-1 ). The most conserved structural feature is the DNA-binding domain that contains zinc “fingers.” The spatial location of cysteine residues within this domain creates a structure coordinately linked to a zinc atom that forms a helix that binds to the major groove of DNA. Because the energy of protein-DNA interaction depends on the area of contact, most proteins bind DNA as complexes. Steroid hormone receptors of the glucocorticoid receptor subfamily bind to DNA as homodimers; receptors of the thyroid hormone receptor subfamily may bind as homodimers, but they more commonly bind as heterodimers with a common partner, the retinoid X receptor (RXR) ( Fig. 240-5 ).

FIGURE 240-5  How steroid hormone receptors work. Left, Glucocorticoid receptor family members bind as homodimers to palindromic DNA sites. Thyroid hormone receptor family members bind primarily as heterodimers with retinoid X receptor to direct repeat DNA sites separated by varying numbers of base pairs. Right, As a result of hormone binding, repressor complexes dissociate and activator complexes bind to nuclear receptors. Repressor complexes contain histone deacetylase (HDAC) and activator complexes contain histone acetylase (CAF). The coactivator and corepressor complexes contain multiple proteins; only a few are shown.

The DNA recognition element consists of two half-sites of six base pairs, each half binding one monomer surface of the dimeric receptor protein. The half-sites are arranged as direct, inverted, or everted repeats. Receptors of the glucocorticoid receptor subfamily most often bind to palindromic sites, whereas receptors of the thyroid hormone receptor subfamily most often bind to sites made up of directly repeated DNA sequences. Small variations in the DNA-binding domain and in the DNA recognition element provide specificity for hormone action. One important determinant for receptor binding and activity is the spacing between the two half-sites for dimeric receptor binding. The spacing rules for DNA recognition elements that are arranged as direct repeats (DR) indicate that a spacing of 1 (DR + 1) directs RXR homodimer binding and 9-cis-retinoic acid responses, DR + 3 directs vitamin D receptor·RXR binding and vitamin D responses, DR + 4 directs thyroid hormone receptor·RXR binding and thyroid hormone responses, and DR + 5 directs retinoic acid receptor·RXR binding and all-trans-retinoic acid responses. RXR binds to the upstream half, and the hormone-specific receptor binds to the downstream half of these DNA response elements to mediate hormone-dependent changes in transcription. Spacing between these sites is crucial for binding homodimeric receptors of the glucocorticoid receptor class, but the sequence of the half-site also provides an essential discriminant. Specificity is quantitative, not absolute. For example, progesterone receptors bind to glucocorticoid response elements, and retinoic acid receptors bind to thyroid hormone receptor DNA response elements. Specificity is sufficient for generating hormone-specific responses but may permit overlapping functions as in ligand-activated progesterone receptor induction of glucocorticoid-regulated genes.

Hormone binding activates the biologic function of the receptor. Cortisol receptors exist in inactive complexes with other proteins; cortisol binding induces an allosteric change that facilitates dissociation, thus allowing the ligand-bound receptor to bind to DNA. Thyroid hormone and retinoic acid receptors exist bound to DNA rather than complexed to protein; hormone binding results in an allosteric change that activates the receptor, so it interacts with other components of the transcription machinery. Binding of triiodothyronine (T3) to the thyroid hormone receptor results in dissociation of a repressor complex that binds to the empty receptor and binding of an activator complex to the liganded receptor. The thyroid hormone receptor and activator interact with proteins such as the CREB-binding protein (CBP) that integrate information from multiple transcription factors, including CREB, STAT, and the thyroid hormone receptor family.

The steroid hormone receptor family is large and includes subfamilies of receptors: at least six for retinoic acid, two for thyroid hormone, several for 1,25(OH)2 vitamin D and for fatty acids or metabolites causing peroxisome proliferation, and a group of “orphans” whose ligands remain to be identified. As ligands are identified, many orphan receptors are being adopted. Among these are the receptors that heterodimerize with RXR and regulate cholesterol, bile acid, and xenobiotic metabolism. Metabolism of cholesterol to bile acids in the liver, the major route for cholesterol catabolism, is stimulated by oxysteroids acting through oxysterol (lipid) receptors (LXR) ( Fig. 240-6 ). LXRα stimulates transcription of the cytochrome P-450 CYP7A gene that catalyzes formation of bile acids. The bile acid receptor farnesoid X receptor (FXR), in turn, blocks this activation to provide a feedback loop between cholesterol catabolism and bile acid formation. This receptor also regulates the transport proteins that facilitate bile acid uptake and egress in gut and liver.

FIGURE 240-6  Steroid receptor family proteins regulate multiple metabolic processes. CYP genes are cytochrome P-450 proteins involved in hepatic metabolism and steroid endocrine gland hormone biosynthesis. Peroxisome proliferator–activated receptors (PPARs) are activated by polyunsaturated fatty acids, prostaglandins, eicosanoids, and thiazolidinediones. Oxysterol (lipid) receptors (LXRs) are activated by oxidized derivatives of cholesterol. Farnesoid X receptor (FXR) is activated by bile acids. Pregnane X receptor (PXR) is the rodent orthologue of human SXR, the steroid xenobiotic receptor that binds a variety of xenobiologic ligands. Constitutive androstane receptor (CAR) responds to phenobarbital-like inducers. Classic steroid receptors for androgens (AR), estrogens (ER), glucocorticoids (GR), mineralocorticoids (MR), progesterone (PR), vitamin D (VDR), and retinoic acid (RAR, RXR) are indicated.  (Adapted from Chawla AJ, Repa J, Evans RM, Mangelsdorf D: Nuclear receptors and lipid physiology: Opening the X-files. Science 2001;294:1866-1870, with permission.)

Two receptors that regulate detoxification and elimination of toxic endogenous substances and xenobiotics are the constitutive androstane receptor (CAR), which mediates responses to phenobarbital-like inducers by enhancing transcription of the CYP2B gene, and steroid and xenobiotic receptor (SXR)/pregnane X receptor (PXR), which senses xenobiotics and induces CYP3A gene expression, thus metabolizing more than 50% of prescribed drugs and toxic lithocholic bile salts (see Fig. 240-6 ). SXR/PXR also induces expression of ABC transporters to export toxic compounds from cells. The steroid receptor family of proteins thus regulates many aspects of metabolism beyond those regulated by products of classic endocrine organs.

Regulation of Gene Transcription

Hormone-activated receptor proteins bound to their DNA response element targets act as cis-active enhancers. They act from various positions relative to the start of transcription and in various combinations with other regulatory proteins to control the rate of initiation of gene transcription. Gene promoters lie upstream of the site where eukaryotic RNA polymerase II initiates transcription of messenger RNA. The best characterized promoter contains a TATA box that binds a protein, transcription factor II-D (TF II-D), which directs accurate transcription by RNA polymerase II approximately 30 base pairs downstream. Seven proteins (TATA-associated factors [TAFs]) associate with TF II-D in a specific complex that provides a molecular surface for interaction with the transcription regulatory proteins, which are bound elsewhere to DNA. Other promoter motifs include a basal initiator and GC-rich regions in which multiple transcription start sites exist. Gene expression is induced by increasing the rate of transcription. Mechanisms involved in enhancing rates of initiation of transcription include summing of multiple weak protein-protein interactions and acetylating histones to change their interaction with DNA at the transcription start site.

Hormone-activated receptors can also repress transcription. Negative feedback loops operate through this process. Activated cortisol receptors repress transcription of the gene encoding the adrenocorticotropic hormone (ACTH) precursor; activated thyroid hormone receptors inhibit transcription of both α and β-TSH subunit genes. The principle of ligand-activated receptors binding to specific DNA target sequences in the regulated gene is the same as that required for inductive responses. The hormone receptor may inhibit transcription by multiple mechanisms, including deacetylating histones, to increase their interaction with DNA.

Many other proteins regulate initiation of transcription, both as inducers and as inhibitors. These bind to DNA through specific sequences, as do steroid receptors, or they may interact with proteins that do. These proteins may be modified in response to hormonal signals initiated at the cell surface. Such alterations account for the changes in gene transcription resulting from hormones acting through surface receptors. Two general and cooperative mechanisms exist: phosphorylation and translocation of transcription factors from cytoplasm to nucleus. Genes regulated by cyclic AMP contain DNA sequences that specify binding of a specific nuclear transcription regulator (CREB). CREB, which undergoes changes in activity on phosphorylation, is a required final mediator of gene induction by peptide hormones that act at the cell surface to activate adenylate cyclase and cyclic AMP–dependent protein kinase. STAT and related proteins are phosphorylated on tyrosine residues and, when phosphorylated, enter the nucleus to activate transcription of specific genes. This chain of effects alters transcription of messenger RNAs and cell protein concentrations to dictate changes in cell function and organ physiology.

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