Cell Signaling PDF

CELL SIGNALING Cell to cell communication = Cell signalling Cell-to-cell communication is essential for multicellular organisms Intercellular signaling describes the mechanisms by which one cell (the "sender" cell) sends a message to change the function of another cell (the "target" or recipient...

CELL SIGNALING Cell to cell communication = Cell signalling Cell-to-cell communication is essential for multicellular organisms Intercellular signaling describes the mechanisms by which one cell (the "sender" cell) sends a message to change the function of another cell (the "target" or recipient cell). Two cells can communicate (i.e., transmit information to each other) in several different ways. One requires that they be in direct physical contact; this mode is juxtacrine or contact dependent. A second and more common mode is contact independent and involves release of secreted molecules from the sender cells. The secreted molecules diffuse through extracellular space to the target cells where they interact with receptor proteins resulting in an activation of intracellular signaling and altered function. This latter mode permitS intercellular communication between cells separated by large distances, for example, cells in different tissues or organs or between cells separated by short distances Contact-independent signaling by secreted molecules includes four modes: endocrine, paracrine, synaptic or neuronal, and autocrine. These modes are distinguished by: (1) the actual diStance between the sender and target cells; and (2) the biochemical nature of the secreted molecules. Juxtacrine, or Contact-Dependent, Signaling In juxtacrine signaling, cells communicate by either of two mechanisms. One is through gap junction channels that allow direct transfer of small ions, metabolites, and second messenger molecules between neighboring cells. The second mechanism involves the interaction of a protein expressed on the surface of the sender cell with a receptor protein expressed on the surface of the target cell. This interaction changes the conformation of the receptor protein and triggers a cascade of intracellular signaling events in the target cell that results in altered function of that cell. Important elements of juxtacrine signaling include a variety of proteins , such as integrins, involved in cell-cell adhesion and regulation of cell shape and motility. Endocrine Signaling This mode involves signaling between cells separated by short or long distances. Specialized sender cells in glandular tissues synthesize, package, and secrete molecules called hormones. By definition, hormones are molecules secreted into the blood for long-distance transport to the target cells in various tissues. Two important aspecrs of endocrine signaling should be noted. First, because a hormone is being secreted into the total blood volume, it will be diluted many fold by the time it reaches target cells in peripheral tissues. The concentration of hormones in blood and interstitial spaces is always low (picomolar to nanomolar concentrations, i. e., 10 - 12 to 10 -9 M) even during maximal rates of secretion from the source gland. Given the low concentration of hormones, the target cell receptors must have a very high affinity and selectivity for a particular hormone. One consequence of this high affinity is that once the hormone is bound, it cannot easily dissociate from its receptor. Second, it takes a considerable amount of time to increase and or to decrease the concentration of a hormone in blood because of the large volume or capacity of the blood. Following secretion, hormone concentrations in the blood and interstitial fluid can remain elevated for many minutes or even for hours. This and the high affinity of hormone-receptor interactions have important consequences for the mechanisms by which target cells turn off their response to a hormone. It also ensures that intracellular signaling by the target cell will last for long periods. Paracrine Signaling In this mode , a sender cell does not secrete signaling molecules into the blood for long distance transport , but only secretes such molecules into itS immediate or local environment. This greatly restricrs the distances over which the molecules can travel (by passive diffusion) and thus lirnitS responses only to target cells in the immediate vicinity of the sender cell. Given their localized action, the secreted molecules used for paracrine signaling are termed local mediators. Because, unlike secreted hormones, the secreted local mediators are not massively diluted, their concentration near target cells can be fairly high (nanomolar to micro molar range, i. e., 10 -9 to 10 -6 M). Thus, target cell receptors for local mediators usually have lower affinity for these secreted molecules. Lower affinity means that the local mediator can rapidly dissociate from the receptor when the local, extracellular concentration of the mediator is reduced. Moreover, because the mediator is secreted locally into a small extracellular volume, its concentration at the target cell will be only transiently elevated due to rapid diffusion and dilution into the larger tissue space. Thus , paracrine signaling is used for rapid and localized communication between cells. Synaptic or Neuronal Signaling This type of paracrine signaling is specific to nerve cells that send out specialized cellular extensions (axons) to the immediate vicinity of a target cell that is usually another neuron or a muscle cell. Neuron secretes signaling molecules called neurotransmitters. Neurotransmitters travel only very short distances to the target cell. Because of this short intercellular distance and the restricted synaptic volume, the local concentration of neurotransmitter reaching the target cell can be very high (micromolar to millimolar range, i. e., 10 - 6 to 10 -3 M). Thus, receptors on the target cell can have relatively low affinity for the neurotransmitter. This low affinity ensures that neurotransmitters can rapidly dissociate from their receptors following decreases in their local concentration. This is essential for the very rapid (millisecond) termination of neurotransmission from the presynaptic neuron to its postsynaptic targets. Autocrine Signaling This type of signaling occurs when one cell type is both the sender cell and the target cell. Autocrine signaling is often used by organisms during tissue growth , organ development , and immune and inflammatory responses. The general features of autocrine signaling are the same as for non synaptic paracrine signaling except that the sender cell provides both the secreted molecule (often a large polypeptide growth factor) and a receptor for that molecule. For example, early tissue development often involves increased expression of a growth factor and the receptor for that growth by a particular progenitor or stem cell. The released growth factor induces autocrine activation of its target receptor to initiate replication of the original stem cell and its progeny cells. The Three Major Stages of Cell Signaling Reception Transduction Response EXTRACELLULAR CYTOPLASM FLUID Plasma membrane Reception Transduction Response Receptor Activation of cellular response Relay molecules in a signal transduction pathway Signal molecule  Signal transduction and amplification Transduction is the conversion of the external signal message, into a sequence of intracellular metabolic events mediating changes in cell activity. Transduction usually involves multiple steps Multistep pathways can amplify a signal: A few molecules can produce a large cellular response Signal transduction comprises the biochemical decoding of that signal on receipt by the target cells, a process that involves stepwise regulation of intracellular signaling proteins that ultimately results in altered function of proteins involved in metabolism , gene regulation, membrane transport, and cell motility.  Signal Termination 1. The first level of termination is the chemical messenger itself. When the stimulus is no longer applied to the secreting cell, the messenger is no longer secreted, and the existing messenger is catabolized. For example, many polypeptide hormones, such as insulin, are taken up into the liver and degraded. 2. Within each pathway of signal transduction, there are also specific steps at which the signal may be turned off. The receptor might be desensitized to the messenger by phosphorylation. Termination can also be achieved through degradation of the second messenger. Second messengers are molecules that relay signals from receptors on the cell surface to target molecules inside the cell. Each of these terminating processes is also highly regulated.. Sites of signal termination. Processes that terminate signals are shown in blue.  Types of Chemical Messengers There are three types of major signaling systems in the body employing chemical messengers: the nervous system, the endocrine system, and the immune system. It is important to realize that each of the hundreds of chemical messengers has its own specific receptor, which will usually bind no other messenger. There are also some compounds normally considered hormones that are more difficult to categorize. For example, retinoids, which are derivatives of vitamin A (also called retinol) and vitamin D (which is also derived from cholesterol) are usually classified as hormones, although they are not synthesized in endocrine cells.  Table 1 Types of Chemical Messengers System Types of Messengers Examples of Messengers Acetylcholine -amino Nervous (neurotransmitters act as messengers) Biogenic amines butyrate (GABA) Neuropeptides Endorphins Neuropeptide-Y Endocrine (molecule secreted by one organ but Polypeptide Insulin Glucagon acts at another organ) Catecholamines Epinephrine Dopamine Steroid hormones Estrogen Cortisol (lipophilic) Immune (alters gene transcription in target Interleukins Colony- Cytokines cells) stimulating factors Interferons Eicosanoids (control cellular function in Primarily 20 carbon Prostaglandins Leukotrienes response to injury) lipids Growth factors (goes across all systems) Proteins PDGF EGF PDGF, platelet-derived growth factor; EGF, epidermal growth factor  Types of Receptors Most receptors fall into two broad categories: 1. Intracellular receptors 2. Plasma membrane receptors (Fig. ). Messengers using intracellular receptors must be hydrophobic molecules able to diffuse through the plasma membrane into cells. In contrast, polar molecules, such as peptide hormones, cytokines, and catecholamines, cannot rapidly cross the plasma membrane and must bind to a plasma membrane receptor. Plasma membrane receptors have extracellular binding domains. Intracellular receptors bind steroid hormones or other messengers able to diffuse through the plasma membrane. Their receptors may reside in the cytoplasm and translocate to the nucleus, reside in the nucleus bound to DNA, or reside in the nucleus bound to other proteins. 1. Intracellular Receptors Some receptor proteins are intracellular, found in the cytosol or nucleus of target cells Small or hydrophobic chemical messengers can readily cross the membrane and activate receptors Examples of hydrophobic messengers are the steroid and thyroid hormones An activated hormone-receptor complex can act as a transcription factor, turning on specific genes  A transcription factor is a protein that binds to a specific site on DNA and regulates the rate of transcription of a gene (i.e., synthesis of the mRNA). External signaling molecules bind to transcription factors that bind to a specific sequence on DNA and regulate the expression of only certain genes; they are thus called gene-specific or site-specific transcription factors. 2. Plasma Membrane Receptors and Signal Transduction All plasma membrane receptors are proteins with certain features in common: an extracellular domain that binds the chemical messenger, one or more membrane-spanning domains that are α-helices, and an intracellular domain that initiates signal transduction.  As the ligand binds to the extracellular domain of its receptor, it causes a conformational change that is communicated to the intracellular domain through the α- helix of the transmembrane domain. The activated intracellular domain initiates a characteristic signal transduction pathway. Signal transduction pathways run in one direction. From a given point in a signal transduction pathway, events closer to the receptor are termed “upstream” and events closer to the response are termed “downstream.”  In many pathways, the signal is transmitted by a cascade of protein phosphorylations This phosphorylation (kinases) and dephosphorylation (phosphatases) system acts as a molecular switch, turning activities on and off Phosphatase - enzymes remove the phosphates  Signal molecule Receptor Activated relay molecule Inactive protein kinase 1 Active protein kinase 1 Inactive protein kinase ATP 2 ADP Active P protein PP kinase Pi 2 Inactive protein kinase ATP ADP Active P 3 protein PP kinase Pi 3 Inactive protein ATP ADP P Active Cellular PP protein response Pi The pathways of signal transduction for plasma membrane receptors have two major types of effects on the cell: 1. rapid and immediate effects on cellular ion levels or activation/inhibition of enzymes and/or 2. slower changes in the rate of gene expression for a specific set of proteins. Often, a signal transduction pathway will diverge to produce both kinds of effects.  Major Classes of Plasma Membrane Receptors 1. ION CHANNEL RECEPTORS 2. RECEPTORS THAT ARE ENZYMES (KINASES) OR THAT BIND TO AND ACTIVATE ENZYMES (KINASES) 3. HEPTAHELICAL RECEPTORS  1. Ion Channel Receptors or Ligand-gated ion channels (LGICs) (ionotropic receptor) or channel-linked receptor. They are opened or closed in response to the binding of a chemical messenger (i.e., a ligand), such as a neurotransmitter. Most small-molecule neurotransmitters and some neuropeptides use ion channel receptors. NOTE The majority of ion channels fall into two broad categories: voltage-gated ion channels (VGIC) and ligand-gated ion channels (LGIC). Signal Gate molecule closed Ions (ligand) Plasma Ligand-gated membrane ion channel receptor Gate open Cellular response Gate closed  These proteins are typically composed of at least two different domains: a transmembrane domain which includes the ion pore, and an extracellular domain which includes the ligand binding domain.  LGIC is regulated by a ligand and is usually very selective to one or more ions like Na+, K+, Ca2+, or Cl-. Such receptors located at synapses convert the chemical signal of presynaptically released neurotransmitter directly and very quickly into a postsynaptic electrical signal. Many LGICs are additionally modulated by allosteric ligands, by channel blockers, ions, or the membrane potential. Ligand-gated ion channels are likely to be the major site at which anaesthetic agents and ethanol have their effects, halthough unequivocal evidence of this is yet to be established  G-Protein-Coupled Receptors The G-protein-coupled receptors (also known as heptahelical receptors) contain seven-membrane spanning domains which are α-helices. Although there are hundreds of hormones and neurotransmitters that work through heptahelical receptors, the extracellular binding domain of each receptor is specific for just one polypeptide hormone, catecholamine, or neurotransmitter (or its close structural analogue). Fig. Heptahelical receptors  Signal-binding site Segment that interacts with G proteins G-protein-linked receptor Out 7 transmembrane NH2 domain receptor In 2nd messengers COOH G  Heptahelical receptors initiate signal transduction through heterotrimeric G-proteins (proteins which are activated upon binding GTP) composed of α-, β-, and γ-subunits. However, different types of heptahelical receptors bind different G- proteins, and different G-proteins exert different effects on their target proteins  G proteins comprise several families of diverse cellular proteins that subserve an equally diverse array of cellular functions.  These proteins derive their name from the fact that they bind the guanine nucleotides guanosine triphosphate (GTP) and guanosine diphosphate (GDP) and possess intrinsic GTPase activity.  G proteins play a central role in signal transduction as well as in a myriad of cellular processes, including membrane vesicle transport, cytoskeletal assembly, cell growth and protein synthesis.  Mammalian G proteins can be divided into two major categories: heterotrimeric G proteins and small G proteins.  HETEROTRIMERIC G PROTEINS  The family of heterotrimeric G proteins is involved in transmembrane signalling in the nervous system, with certain exceptions.  Heterotrimeric G proteins consist of three distinct subunits, α, β and γ.  These proteins couple the activation of diverse types of plasmalemma receptor to a variety of intracellular processes. In fact, most types of neurotransmitter and peptide hormone receptor, as well as many cytokine and chemokine receptors, fall into a superfamily of structurally related molecules, termed G protein–coupled receptors.  Consequently, numerous effector proteins are influenced by these heterotrimeric G proteins: ion channels; adenylyl cyclase; phosphodiesterase (PDE); phosphoinositide-specific phospholipase C (PI-PLC), which catalyzes the hydrolysis of phosphatidylinositol 4,5-bisphosphate (PIP2); and phospholipase A2 (PLA2), which catalyzes the hydrolysis of membrane phospholipids to yield arachidonic acid.  In addition, these G proteins have been implicated in several other intracellular processes, such as vesicular transport and cytoskeletal assembly. G proteins are now divided into four main categories: 1.the Gs family stimulates adenylyl cyclase; 2.The Gi family (which includes Go, Ggust, Gz) can inhibit adenylyl cyclase, activate a certain type of K+ channel, inhibit voltage gated Ca 2+ channels, activate the MAP-kinase pathway or activate phosphodiesterase; 3.the Gq family activates PI-PLC; and 4.the G12 family (composed of G11-16) activates a group of proteins termed Rho-GEFs (guanine nucleotide exchange factors) The different types of G protein contain distinct α subunits, which confer part of the specificity of functional activity. G protein α subunits, are categorized on the basis of their structural and functional homologies. Current nomenclature identifies several subfamilies of G protein α subunit: Gαs, Gαi, Gαq and Gα12. The Mr of these proteins varies between 38,000 and 52,000. As a first approximation, these distinct types of α subunits share common β and γ subunits. However, multiple subtypes of β and γ subunits are known: five β subunits of Mr 35,000–36,000 and 11 γ subunits of Mr 6,000–9,000. These proteins show distinct cellular distributions, and differences in their functional properties.  The five known forms of G protein β subunit, whose structures are highly similar, are divided into two families comprising Gβ1–4 and Gβ5.  The 11 forms of γ subunit are more divergent structurally. Some show striking regional distributions in the brain: for example, Gγ7 is highly enriched in striatum.  All G protein α subunits are modified in their N-terminal domains by palmitoylation or myristoylation. These modifications may regulate the affinity of the α subunit for its βγ subunits and, thereby, the likelihood of dissociation or reassociation of the heterotrimer. The modifications also may help determine whether the α subunit, released upon ligand-receptor interaction, remains associated with the plasma membrane or diffuses into the cytoplasm.  G protein γ subunits are modified on their C-terminal cysteine residues by isoprenylation Heterotrimeric G Protein α-Subunits in Brain  G protein activation mechanism In the resting state, G proteins exist as heterotrimers that bind GDP and are associated with extracellular receptors (Fig. A). When a ligand binds to and activates the receptor, it produces a conformational change in the receptor, which in turn triggers a dramatic conformational change in the α subunit of the G protein (Fig. B). This conformational change leads to (a) a decrease in the affinity of the α subunit for GDP, which results in the dissociation of GDP from the α subunit and the subsequent binding of GTP because the cellular concentration of GTP is much higher than that of GDP; (b) dissociation of a βγ subunit dimer from the α subunit; and (c) release of the receptor from the G protein (Fig. B, C). This process generates a free α subunit bound to GTP as well as a free βγ subunit dimer, both of which are The system returns to its resting state when the ligand is released from biologically active and regulate the functional activity the receptor and the GTPase activity that resides in the α subunit of effector proteins within the cell. The GTP-bound α hydrolyzes GTP to GDP (Fig. D). The latter action leads to reassociation of subunit is also capable of interacting with the receptor the free α subunit with the βγ subunit complex to restore the original and reducing its affinity for ligand. heterotrimers.  In addition to mediating signal transduction at the plasma membrane, evidence suggests that certain heterotrimeric G proteins are implicated in processes that involve the trafficking of cell membranes. For example, the Gαi subunit has been detected at relatively high concentrations in intracellular membranes, including the Golgi complex, trans-Golgi network and endoplasmic reticulum.  It was assumed that Gαi may regulate the budding of membrane vesicles through these organelles. It also has been suggested that Gαi could be involved in the process by which portions of the plasma membrane are vesicularized into the cytoplasm via endocytosis.  One class of protein kinase that binds βγ subunits is called G -protein–receptor kinases (GRKs). These kinases phosphorylate G-protein–coupled receptors that are occupied by ligand and thereby mediate one form of receptor desensitization.  The βγ subunits also bind to several other proteins, including certain protein kinases as well as phosducin (phosphoprotein, which is located in the outer and inner segments of the rod cells in the retina) and Ras-GEFs; Schematic illustration of the role of G protein β subunits in intracellular targeting of proteins. (A) Under resting conditions, the receptor is associated loosely with a heterotrimeric G protein and G protein- receptor kinases (GRK) are cytoplasmic and therefore unable to phosphorylate the receptor. (B) Upon activation of the receptor and G protein, free α subunit is generated, which can lead to a variety of physiological effects. In addition, a free β subunit dimer is generated, which can bind to the GRK and draw it toward the membrane, where it can phosphorylate the ligand-occupied receptor. In this way, β subunits can direct GRKs specifically to the targets, which are those receptor molecules occupied by ligand. Free β also produces other physiological effects by interacting with other cellular proteins. The β complex is tethered to the membrane by an isoprenyl group on the  subunit, as depicted.  Another important role for βγ subunits is regulation of the mitogen-activated protein kinase (MAP- kinase) pathway, including the MAP-kinase called ERK (extracellular-regulated kinase); MAP-kinases are the major effector pathway for growth factor receptors. However, signals that act through G protein–coupled receptors, particularly those coupled to Gi, can modulate growth factor activation of the MAP-kinase pathway. This is mediated via βγ subunits. Activation of the receptors leads to the generation of free βγ subunits, which then activate the MAP- kinase pathway at some early step in the cascade. Some possibilities include direct action of the βγ subunits on Ras or on one of several ‘linker’ proteins between the growth factor receptor itself and activation of Ras. It was observed Gβγ subunits in mediating G-protein signalling in the absence of activation of the G protein’s associated receptor. This mechanism involves a newly discovered modulatory protein, called GoLoco, which triggers the release of free βγ dimers from G protein–receptor complexes without receptor activation and leads to βγ regulation of its several effector proteins.  The activity of G protein βγ subunits is modulated by phosducin, that is a cytosolic protein enriched in retina and pineal gland but also expressed in brain and other tissues. Phosducin binds to G protein βγ subunits with high affinity. The result is prevention of βγ subunit reassociation with the α subunit. In this way, phosducin may sequester βγ subunits, which initially may prolong the biological activity of the α subunit. However, eventually this sequestration may inhibit G protein activity by preventing the direct biological effects of the βγ subunits as well as preventing regeneration of the functional G protein heterotrimer. The ability of phosducin to bind to βγ subunits is altered upon its phosphorylation by cAMP- or Ca2+-dependent protein kinases  It has been known for years that the activity of small G proteins is modulated by proteins that bind to the G proteins and stimulate their intrinsic GTPase activity. These proteins are termed GTPase-activating proteins (GAPs). Analogous proteins for heterotrimeric G proteins were first identified in yeast but subsequently found in mammalian tissues, are termed RGS proteins. RGS proteins bind to G protein α subunits and stimulate their GTPase activity. This action hastens the hydrolysis of GTP to GDP and more rapidly restores the inactive heterotrimer; Thus, RGS proteins inhibit the biological activity of G proteins. All RGS proteins contain a core RGS domain, which is responsible for regulating G protein α subunit GTPase activity. However, several other domains contained within the RGS protein subtypes may control the protein’s localization, stability and confer other diverse functions on these proteins, in addition to GAP. There is also growing evidence that alterations in the activity of specific RGS proteins, for example via changes in their expression or association with interacting partners, modulate the activity of specific G proteins and, consequently, the sensitivity of specific G-protein–coupled receptors. Such mechanisms have been implicated, for example, in disorders as diverse as hypertension, drug addiction, schizophrenia and Parkinson’s disease  Another example is provided by the R12 subfamily of RGS proteins. These subtypes contain the Go-Loco motif described earlier. The name of the motif comes from the fact that it interacts with G proteins and the fact that the Drosophila homolog of RGS12 is called Loco. Go-Loco binds directly to Gi and stabilizes it in its GDP-bound form. At the same time, it leads to the dissociation of Gβγ subunit dimers, which then activate numerous effectors. This occurs independently of ligand activation of the associated G protein– coupled receptor. In this way, RGS12 proteins can stimulate receptorindependent G protein signalling. Another family of proteins interacting with G protein signalling concerns the activators of G protein signalling (AGS). This group contains structurally diverse proteins that serve as binding partners of G protein components independent of receptor activation G proteins, as well as their associated receptors and RGS proteins, have been reported to undergo phosphorylation by a most of protein serine/threonine kinases and protein tyrosine kinases  SMALL G PROTEINS In addition to the heterotrimeric G proteins, other forms of G protein play important roles in cell function. These proteins belong to a large superfamily often referred to as ‘small G proteins’ because of their low Mr (20,000–35,000). The small G proteins, like the heterotrimeric G proteins, bind guanine nucleotides, possess intrinsic GTPase activity, and cycle through GDP- and GTP-bound. One unifying feature of the various classes of G protein is that the binding of GTP versus GDP dramatically alters the affinity of the protein for some target molecule, apparently by inducing a large conformational change. Small G proteins appear to function as molecular switches that control several cellular processes. Examples of small G proteins and their possible functional roles are given in Table. Examples of Small G Proteins  Schematic illustration of proteins that modulate the functioning of G proteins. The functional activity of G proteins is controlled by cycles of binding GDP versus GTP. This is associated with a major conformational change in the protein. There are several proteins that regulate this cycle and thereby regulate the functional activity of G proteins. Analogous modulator proteins exist for heterotrimeric G protein α subunits and for small G proteins. There are proteins that facilitate the release of GDP from the G protein and thereby enhance G protein function. Examples of such guanine nucleotide exchange factors (GEFs) are receptors for heterotrimeric G proteins or a large number of GEFs specific for various small G proteins. There are proteins, GTPase-activating proteins (GAPs), that activate the GTPase activity intrinsic for the G proteins and thereby inhibit G protein function. Examples are the regulators of G protein-signaling (RGS) proteins for heterotrimeric G proteins and a series of GAPs specific for various small G proteins. There also may be GTPase-inhibitory proteins (GIPs) that exert the opposite effects. Heterotrimeric βγ subunits can be viewed as such; analogous proteins have been proposed for the small G proteins. Phosducin, by binding to βγ subunits, would represent yet another regulatory protein that modulates G protein function. 2. RECEPTORS THAT ARE KINASES OR THAT BIND TO AND ACTIVATE KINASES Signal transduction may involve two types of tyrosine kinase: 1. receptors with inherent tyrosine kinase activity (RTKs), for example insulin receptor; Receptors with tyrosine kinase (RTK) activity 2. receptors, which recruit cytoplasmic tyrosine kinases, for example growth hormone, leptin operating through Janus kinase (JAK). Receptors linked to tyrosine kinases, The tyrosine kinase receptors generally exist in the membrane as monomers with a single membrane- spanning helix. One molecule of the growth factor generally binds two molecules of the receptor and promotes their dimerization (Fig.). Once the receptor dimer has formed, the intracellular tyrosine kinase domains of the receptor phosphorylate each other on certain tyrosine residues (autophosphorylation). The phosphotyrosine residues form specific binding sites for signal transducer proteins.  RAS AND THE MAP KINASE PATHWAY The Ras–MAP kinase pathway provides a nice example to demonstrate how receptor tyrosine kinases can activate signaling through the assembly of protein complexes, due to protein– protein interactions. In this pathway, one of the domains of the receptor containing a phosphotyrosine residue forms a binding site for intracellular proteins with a specific three- dimensional structure known as the SH2 domain (named for the first protein in which it was found, the src protein of the Rous sarcoma virus). The adaptor protein Grb2 is one of the proteins with an SH2 domain that binds to phosphotyrosine residues on growth factor receptors. Binding to the receptor causes a conformational change in Grb2 that activates another binding site called an SH3 domain. These activated SH3 domains bind the protein SOS (SOS is an acronym for “son of sevenless,” a name that is not related to the function or structure of the compound). SOS is a guanine nucleotide exchange factor (GEF) for Ras, a monomeric G protein located in the plasma membrane SOS catalyzes exchange of guanosine triphosphate (GTP) for guanosine diphosphate (GDP) on Ras, causing a conformational change in Ras that promotes binding of the protein Raf. Signal Transduction through Tyrosine Kinase Receptors Raf is a serine protein kinase that is also called MAPKKK (mitogen-activated protein kinase kinase kinase). Raf begins a sequence of successive phosphorylation steps called a phosphorylation cascade. When one of the kinases in a cascade is phosphorylated, it binds and phosphorylates the next enzyme downstream in the cascade. The MAP kinase cascade ultimately leads to an alteration of gene transcription factor activity, thereby either upregulating or downregulating transcription of many genes involved in cell survival and proliferation. Although many different signal transducer proteins have SH2 domains and many receptors have phosphotyrosine residues, a signal transducer protein may only be specific for one type of receptor. This specificity of binding results from the fact that each phosphotyrosine residue has a different amino acid sequence around it that forms the binding domain. Similarly, the SH2 domain of the transducer protein is only part of its binding domain. Conversely, however, several transducer proteins will bind to multiple receptors (such as Grb2). Many tyrosine kinase receptors (as well as heptahelical receptors) also have additional signaling pathways involving intermediates such as phosphatidylinositol phosphates. Thank you for your attention