Signal Transduction

• Signal transduction is the conversion of information into a chemical change within a cell.
• Signals detected by specific receptors initiate the transduction process.
• The conversion of a signal to a cellular response involves a chemical process.

Specificity in Signal Transduction

• Precise molecular complementarity between the signal and receptor molecules ensures specificity.
• Weak, noncovalent forces mediate this specific interaction.

Common Features of Signal-Transducing Systems

• Sensitivity arises from the high affinity of signal receptors for their ligands.
• The dissociation constant (Kď) is commonly < 10-7 M, indicating high affinity.
• Amplification occurs when a signal receptor activates an enzyme, which then catalyzes the activation of multiple molecules of another enzyme in an enzyme cascade.
• Modular proteins have multiple domains that recognize specific features allowing cells to mix and match signaling molecules for diverse responses.
• Scaffold proteins are nonenzymatic proteins that bring together interacting enzymes in cascades, facilitating efficient signal transduction.
• Desensitization happens when a receptor is no longer responsive to a signal that persists continuously which prevents overstimulation.
• Integration is the capacity of a system to receive multiple signals and produce a unified response allowing cells to make comprehensive decisions based on various inputs.
• Divergence occurs where signaling pathways branch, resulting in multiple downstream effects from a single signal, which allows for complex cellular responses.
• Response localization refers to the confinement of signaling system components to specific locations, regulating processes locally and preventing widespread effects.
• Cells can respond to a myriad of different biological signals such as antigens, cell surface glycoproteins, growth factors, and hormones.

Basic Protein Components of Signal Transduction

• Plasma membrane receptors with 7 transmembrane (7tm) helices commonly initiate signal transduction.
• G proteins, binding GTP or GDP, interface with membrane receptors to mediate downstream signaling.
• Membrane enzymes, with cyclic nucleotides as substrates or products, play roles in signal amplification and diversification.
• Protein kinases phosphorylate GPCR receptors, modulating their activity and downstream signaling.
• Membrane protein tyrosine kinases add phosphate groups to tyrosine residues on target proteins to regulate protein activity.
• Cyclic nucleotide-dependent protein kinases are activated by cyclic nucleotides, mediating various cellular responses.
• Ca2+-binding proteins and Ca2+-dependent protein kinases are involved in calcium-mediated signaling pathways.
• Protein kinases activated during cell division regulate cell cycle progression.
• Nonenzymic protein scaffolds organize signaling components into functional modules, improving efficiency and specificity.

General Steps of Signal Transduction in Animals

• A signal (ligand) interacts with a receptor.
• The activated receptor interacts with cellular machinery to produce a second signal or change in protein activity.
• Cellular metabolic activity changes.
• The transduction event ends.

Four General Types of Signal Transducers

• G protein-coupled receptors activate intracellular GTP-binding protein to regulate an enzyme producing a second messenger.
• Receptor enzymes (tyrosine kinase) activate tyrosine kinase activity via autophosphorylation upon ligand binding.
• Gated ion channels open or close in response to signal ligand concentration or membrane potential.
• Nuclear receptors enable hormones to regulate specific gene expression.

GPCRs

• GPCRs act through guanosine nucleotide-binding proteins (G proteins).
• Three essential components of GPCRs include: a plasma membrane receptor with seven transmembrane helical segments, a G protein cycling between active (GTP-bound) and inactive (GDP-bound) forms, and an effector regulated by the activated G protein.
• A "first messenger" is an extracellular signal that activates a receptor on the cell surface.
• A "second messenger" is a low molecular weight metabolite or inorganic ion whose concentration changes due to the effector enzyme, which then activates or inhibits downstream targets.
• Adrenergic receptors are protein receptors in the plasma membrane that bind epinephrine, categorized into four general types: α1, α2, β1, β2.
• β-adrenergic receptors specifically refer to the β1 and β2 subtypes.
• An agonist is a molecule that binds a receptor and produces the effects of the natural ligand.
• An antagonist is an analog that binds the receptor, blocking the effects of agonists and the natural ligand.
• GPCRs (seven-transmembrane (7tm) or heptahelical receptors) span the membrane seven times and interact with heterotrimeric G proteins.
• Heterotrimeric G proteins has three subunits; α, β, and γ and the α subunit is the binding site for GDP or GTP.
• A hormone-bound GPCR acts as a guanosine nucleotide-exchange factor (GEF)
• Because the G protein stimulates its effector, it is referred to as a stimulatory G protein (Gs)
• Adenylyl cyclase is an intergral protein in the plasma membrane that catalyzed the synthesis of cAMP from ATP when associated with active Gsα.
• Adenylyl cyclase can also be inhibited by inhibitory G protein (Gi).
• Inactive G protein (Gi) can be transformed if activated by the binding of somatostatin to its receptor and it structurally homologous to Gs.
• Gα possesses intrinsic GTPase activity, converting Gα to its inactive state by hydrolyzing bound GTP to GDP.
• cAMP activates cAMP-dependent protein kinase (protein kinase A or PKA), which catalyzes the phosphorylation of specific Ser or Thr residues on target proteins.
• The R2C2 complex (PKA) is catalytically inactive until cAMP binds.
• Cyclic AMP is an allosteric activator of PKA and bining of cAMP creates two active C subunits
• Consensus sequences commonly contain the target for phosphorylation (Ser or Thr) by protein kinases.
• Fluorescence Resonance Energy Transfer (FRET) measures the nonradiative transfer of energy between fluorescent probes and can be used to detemine if two proteins interact.
• When [cAMP] is low, R and C subunits of PKA are associated and FRET is exhibited.
• When [cAMP] rises, R and C subunits of PKA dissociate and FRET ceases.
• When PKA is inactive the Ser residue is not phosphorylated, 14-3-3 has no affinity for the Ser residue, and FRET is not observed.
• When PKA is active in the cell, the Ser residue is phosphorylated, 14-3-3 binds the Ser residue, and FRET is observed
• GPCRs are also often involved in vision, olfaction, and gustation
• Olfactory receptors are GPCR and bind a wide variety of odorants
• Rhodopsin is a GPCR in the disk membranes of rod cells of the vertebrate eye
• Amplification is the function of the enzyme cascade
• The cascade amplifies the hormonal signal by orders of magnitude and accounts for the low concentration of epinephrine required for activity.
• The signal leads to intracellular changes within fractions of a second.
• Termination methods: epinephrine concentration drops below the Ka for its receptor, the GTPase activity of the G protein hydrolyzes the GTP bound to the Go subunit, or cAMP is hydrolyzed to 5' AMP by cyclic nucleotide phosphodiesterase.
• GTPase activator proteins (GAPs) strongly stimulate GTPase activity.
• Phosphoprotein phosphatase hydrolyzes phosphorylated Tyr, Ser, or Thr residues, releasing inorganic phosphate (Pi) which reverses the metabolic effects that result from PKA activity.
• Desensitization decreases the response while the signal persists.
• β-adrenergic receptor kinase (βARK) phosphorylates several Ser residues near the receptor’s carboxyl terminus when epinephrine is bound to the receptor.
• β-arrestin (βarr or arrestin 2) binds to the phosphorylated carboxyl-terminal domain of rhodopsin to prevent further interaction between activated rhodopsin and transducin.
• G protein-coupled receptor kinases (GRKs) are kinases that phosphorylates GPCRs and play a role similar to that of BARK in receptor desensitization and resensitization
• Activation in the guanosine nucleotide-binding protein occurs when GTP bind to the α subunit
• Cyclic AMP (cAMP) acts as a second messenger for many regulatory molecules
• cAMP acts by changing intracellular [cAMP] and thus the activity of PKA.
• In some cells, the catalytic subunit of PKA can also move into the nucleus, where it phosphorylates cAMP response element binding protein (CREB)
• cAMP response element binding protein (CREB) = alters the expression of specific genes regulated by cAMP
• extracellular signals have different effects on different tissues or cell types, depending on the type of receptor in the tissue, the type of G protein (Gs or Gi ) coupled to the receptor, and the set of PKA target enzymes in the cell.
• Adaptor proteins are noncatalytic proteins that hold together other protein molecules that function in concert.
• Heterotrimeric G proteins play roles in cellular processes, including, sensory perception, signaling for cell division, growth and differentiation, intracellular movements of proteins and membrane vesicles, or protein synthesis.
• AKAPs (A kinase anchoring proteins) = have multiple, distinct protein-binding domains

cGMP Signaling

• Guanylyl cyclase converts GTP to cGMP when it is activated.
• cGMP (guanosine 3',5'-cyclic monophosphate) has role as second messenger.
• cGMP-dependent protein kinase = protein kinase G (PKG) = mediates many of the actions of cGMP
• Atrial natriuretic factor (ANF) = peptide hormone that activates guanylyl cyclase in the kidney and is released when blood volume increases.
• A rise in [cGMP] triggers increased renal excretion of Na+ and consequently of water, which reduces blood volume.
• G proteins act as self-limiting switches in many processes, including sensory perception and cell division.
• Ras is a ~20 kDa minimal signaling unit
• GTPase activator proteins (GAPs) = increase the GTPase activity of G proteins
• Guanosine nucleotide-exchange factors (GEFs) = catalyze the exchange of bound GDP with GTP and example is the β-adrenergic receptor
• A mutation in Ras eliminates its GTPase activity, causing the protein to remain constantly active and occurs in ~25% of human cancers.
• "Activating” mutations = lead to a continuously elevated [cAMP] found in ~40% of adenomas.
• "Inactivating” mutations = cause individuals to be unresponsive to hormones that act through cAMP.
• The enzyme, cholera toxin, catalyzes the transfer of the ADP-ribose moiety of NAD+ to an Arg residue of Gsa and is secreted by Vibrio cholerae
• Phospholipase C (PLC) = catalyzes cleavage of the membrane phospholipid phosphatidylinositol 4,5-bisphosphate, or PIP2
• Gq = associated trimeric G protein that activates the PIP2-specific PLC when an agonist binds its specific receptor
• diacylglycerol and inositol 1,4,5- trisphosphate (IP3) = potent second messengers
• IP3-gated Ca2+ channel = receptor-gated channel in the endoplasmic reticulum that opens to release sequestered Ca2+ into the cytosol.
• Elevated [Ca2+] and diacylglycerol activate protein kinase C (PKC)
• protein kinase C (PKC) = binds and phosphorylates proteins that contain a PKC consensus sequence
• In unstimulated cells, cytosolic [Ca2+] is kept very low by Ca2+ pumps.
• Hormonal, neural, or other stimuli cause either an influx of Ca2+ into the cell through Ca2+ channel or the release of sequestered Ca2+ into the cytosol.
• Oscillates in concentration with both time and space
• Calmodulin (CaM) = acidic protein that undergoes a conformational changes when Ca2+ binds and associates with a variety of proteins to modulates their activities.
• Ca2+/calmodulin-dependent protein kinases (CaM kinases) phosphorylates target enzymes following activation by Ca2+-bound calmodulin
• Gustducin is a heterotrimeric G protein coupled to GPCRs in taste sensory neurons and stimulates cAMP production by adenylyl cyclase
• a tumor suppressor gene NF1 can cause Ras to stay active for an extended period The closing of K+ channels in the plasma membrane following phosphorylation by PKA sends an electrical signal to the brain

GPCRs in Special Senses

• In vision, light is a trigger for a GPCR
• In vertebrate olfaction, neuronal action potentials can occur due to small depolarizations produced by increasing Na+ and Ca2+
• Rhodopsin acts via retinal to receive sensory information involving light
  • Rhodopsin kinase = phosphorylates Thr and Ser residues in the carboxyl-terminal domain of rhodopsin and functionally and structurally homologous to the βARK that desensitizes the β-adrenergic receptor

GPCR Features

• GPCRs are encoded in vertebrates, Drosophila, Caenorhabditis elegans, Saccharomyces
• Approxmiately 800 GPCRs are encoded from the ~20,000 genes in the human genome
• GPCRs can detect other molecules inlcuding amines, peptides, and protein hormones
• Heterotrimeric G proteins play roles in cellular processes, including, sensory perception, signaling for cell division, growth and differentiation, intracellular movements of proteins and membrane vesicles, or protein synthesis

Receptor Tyrosine Kinases (RTKs)

• Receptor tyrosine kinases (RTKs) = family of plasma membrane receptors with protein kinase activity and have a cytoplasmic Tyr kinase domain
• Active insulin receptor protein (INSR) = a dimer of αβ monomers, where α subunits contain the insulin-binding domain, and intracellular domains of the β subunits contain the protein kinase activity

Insulin Signal Transduction

• Each β subunit phosphorylates three essential Tyr residues

Raf-1, MEK, and ERK

• ERK resides in the MAPK family (mitogen-activated protein kinases) and is specific for Ser or Thr residues
• MEK is of MAP kinase kinase (MAPKK) family that activates the MAP kinase and phosphorylates both Ser and Try residues
• Raf-1 resides in the MAP kinase kinase kinase (MAPKKK) family , activates the MAP kinase kinase, and is specific for Ser or Thr residues
• PIP3 (Phosphatidylinositol) mediates a lipid signal, with kinase function
• PIP2 = Phosphatidylinositol 4,5-bisphosphate
• PIP3 = Phosphatidylinositol 3,4,5-trisphosphate