Y1U02W1 – Basics of Cell Signalling 24-25.pptx PDF

Unit 2 Week 1 – Basics of Cell Signaling 1st October 2024 Upcoming event… Where: Boiler House, Newcastle University When: 6pm on the 9th of October for a night of presentations, free food and drink! Register here using this link - https://forms.office.com/e/aPVCedVzbuLinks to an e xtern...

Unit 2 Week 1 – Basics of Cell Signaling 1st October 2024 Upcoming event… Where: Boiler House, Newcastle University When: 6pm on the 9th of October for a night of presentations, free food and drink! Register here using this link - https://forms.office.com/e/aPVCedVzbuLinks to an e xternal site. INSPIRE is dedicated to supporting medical students interested in research through workshops, events, and studentships. This symposium kicks off our year of activities, and we’d love to have you with us! We will also use this opportunity to ask you how we can support you throughout the coming year. If you have any questions, please email [email protected] directly. What should you be able to do after this lecture… Define modalities cells use to signal each other Contrast the action of intracellular receptors to cell surface receptors Identify a variety of second-messenger systems and explain how they result in signal amplification Describe the roles of phosphorylation Explain how signalling pathways interact Introduction The big picture Activation/modulation of intracellular signalling cascades amplify and integrate signals from receptors to enable… CONTROLLED RESPONSES to events Cell signalling is dependent on: the incredible specificity of protein-protein interactions the ability of ligand-binding or covalent modification to induce conformational change  modulates activity/target specificity/protein levels – transcription/translation/ stability This underpins both the The parts involved How do cells signal each other? Hydrophilic Hydrophobi c Ligands: the signalling molecule The parts involved How do cells signal each other? Membran e receptor Intracellular receptor Receptor proteins: the place a signal binds The parts involved How do cells signal each other? Signal Cellular transduction respons pathway e Signal transduction pathway Cellular response Signal transduction pathway : converting the signal into a cellular response Several modes of cell communication A signal travels to its target Direct Paracrine Endocrine Synaptic contact signaling signaling Signaling (Autocrine) … defined by the distance a signal travels to its target Several modes of cell communication A signal travels to its target Adjacent plasma membran e Direct contact Gap junctions provides direct contact Plasma membrane Several modes of cell communication A signal travels to its target Secretory cell Paracrine signaling (Autocrine signaling) Adjacent target cells Several modes of cell communication A signal travels to its target Hormone secretion into blood by endocrine gland Endocrine signaling Bloo d vess el Distant target cells Several modes of cell communication A signal travels to its target Nerve Target cell cell Neurotransmit ter Synaptic signaling Synapt ic gap Signal transduction pathways inside the target cell What occurs in the cell after the signal arrives? When the ligand binds Glucag to the protein on receptor, there are many possibilities. Glucag A single response recept on or (e.g. glucagon) ATP cAMP A variety of responses (e.g. Many adrenaline/epinephrine) substrates phosphorylat ed Some are second messenger systems Phosphorylation Two directions Phosphorylation Can activate or deactivate Can act as an on/off switch proteins in signal transduction for protein activity pathways Phosphorylation Activation or deactivation of proteins Energy input Phosphorylated ADP = ATP ADP + Pi AT Dephosphorylated ATP P Energy release = ADP ATP (adenosine triphosphate) – the energy Adenosine triphosphate (ATP) Adenosine triphosphate (ATP) Phosphorylation A on/off switch for protein activity AT ADP P Protein kinase O OH O P O O Protein phosphatase Phosphorylate d protein Protein kinase: An enzyme that takes phosphate from ATP and gives it to a protein (Phosphorylates proteins) Phosphorylation A on/off switch for protein activity AT ADP P Protein kinase OH Pi Protein Protein phosphatase Phosphorylate d protein Pi Protein phosphatase: reverse the process of a kinase (Dephosphorylates proteins ) Receptor types: cell surface receptors Conversion from extracellular signal to intracellular signal Chemically gated ion Enzymatic G-protein coupled channels receptors receptors Three super families of cell surface receptors Receptor types: cell surface receptors Chemically gated ion channels act to transport ions in and out of cells Open when Multi-pass ligand transmembrane is present protein Central channel (hydrophilic passage) Ions Chemically gated ion channels act to transport ions in and out of cells Receptor types: cell surface receptors Enzymatic receptors often use a second messenger Single pass Enzymatic receptors often use a second transmembrane protein messenger Receptor protein kinases Receptor types: cell surface receptors G-protein coupled receptors G-Protein Coupled Receptors Also Use Second Messenger Systems GTP GTP Indirect action on ion channels or enzymes from inside the Activated by GTP (guanosine cell triphosphate) Receptor types: intracellular receptors Passing the cell membrane The ligand must be able to pass through the cell membrane to reach these. Affects a change in gene expression Receptor types: intracellular receptors Passing the cell membrane Some ligands bind to intracellular receptors in the cytoplasm Receptor types: intracellular receptors Passing the cell membrane Others bind to receptors within the nucleus Receptor types: intracellular receptors Steroids act on intracellular receptors Each steroid receptor has three functional domains: Receptor types: intracellular receptors Steroids act on intracellular receptors Hormone- binding domain Hormone receptor complex Hormone response element Receptor types: intracellular receptors Steroids act on intracellular receptors DNA DNA-binding Protein domain mRNA mRN A Receptor types: intracellular receptors Steroids act on intracellular receptors Domain that can interact with cofactors that affect activity Mechanisms of DNA binding and transcriptional activation Homodimeric receptors: (–) Hormone – in the cytoplasm (+) Hormone – receptors translocate to the nucleus and bind to response elements Heterodimeric nuclear receptors (e.g., RXR-VDR, RXR-TR, and RXR-RAR): Located exclusively in the nucleus. (–) Hormone – repress transcription when bound to their cognate sites in DNA by directing histone deacetylation (positively acting on transcription) at nearby nucleosomes (+) Hormone – conformational change – binds histone acetylase complexes – reverses repressing effects Receptor tyrosine kinases Signal molecules Activated tyrosine- Activated proteins kinase receptor (phosphorylated dimer) Regulate normal cell processes Critical role in cancers Neurodegenerative diseases Over 90 RTK genes identified Receptor tyrosine kinase pathways Intracellular or embedded receptor Virtually all aspects of a cell are affected by tyrosine kinases. Protein kinases are enzymes that phosphorylate proteins. Some are Others are embedded in the intracellular membrane as receptors (RTKs) Receptor tyrosine kinase pathways Protein kinase serine, ATP ADP threonine, or tyrosine side Protein chain kinase O– OH O P O– Protei O n Protein Phosphorylated Phosphatase Protein Pi Protein kinases are enzymes that phosphorylate proteins, some are intracellular others are embedded in the membrane Receptor tyrosine kinase pathways Functions Cell cycle Cell growth Cell migration Cell metabolism Cell proliferation Many growth factors Receptor tyrosine kinase pathways Two transmembrane RTKs are involved Extracellular ligand-binding domain Ligan ds Single transmembrane domain Intracellular kinase domain Trans- membrane RTK proteins Receptor Tyrosine Kinase Pathways RTKs are regulated by autophosphorylation Dimerization: a pair of RTKs associate Dimerization and autophosphorylation Receptor tyrosine kinase pathways One signal molecule can elicit multiple responses Phosphotyrosines act as docking sites for other proteins involved in signal transduction Dependent on type of response proteins (often enzymes) Docking Proteins: help other proteins dock on phosphotyrosines to become phosphorylated Adaptor Proteins: facilitate downstream Phosporylated signaling events, do not protein participate in signal transduction Receptor tyrosine kinase pathways The insulin receptor: background We will discuss the role of Adrenaline adrenaline (epinephrine) in the control of blood glucose levels later Receptor tyrosine kinase pathways The insulin receptor: background Receptor tyrosine kinase pathways The insulin receptor: glycogen synthase signaling pathway Insulin receptor Insulin α α Disulfide bridge binds the Insulin extracellular domain Insulin β response β Intracellular domains protein (docking autophosphorylate Activation of protein) glycogen insulin Allows the synthase response protein to Glycogen bind (docking protein) Glucose → synthase glycogen = Passes signal (P) by ↓ blood Glycogen sugar Glucose binding other proteins Receptor tyrosine kinase pathways Kinase cascades Mitogen-activated protein (MAP) kinases are activated by a series of protein kinases MKK Module of protein kinases MKK K K Phosphorylate each other MKK MKK Finally phosphorylates MAP MK kinase MK Amplification of signal is accomplished by kinase cascades Receptor tyrosine kinase pathways Kinase cascades Amplification Each enzyme can act on multiple substrates Large amount of final product = large response Cellular response Cellular response Receptor tyrosine kinase pathways Scaffold proteins Signaling module Scaffold protein Kinase cascade Response proteins Scaffold proteins organize the kinase cascade for ultimate efficiency Multivalent adapter proteins allow modular and adaptable signal transduction SH2 Domains Bind Proteins with Phosphotyrosine Residue G Protein−Coupled Signaling G protein−coupled receptors (GPCRs) are -helical integral membrane proteins G-proteins are heterotrimeric ( ) membrane-associated proteins that bind GTP G-proteins mediate signal transduction from GPCRs to other target proteins The β2-adrenergic receptor (β 2AR) is a GPCR – seven transmembrane domains G-protein coupled receptors The largest category of receptor G-proteins link the Effecto Ligand r receptor proteins to the effector proteins Effector proteins produce second messengers Active second messenger Inactive second messenger GTP: guanosine A c t i v Cascade of a cellular responses t ATP: adenosine e Second messenger systems Overview Many signaling Cytoplas Nonsteroid hormone pathways involve m (first messenger) multiple steps and amplify the signal Enzym e Second messenger Receptor protein Plasma membrane Effect on cellular of target cell function, such as glycogen breakdown G-protein coupled receptors Effector proteins produce second messengers Background: Adrenaline (ligand) Adrenaline is a hormone Adenylyl cyclase made in adrenal glands (pair GPCR of organs on top of kidneys) Mediates stress response: mobilization of energy cAMP Binding to receptors in muscle or liver cells induces breakdown of glycogen Activates PKA Binding to receptors in Respons adipose cells induces lipid e Nucleus hydrolysis proteins Binding to receptors in heart cells increases heart rate GPCR: β2-adrenergic receptor (β2AR) G-protein coupled receptors Effector proteins produce second messengers Things to note: Many Gα subunits may be Adrenaline (ligand) activated by one occupied Adenylyl cyclase receptor GPCR Adenylyl cyclase catalyses the formation of cyclic AMP second messenger cAMP cAMP is degraded – reverses activation of Protein kinase A (PKA) Activates PKA Respons ATP e Nucleus proteins cAMP GPCR: β2-adrenergic receptor (β2AR) Signal Amplification Self-Inactivation in G-protein Signaling Points to note: G proteins cycle between GDP-bound (off) and GTP-bound (on). The protein’s intrinsic GTPase activity, in many cases stimulated by RGS proteins (regulators of G-protein signaling), determines how quickly bound GTP is hydrolyzed to GDP and thus how long the G protein remains active. Desensitization of -Adrenergic Receptors Points to note: Process is mediated by two proteins: β-adrenergic protein kinase (βARK) β-arrestin (βarr; also known as arrestin 2) G-protein coupled receptors GPCRs can use other secondary messenger molecules Ligand For example: cGMP, inositol-1,4,5- GPC Phospholipase C (PLC) triphosphate (IP3) and/or calcium R GTP-bound Gα subunit activates PLC Activated PLC cleaves Phosphatidylinositol 4,5-bisphosphate (PIP 2) to generate Inositol trisphosphate (IP3), which can activate a IP3 ligand-gated ion channel Ca2+- A ligand-gated ion channel releases Ca 2+, binding which acts as another secondary messenger by activating calcium-sensing protein ER proteins like protein kinase C (PKC) or Ca2+ calmodulin PLC cleavage also produces diacylglycerol (DAG) – also activates PKC G-protein coupled receptors Different G-proteins can activate the same transduction pathways… Fight or flight response Let’s say we need to escape a dangerous situation: Adrenaline is present and we need some sugar to get away… Different receptors can produce the same second messengers. One signal molecule could elicit G-protein coupled receptors Different G-proteins can activate the same transduction pathways… Fight or flight response Release of glucose Adrenaline Adenylyl cyclase Adrenaline binds to its Glucagon membrane receptor and a G- GPCR protein activates the second messenger cAMP glucagon also binds to its receptor and has the same PKA effect Phosphorylase kinase Glycogen phosphorylase Glycogen Glucose 6-phosphate G-protein coupled receptors Different G-proteins can activate the same transduction pathways… Fight or flight response So… we have increased our Adrenaline Adenylyl cyclase heart rate but we are not interested in digesting food at GPCR Glucagon this point in time… cAMP PKA Phosphorylase kinase Glycogen phosphorylase Glycogen Glucose 6-phosphate G-protein coupled receptors Or one signal molecule could have different effects… Fight or flight response So… we have increased our heart rate and mobilized glucose production …but we are not interested in digesting food at this point in time Adrenaline Heart Smooth muscle of intestine Adenylyl cyclase increase cAMP G-protein inhibits adenylyl cyclase increase contraction strength relaxation G-protein coupled receptors Different G-proteins can have opposite effects on the same transduction pathways… Breakdown of triacylglycerols to fatty acids and glycerol (lipolysis): Stimulated by binding of adrenaline, glucagon, or adrenocorticotropic hormone (ACTH) to distinct GPCRs, all of which activate adenylyl cyclase via G αs Inhibited by binding of prostaglandin E1 (PGE 1) and adenosine – inhibit adenylyl cyclase via Crosstalk between a tyrosine kinase receptor and a GPCR during insulin signalling is an example of integration of signals Summary: model of cell signaling Six steps of cell-cell communication 1. Synthesis of signal 2. Release of the signaling molecule by the signaling cell: exocytosis, diffusion, cell-cell contact 3. Transport of the signal to the target cell 4. Detection of the signal by a specific receptors 5. Change in cellular metabolism, function or development triggered by the receptor-signal Summary of receptor-mediated signaling Receptors that utilize a nonreceptor tyrosine kinase Receptor tyrosine kinase G-protein coupled receptor (GPCR) seven-transmembrane protein linked to heterotrimeric G proteins Steroid or nuclear receptors that bind ligands and can then influence transcription Notch, which recognizes a ligand on a distinct cell and is cleaved yielding an intracellular Notch (IC Notch) fragment that can enter the nucleus and influence transcription of specific target genes Wnt/Frizzled pathway, where activation releases intracellular β- catenin from a protein complex that normally drives its constitutive degradation. The released β-catenin can then migrate to the nucleus Summary of receptor-mediated signaling Receptors that utilize a nonreceptor tyrosine kinase Receptor tyrosine kinase G-protein coupled receptor (GPCR) seven-transmembrane protein linked to heterotrimeric G proteins Steroid or nuclear receptors that bind ligands and can then influence transcription Notch, which recognizes a ligand on a distinct cell and is cleaved yielding an intracellular Notch (IC Notch) fragment that can enter the nucleus and influence transcription of specific target genes Wnt/Frizzled pathway, where activation releases intracellular β- NICD (Notch intracellular catenin from a protein complex that normally drives its constitutive domain) CSL (CBF1/Suppressor of degradation. The released β-catenin can then migrate to the nucleus Summary of receptor-mediated signaling Receptors that utilize a nonreceptor tyrosine kinase Receptor tyrosine kinase G-protein coupled receptor (GPCR) seven-transmembrane protein linked to heterotrimeric G proteins Steroid or nuclear receptors that bind ligands and can then influence transcription Notch, which recognizes a ligand on a distinct cell and is cleaved yielding an intracellular Notch (IC Notch) fragment that can enter the nucleus and influence transcription of specific target genes Wnt/Frizzled pathway, where activation releases intracellular β- catenin from a protein complex that normally drives its constitutive degradation. The released β-catenin can then migrate to the nucleus Summary: biological roles of signal transduction Signal transduction - Reception of an environmental stimulus by a cell, which leads to metabolic changes that adapt the cell to the stimulus Cells receive signals from the environment beyond the plasma membrane. Types of signals include: antigens hormones neurotransmitters light touch pheromones Signals can originate from diverse sources: Hormones (act at a distance) Growth factors (action is long-lasting) Neurotransmitters (secretion close to target cells) Pheromones (act upon cells in a different organism) These signals cause changes in the cell’s composition and function, such as: differentiation and antibody production growth in size or strength sexual versus asexual cell division Further Reading Berne and Levy: Physiology 7th Ed Chapter 3: Signal Transduction, Membrane Receptors, Second Messengers, and Regulation of Gene Expression https://www.clinicalkey.com/student/content/book/3-s 2.0-B9780323847902000035