Final Exam DOC

• Enzyme-coupled receptors


  Cell surface receptors activated by specific ligands that initiate intracellular signaling events through intrinsic enzymatic activity
• Enzyme-coupled receptors


  • Possess both receptor and enzymatic activities in the same protein structure
  • When ligands bind to them they undergo conformational changes to activate enzymatic functions → signaling events
• Receptor tyrosine kinases (RTKs)


  A type of enzyme-coupled receptor that has the ability to phosphorylate tyrosine on target proteins
• RTKs


  • Extracellular ligand-binding domains
  • Transmembrane helix
  • Intracellular tyrosine kinase domain
• Ras-mediated signaling


  1. RTK activation causes the activation of GTPase Ras

  2. Ras GEF

  3. ~30% of human tumors express hyperactive mutant forms of Ras
• Detection of Ras activation

  1. FRET is used to monitor the activation of Ras

  2. Experiment: add "EGF" and detect red light emission under UV
• Names & functions of MAP kinase molecules


  • MAPKKK = Raf =Kinases A
  • MAPKK = MEK
  • MAPK = ERK
  • MAP kinase domain = Phosphorylate other proteins, Osmotic response
  • MAP kinase module = Pass signals to the next kinase, mating response
• MAP kinase signaling


  1. Ras activates MAP kinase pathways which cause changes in gene expression and cell behavior

  2. Stimulates cell division

  3. Ras recruits Raf to the plasma membrane and helps activate it
• MAP kinase Mating Response


  1. Mating factor from yeast binds to a G protein-coupled receptor (GPCR) and activates the G protein

  2. Activated G protein BY complex indirectly activates MAPKKK, referred to as Kinase A

  3. Kinase A sends a signal to MAP kinase module

  4. MAP kinase referred to as Kinase C phosphorylates and activates proteins downstream mediating a mating response for cell arrest and cell fusion

  5. MAPPKKK, MAPKK, MAPK are bound to scaffold protein 1 to ensures signal transduction
• MAP kinase Response to High osmolarity


  1. Yeast cells sense a change in osmolarity-sensing receptor proteins and activate the same MAPKKK [Kinase A] as the mating response

  2. Receptor proteins and kinase are bound to the same scaffold protein

  3. Kinase A relays the signal downstream including the MAP kinase module

  4. Scaffold protein 2 contains MAP kinase domain and provides MAPKK activity

  5. Kinase are bound to scaffold protein 2 and ensure signaling specifically for osmolarity response pathways
• MAP kinase


  • Kinase in each molecule are bound to different scaffold proteins [no cross-talk] between mating response and response to high osmolarity
  • Sequentially phosphorylate and activate each other; amplifying the signal
  • MAPKKK phosphorylates and activates MAPKK which phosphorylates and activates MAPK
  • Scaffold-mediated organization: Scaffold proteins bring together MAP kinases modules so they can efficiently phosphorylate target proteins, increases the chances of kinase-substrate interactions and signal transduction efficiency, facilitate sequential phosphorylation process and amplifies the signal
• Cytokine receptors


  A type of enzyme-coupled receptor that regulates immune responses and hematopoiesis
• Cytokine receptors


  • Immunoregulatory proteins
  • Activate gene regulatory proteins held in laten state by binding to cytokine receptors
  • DO NOT have intrinsic enzymatic activity
  • Ligand binding causes cytokine receptors to activate intracellular kinases (JAKs) leading to the phosphorylation of target proteins and initiation of signaling events
• JAK-STAT signaling pathway activated by cytokines

  1. Cytokine binding to its receptor initiates a cascade of events that ultimately lead to the activation of JAKs, phosphorylation of receptor tyrosine residues, and subsequent phosphorylation and nuclear translocation of STAT proteins

  2. This signaling pathway plays a crucial role in controlling gene expression and regulating various cellular processes, including immune responses, cell growth, and differentiation

  3. Signal off by protein phosphatases
• JAK-STAT signaling pathway


  1. Cytokine binding to receptor

  2. Receptor dimerization or re-orientation

  3. Activation of JAKs

  4. Phosphorylation of receptor

  5. Phosphorylation of STAT proteins

  6. Nuclear translocation of STAT dimers
• Receptor serine/threonine kinases


  • Possess intrinsic kinase activity
  • Phosphorylate serine and threonine on target proteins
  • Crucial to cell growth and development
  • Cell surface receptors that respond to binding to signaling molecules like hormones and local mediators [TGF-B] superfamily
  • Directly initiate signaling pathways within a cell upon ligand binding
• TGF-β Superfamily Signaling


  1. When a TGF-B superfamily ligand binds to a receptor it activates the receptor serine/threonine kinase activity

  2. The activations triggers events that lead to the regulation of gene expression within the cell

  3. An important component of this pathway is a group of cytoplasmic gene regulation proteins called SMADs

  4. SMAD proteins act as mediators of TGF-B signaling by translocation into the nucleus and expression of target genes
• Receptor Serine/Threonine Kinases in Plants

  • Largest class of cell-surface receptors in plants
  • Possess a cytoplasmic domain with typical serine/threonine kinase activity
  • Feature LRR structures known as LRR receptor kinases
  • LRR receptor kinases allow them to recognize and respond to various extracellular signals
• Important Examples of Signaling in Plants

  • Ethylene Signaling
  • Auxin Signaling
  • Phytochrome-Mediated Light Signaling
• Ethylene Signaling


  • Key plant hormone involved in various physiological processes, including fruit ripening, senescence, and response to stress
  • Ethylene signaling pathways involve ethylene receptors and downstream components that regulate gene expression and responses
• Auxin Signaling


  • Key plant hormone that regulates processes such as cell elongation, root development, and tropic responses
  • Auxin signaling relies on receptors, auxin-responsive transcription factors, and auxin-responsive genes to mediate cellular responses to auxin stimuli
• Phytochrome-Mediated Light Signaling


  • Phytochromes are photoreceptors in plants that perceive red light signals and regulate various developmental processes, including seed germination, seedling growth, and photomorphogenesis
  • Phytochrome-mediated light signaling involves the activation of phytochrome receptors, which in turn initiate signaling cascades leading to changes in gene expression and plant growth and development
• Cytoskeleton


  • Support cell shape
  • Cell motility
  • Intracellular movement (transport of organelles)
  • Cell division (mitosis & cytokinesis)
• Types of cytoplasmic fibers

  • Intermediate filaments (Ф ~10 nm)
  • Microtubules (Ф ~25 nm)
  • Microfilaments (Ф ~ 8 nm)
• Dynamic Instability


  • Rapid switching between phases of growth [polymerization] and shrinkage [depolymerization] at the end of microtubules
  • This dynamic behavior allows the microtubules to adapt to changing intracellular conditions quickly and reorganize their cytoskeleton
  • For example, during cell division, dynamic microtubules participate in the formation of the mitotic spindle, facilitating the segregation of chromosomes
  • In intracellular transport, dynamic microtubules serve as tracks for motor proteins to transport cargo such as organelles and vesicles to different regions within the cell
• Fiber-associated (binding) Proteins


  • Molecules that interact with cytoskeletal fibers, regulating their organization, stability, and function
  • Include motor proteins that facilitate intracellular transport along cytoskeletal tracks, as well as cross-linking proteins that stabilize cytoskeletal networks
• Actin Filaments (a.k.a. microfilaments)


  • Found in all cell types, play a central role in muscle contraction
  • Responsible for movement of cells over the substrate to which they are attached
  • Provide "highways" along which intracellular organelles and vesicles can be trafficked
  • Generate the cleavage furrow that divides the cytoplasm during cytokinesis
• Actin monomer


  ~375aa; highly conserved among eukaryotes
• Actin polymerization


  • Actin monomers bound to either ATP or ADP
  • Fiber polarity: plus and minus ends
• Dynamics of actin assembly and disassembly

  1. Nucleation and actin-nucleating factors

  2. Monomer availability and actin monomer binding proteins

  3. Actin filament-binding proteins affect filament dynamics
• Myosins


  • Actin-binding motor proteins in eukaryotic cells
  • Consist of heavy chains and light chains
  • Heavy chains contain motor domain for ATP hydrolysis & actin binding
  • Light chains regulate myosin activity & localization within the cell
• Myosin types in the myosin superfamily

  • Myosin I - involved in membrane and vesicle transport
  • Myosin II - Forms bipolar filaments, helps with muscle contractions, & cytokinesis
  • Myosin V -Intracellular transport, moving vesicles and organelles along actin filaments
  • Myosin VI - Functions in endocytosis and intracellular transport, moving cargo towards the minus end of actin filaments
• Myosin II functions in muscle contraction and cytokinesis


  1. Muscle contraction: Myosin II forms a thick filament that interacts with actin filaments during muscle contraction, ATP hydrolysis by myosin II powers the movements of actin filaments [relative to myosin filaments] causing muscle contraction

  2. Cytokinesis: Myosin II is vital to the contraction of the contractile ring; a structure formed by actin and myosin filaments, the contraction of the ring pinches the cell membrane inward causing the separation of two daughter cells
• Muscle structure


  • Muscles are composed of sarcomeres
  • Skeletal muscle tissue is composed of many, long muscle cells (muscle fibers) arranged in parallel
  • Inside each muscle fiber are numerous myofibrils arranged in parallel along the long axis of the muscle cell
  • Each myofibril consists of many sarcomeres (the fundamental unit of contraction) arranged end-to-end
• Sarcomere components


  • Thin filaments: actin plus actin-binding proteins
  • Thick filaments: bipolar myosin II filaments
• Excitation-contraction coupling


  1. Motor neuron depolarizes the muscle cell

  2. Neurotransmitter (acetylcholine) binds to a receptor on the muscle cell membrane

  3. Neuromuscular junction where motor neurons and muscle fibers communicate
• Muscle cells


  • Contain contractile proteins actin and myosin, which interact to produce muscle contractions
• Sarcomere components


  • Thin filaments: actin plus actin-binding proteins
  • Thick filaments: bipolar myosin II filaments
• Thin filaments


  Actin filaments are thin and serve as attachment sites for myosin during muscle contraction
• Thick filaments


  Myosin interacts with actin filaments, causing the sliding of thin filaments past thick filaments, shortening of the sarcomeres resulting in muscle contraction