bio230 exam
Cell Polarity
Polarized cells have distinct regions with specialized functions:
Apical domain: Faces outward (e.g., external environment or lumen).
Basolateral domain: Faces inward (e.g., towards other tissues).
Functions:
Signal transmission, internal-external distinction, and region-specific tasks.
Membrane Trafficking and Protein Localization
Trafficking Routes
Direct exocytosis to target domain.
Indirect exocytosis: Proteins are exocytosed, endocytosed, then recycled to specific domains.
Sorting Stations
Sorting occurs in compartments like the Trans-Golgi Network (TGN) or endosomes.
Balanced Retrieval Pathways
Retrieval ensures efficiency and prevents misplacement.
Secretory Pathway
Default Pathway: Constitutive secretion.
No specific signals required; proteins are continuously transported to the plasma membrane.
Regulated Secretion:
Vesicles wait for a signal to release contents.
Example: Mast cells releasing histamine in response to allergens.
Endocytosis
Process: Proteins are internalized from the plasma membrane.
Sorting Routes:
Recycling back to the same plasma membrane domain.
Transcytosis to another domain.
Degradation in the lysosome.
Example: Cholesterol uptake via receptor-mediated endocytosis.
Vesicle Trafficking
Three Main Processes:
Vesicle budding from donor membrane into the cytoplasm.
Vesicle fusion with a target membrane.
Vesicle budding away from the cytoplasm (e.g., into extracellular space).
Key Components:
Clathrin-coated vesicles: Aid vesicle formation.
SNARE proteins: Facilitate vesicle fusion by pairing v-SNAREs (vesicle) with t-SNAREs (target membrane).
ESCRT proteins: Form vesicles away from the cytoplasm.
Membrane Identification and Specificity
Phosphoinositides (PIPs):
Lipids marking different membrane domains.
Can be phosphorylated at specific positions for unique signaling.
Example: PI(4,5)P2 targets clathrin coat assembly for vesicle formation.
Rab GTPases:
Molecular switches for vesicle targeting.
Work in tandem with PIPs for membrane identity.
Rabs & SNAREs:
Coordinate vesicle docking and fusion at the correct target membrane.
Applications of Membrane Trafficking
Both exocytosis and endocytosis establish and maintain cell polarity.
Retrieval pathways and sorting ensure functional distribution of proteins across domains.
Key Takeaways
Mechanisms like endocytosis, exocytosis, and vesicle trafficking are critical for cell polarity and function.
Sorting stations and balanced retrieval maintain cellular efficiency.
Phosphoinositides and Rab GTPases provide specificity for vesicle targeting.
Overview of the Cytoskeletal Network
Purpose of the Cytoskeleton:
Defines cell shape, movement, and intracellular organization.
Coordinates with the cell cycle (interphase, mitosis, cytokinesis).
Dynamic Rearrangements:
Interphase: Microtubules radiate from the center; actin at the cortex.
Mitosis: Microtubules form the spindle; actin disassembles.
Cytokinesis: Actin forms the contractile ring; microtubules separate components.
Microtubules
Structure:
Composed of α- and β-tubulin dimers.
Polarized: α-tubulin (-) at one end and β-tubulin (+) at the other.
13 protofilaments form a hollow tube.
Dynamic Instability:
Growth at GTP-bound (+) end; depolymerization at GDP-bound (-) end.
Rapid disassembly when D-form dimers are exposed.
Nucleation by γ-Tubulin:
Found at minus ends near centrioles in animal cells or along microtubules in plants.
Stabilizes microtubule minus ends and promotes polymerization at the plus end.
Functions:
Vesicle Transport:
Kinesins move vesicles toward the plus end; dyneins move toward the minus end.
Both motors utilize ATP hydrolysis.
Example: Tilapia pigment vesicles move to change coloration.
Actin Filaments
Structure:
Composed of asymmetric monomers that bind and hydrolyze ATP.
Two strands form helical, polarized filaments.
Treadmilling:
Simultaneous addition at the plus end and removal at the minus end.
Regulated by proteins like cofilin and ARP2/3.
ARP2/3 Complex:
Nucleates new filaments from pre-existing ones, forming a branched network.
Protects minus ends from depolymerization and drives filament growth at plus ends.
Functions:
Cell Migration:
Actin pushes the cell forward; myosin contracts the rear.
Integrins anchor actin to the extracellular matrix for traction.
Force Generation:
Myosins use ATP to "walk" along actin filaments.
Critical for processes like cell crawling and muscle contraction.
Cytoskeleton and Cell Polarity
Cytoskeletal asymmetry helps define cellular front-back orientation:
Rho Family GTPases (Rho, Rac, Cdc42):
Act as molecular switches to organize actin filaments.
Rac-GTP drives forward movement; Rho-GTP contracts the rear.
Example: C. elegans fertilization establishes polarity by cytoskeletal reorganization.
Interactions Between Cytoskeletal Components
Actin and microtubules influence each other’s organization.
Symmetry-breaking events, like sperm entry in fertilization, can polarize the cytoskeleton.
Overview: Cell Adhesion
Cell adhesion is essential for the cohesion and organization of multicellular organisms.
Major tissue types:
Epithelial tissue: Cells directly connected with minimal extracellular matrix (ECM).
Connective tissue: Cells dispersed within the ECM.
Epithelial Tissue and Cell Adhesion
Structure and Function:
Lines surfaces, cavities, and organs (e.g., skin, digestive tract).
Polarized cells define inside vs. outside through distinct apical and basolateral domains.
Junctional Complexes:
Link epithelial cells to one another and to the ECM.
Interact with the cytoskeleton for structural support.
Cell-Cell Junctions
Adherens Junctions (Cadherins):
Mediated by cadherins, transmembrane proteins requiring Ca²⁺ for homophilic interactions.
Cadherins sort cells based on expression (e.g., E-cadherin and N-cadherin).
Intracellular domains interact with actin filaments, linking adjacent cells' cytoskeletons.
Role in Morphogenesis:
Adhesion belts contract to form structures like the neural tube during development.
Tight Junctions (Occludins & Claudins):
Define and maintain apical and basolateral domains.
Form homophilic interactions to seal adjacent cells, limiting diffusion of molecules and proteins.
Regulate glucose transport:
Apical domain: Active glucose transport into cells.
Basolateral domain: Passive glucose diffusion into connective tissue.
Cell-ECM Junctions
Mediated by integrins:
Transmembrane proteins that bind ECM components and indirectly interact with actin filaments.
Provide adhesion for both epithelial and connective tissues.
Functions:
Anchor epithelial cells to the basal lamina.
Support cell movement in connective tissue.
Establishing and Maintaining Cell Polarity
Sequence of Junction Formation:
Adherens junctions form first, providing polarity cues.
Tight junctions form apical to adherens junctions.
Polarity Cues:
External signals (e.g., sperm entry) or internal signals (e.g., asymmetric cell division) establish polarity.
Cytoskeleton, intracellular trafficking, and cell cohesion maintain a functional epithelium.
Cell Adhesion in Development and Disease
Critical for processes like morphogenesis (e.g., neural tube formation).
Abnormal adhesion is linked to developmental defects and diseases like cancer.
Key Components of a Functional Epithelium
Intracellular trafficking.
Cytoskeletal organization.
Cell cohesion.
Overview: Tissue Morphogenesis
Morphogenesis is the process that generates shape during development.
Part of multicellular development, which involves:
Cell proliferation (increase in cell numbers).
Cell differentiation (changes in gene expression and cell fate).
Morphogenesis (cell shape, interactions, and location changes).
Processes of Morphogenesis
Cell Internalization
Ingression/Delamination:
Cells detach from epithelial layers and migrate inward.
Example: Formation of mesoderm during epithelial-to-mesenchymal transition (EMT).
Invagination/Involution:
Epithelial sheets fold inward while remaining attached.
Example: Formation of endoderm during gastrulation.
Elongation
Convergent Extension:
Cells crawl together and extend to form elongated structures.
Example: Neural tube elongation.
Mass Cell Migration:
Groups of cells move collectively in chains (less adherent) or sheets (more adherent).
Asymmetric Cell Growth, Division, and ECM Deposition:
Plant cells grow directionally due to the orientation of cellulose fibers in the cell wall.
Microtubules guide cellulose deposition.
Fine Repositioning of Cells
Whole Cell Migration:
Example: Neurons in the cerebral cortex migrate along radial glial cells, with earlier neurons in deeper layers and later neurons in outer layers.
Cell Extension Migration:
Example: Axonal growth cones respond to extracellular signals and chemoattractants to guide extension.
Key Morphogenetic Events in Development
Gastrulation: Formation of three germ layers:
Ectoderm: Epidermis and nervous system.
Mesoderm: Muscles, connective tissue, bones, kidneys, etc.
Endoderm: Gut, lungs, pancreas, liver, etc.
Neural Tube Formation:
Upstream signals lead ectodermal cells to differentiate into neural plate cells.
Neural plate invaginates and closes into the neural tube.
Plant Morphogenesis
Plant cells elongate by directional growth, constrained by cellulose fibers in the cell wall.
Organized microtubules guide cellulose deposition for controlled elongation.
Disorganized microtubules disrupt elongation, causing swelling instead.
Multicellular Development Beyond Embryogenesis
In Adults:
Stem cells contribute to continuous tissue renewal.
New developmental processes (e.g., pregnancy) can occur.
Control Mechanisms:
Careful regulation of cell shape, signaling, and gene expression is essential to maintain organized development.
Overview of Tissue Patterning
Tissue patterning involves the spatial organization of cells into defined structures during development.
Key processes:
Increase in cell numbers via cell division.
Change in cell fates via cell signaling and differential gene expression.
Change in cell shape, interactions, and/or location.
Cell Fate Determination
Differentiation:
Cells acquire specialized functions through differential gene expression.
Ectoderm, mesoderm, and endoderm form specific tissues and organs.
Mechanisms of Cell Fate Acquisition:
Asymmetric Division:
Cell fate determinants are unevenly distributed before division.
Spindle alignment ensures differential inheritance by daughter cells.
Symmetric Division + External Signals:
Daughter cells are initially identical but acquire different fates based on environmental signals.
Mechanisms of Pattern Formation
Lateral Inhibition:
Amplifies small differences between cells, leading to distinct fates.
Example: Notch-Delta signaling in Drosophila, where isolated sensory bristles form in a field of undifferentiated epidermal cells.
Induction by Diffusible Signals:
Organizer tissues secrete morphogens that diffuse and influence cell fate in nearby cells.
Cells respond differently to morphogens depending on concentration, creating patterns like bands or rings of differentiation.
Example: Morphogen gradients in vertebrate body axis formation.
Regulatory Hierarchies:
Sequential signaling events divide tissues into distinct regions.
Example: Anterior-posterior segmentation and further subdivision into head, thorax, abdomen, and polarity within segments.
Hox Genes and Body Patterning
Role of Hox Genes:
Encode transcription factors that determine which body parts develop in specific segments.
Hox gene order on chromosomes corresponds to their spatial expression patterns.
Key Points:
Mutations can lead to body parts forming in the wrong segments (e.g., legs instead of antennae in flies).
Humans have four Hox complexes, and similar mechanisms control body segmentation.
Experimental Approaches to Patterning
Transplant Experiments:
Organizer tissues can induce additional structures when transplanted.
Demonstrates the importance of tissue-specific signaling in development.
Morphogen Stability and Range:
The extent of morphogen diffusion depends on its production, diffusion rate, and stability.
Stages of Cell Potency in Development
Totipotent Cells: Can develop into any cell type, including extra-embryonic tissues.
Pluripotent Cells: Can differentiate into any adult cell type.
Multipotent Cells: Restricted to a specific subset of cell types.
Key Examples
Stomatal Patterning in Plants:
Regulated by lateral inhibition, ensuring evenly spaced stomata surrounded by undifferentiated cells.
Axial Segmentation in Animals:
Driven by overlapping gradients of morphogens and transcription factors like Hox genes.
Overview: Tissue Renewal and Stem Cells
Tissue maintenance involves:
Molecular turnover: Replacement of molecules within cells.
Cellular turnover: Division and death of cells to maintain tissue integrity.
Cellular Turnover
Rates of Turnover:
Rapid turnover: In tissues exposed to harsh environments (e.g., skin, gut lining, blood cells).
No turnover: Specialized cells like auditory hair cells and photoreceptors.
Sources of New Cells:
Stem cells: Divide indefinitely to renew tissues.
Differentiated cells: Rarely divide (e.g., liver hepatocytes, pancreatic β-cells).
Stem Cells
Characteristics:
Divide indefinitely.
Not terminally differentiated.
Capable of self-renewal or differentiation.
Regulation of Division:
Asymmetric division: Uneven distribution of cell fate determinants.
Symmetric division: External signals determine cell fate.
Stem-Cell Niche:
Microenvironment promoting self-renewal via:
Secreted signals.
Cell-cell contact.
Types of Stem Cells
Totipotent: Can form all cell types, including extra-embryonic tissues.
Pluripotent: Can form any adult cell type.
Multipotent: Can form multiple cell types.
Unipotent: Restricted to one cell type.
Terminally Differentiated: No further division.
Applications in Tissue Renewal
Epidermis:
Stem cells divide in the basal layer.
Transit-amplifying cells proliferate and migrate to upper layers.
Dead cells shed from the surface.
Blood:
Hematopoietic stem cells (HSCs) give rise to all blood cell types.
Transit-amplifying cells rapidly increase cell numbers before differentiation.
Stem Cell-Based Therapies
Bone Marrow Transplants:
Used to treat leukemia.
Multipotent HSCs restore blood cell production after irradiation.
Induced Pluripotent Stem Cells (iPS):
Adult cells reprogrammed into pluripotent stem cells using OSKM (Yamanaka) factors.
Avoids ethical issues associated with embryonic stem cells.
Challenges and Innovations
Embryonic Stem Cells:
Full developmental potential but face ethical concerns and risk of immune rejection.
iPS Cells:
Derived from adult cells and retain the same genome as the donor.
Differentiation into desired cell types achieved by exposing iPS cells to specific factors.
Overview of Cell Signaling
Cell signaling enables cells to:
Sense and respond to their environment.
Communicate with other cells.
Signaling occurs in all life forms, from bacteria to humans.
Key Definitions
Extracellular Signaling Molecules:
Molecules like ions, hormones, proteins, or gases that transmit signals between cells.
Receptor:
A protein that binds a signaling molecule (ligand) to initiate signaling.
Ligand:
A molecule that activates a receptor.
Intracellular Signaling Molecules:
Transmit signals inside the cell (e.g., proteins, ions, metabolites).
Secondary Messengers:
Small molecules like cAMP, Ca²⁺, or IP₃ that amplify signals.
Effectors:
Execute cellular responses to signals.
Modes of Signaling
Short Distance:
Contact-dependent: Membrane-bound signals connect neighboring cells.
Paracrine: Local diffusion of signals to nearby cells.
Autocrine: Signals act on the same cell that secreted them.
Long Distance:
Synaptic: Neurons transmit signals across synapses to distant targets.
Endocrine: Hormones travel through the bloodstream to target cells.
Intracellular Signal Transmission
Protein Phosphorylation:
Kinases add phosphate groups; phosphatases remove them.
Phosphorylation changes protein activity, interactions, or localization.
GTP-Binding Proteins:
Active when bound to GTP, inactive with GDP.
Regulated by GEFs (activate) and GAPs (deactivate).
Ubiquitination:
Attaches ubiquitin to proteins, altering their stability, localization, or activity.
Secondary Messengers:
Produced in large quantities to amplify signals.
Examples: cAMP, Ca²⁺, DAG, IP₃.
Signal Regulation
On/Off Mechanisms:
Signals must be reversible to allow dynamic responses.
Examples of inhibitors: phosphatase inhibitors or kinase inhibitors.
Feedback Loops:
Positive Feedback: Amplifies signals (e.g., all-or-none response).
Negative Feedback:
Short delay: Reduces output intensity (desensitization).
Long delay: Creates oscillatory outputs.
Signal Specificity
Multiple Signals:
Cells integrate multiple signals for survival, growth, division, or differentiation.
The same signal can elicit different responses depending on the cell type or receptor.
Signaling Complexes:
Scaffold Proteins: Pre-assembled or signal-induced protein complexes ensure efficient signaling.
Phosphoinositides (PIPs): Lipid modifications recruit specific signaling proteins.
Protein Interaction Domains:
SH3: Binds proline-rich sequences.
SH2/PTB: Bind phosphorylated tyrosines.
PH: Binds phosphoinositides.
Coincidence Detectors:
Proteins activated only by multiple simultaneous inputs (e.g., dual phosphorylation).
Speed of Signaling
Fast Responses: Changes in protein activity or membrane potential.
Slow Responses: Transcriptional or translational changes.
Overview of Small Molecule Signaling
Three Mechanisms:
Independent of plasma membrane proteins.
Through ion channels.
Downstream of G-protein-coupled receptors (GPCRs).
Signaling Independent of Plasma Membrane Proteins
Small, Hydrophobic Molecules:
Passively diffuse across membranes without ion channels or receptors.
Examples: Steroid hormones, nitric oxide (NO).
Intracellular Receptors:
Bind small hydrophobic molecules like steroid hormones.
Structure:
N-terminal: Transcription-activating domain.
Middle: DNA-binding domain (binds gene promoters).
C-terminal: Ligand-binding domain.
Ligand binding activates transcription by freeing DNA-binding domains and recruiting coactivators.
Nitric Oxide (NO):
Made from arginine.
Diffuses rapidly to act locally.
Activates guanylyl cyclase, producing cGMP for smooth muscle relaxation.
Signaling Through Ion Channels
Mechanism:
Ion-channel-coupled receptors are gated by signaling molecules.
Channels mediate passive ion flow down electrochemical gradients.
Synaptic Signaling:
Resting synapse: Synaptic vesicles with neurotransmitters wait near the pre-synaptic membrane.
Nerve impulse triggers neurotransmitter release.
Neurotransmitters bind ligand-gated ion channels on the target cell.
Open channels allow ion influx, triggering cellular responses.
Signaling Downstream of GPCRs
G-Protein-Coupled Receptors (GPCRs):
7-transmembrane domain proteins activated by small molecules, proteins, or light.
Humans have over 800 GPCRs; nearly half of pharmaceuticals target these pathways.
Heterotrimeric G Proteins:
Three subunits: G⍺, Gβ, G𝛾.
GPCR acts as a GEF, exchanging GDP for GTP on G⍺, activating it.
GTP-bound G⍺ dissociates from Gβ𝛾 to activate downstream targets.
GTP hydrolysis by G⍺ terminates signaling.
Examples of GPCR Pathways
cAMP Pathway:
Steps:
GPCR activates G⍺ in Gs.
G⍺ activates adenylyl cyclase.
Adenylyl cyclase converts ATP to cAMP.
cAMP binds protein kinase A (PKA), releasing its active catalytic subunits.
PKA phosphorylates CREB, which activates transcription of cAMP-responsive genes.
Applications:
Olfactory signaling: Odorant GPCRs activate Golf, triggering cAMP production and ion channel opening.
Phospholipase C-β (PLCβ) Pathway:
Steps:
GPCR activates G⍺ in Gq.
G⍺ activates PLCβ.
PLCβ cleaves PI(4,5)P2 into DAG and IP3.
IP3 opens ER Ca²⁺ channels, increasing cytosolic Ca²⁺.
DAG and Ca²⁺ activate protein kinase C (PKC).
Applications:
Signals like Ca²⁺ can regulate numerous downstream processes.
Key Features of Small Molecule Signaling
Amplification:
Small molecules like cAMP and IP3 are produced in large amounts to amplify signals.
Speed:
Ion channel signaling is fast.
Transcriptional changes are slower.
Versatility:
GPCRs integrate diverse signals and regulate complex cellular processes
Overview of Signaling via Protein Modifications
Enzyme-coupled receptors transmit signals through two main mechanisms:
Phosphorylation: Activation of receptor kinases or receptor-associated kinases.
Proteolysis: Signal perception triggers protein cleavage or degradation.
Mechanisms of Phosphorylation Signaling
Receptor Tyrosine Kinases (RTKs):
Structure:
Extracellular domain binds specific ligands.
Transmembrane domain anchors the receptor.
Intracellular kinase domain phosphorylates tyrosine residues.
Activation:
Ligand binding induces dimerization and transautophosphorylation.
Phosphorylation creates binding sites for SH2 or PTB domain-containing proteins.
Pathways:
RTKs recruit Ras-GEFs to activate Ras, a monomeric GTPase.
Ras activates MAP kinase cascades, leading to changes in gene expression.
RTKs can also activate PI3-kinase to produce PI(3,4,5)P3, triggering survival pathways (e.g., Akt activation).
Tyrosine-Kinase-Associated Receptors:
Receptors without intrinsic kinase activity associate with cytoplasmic tyrosine kinases.
Mechanisms resemble RTKs (transautophosphorylation and SH2 domain recruitment).
Receptor Serine/Threonine Kinases:
Similar structure to RTKs but phosphorylate serine/threonine residues.
Signaling via Proteolysis
Notch-Delta Pathway:
Mechanism:
Notch interacts with Delta on adjacent cells.
Interaction triggers Notch cleavage at multiple sites.
The Notch intracellular domain (NICD) translocates to the nucleus, promoting gene transcription.
Applications:
Involved in tissue patterning, neurogenesis, and stem cell renewal.
Hedgehog Pathway:
No Hedgehog Signal:
Patched inhibits Smoothened.
Gli3 is phosphorylated by PKA and undergoes proteolytic processing.
Gli3 fragment represses target gene expression.
With Hedgehog Signal:
Hedgehog binds and inactivates Patched.
Smoothened activates and prevents Gli3 proteolysis.
Gli2 moves to the nucleus to activate target genes.
Applications:
Critical in neural tube closure, limb development, and stem cell renewal.
Specific Examples
Sevenless Signaling in Drosophila:
RTK: Sevenless (Sev).
Ligand: Bride of Sevenless (Boss).
Adaptor Protein: Grb2 (Drk).
Ras-GEF: Son of Sevenless (Sos).
Outcome: Activation of a MAP kinase cascade for photoreceptor R7 cell differentiation.
PI3-Kinase Survival Pathway:
Steps:
RTK activation recruits PI3-kinase.
PI3-kinase converts PI(4,5)P2 to PI(3,4,5)P3.
PI(3,4,5)P3 recruits PDK1 and Akt.
PDK1 and mTORC2 phosphorylate Akt.
Akt phosphorylates Bad, releasing Bcl2 to inhibit apoptosis.
Additional Notes
Scaffold Proteins:
Provide specificity by organizing signaling complexes.
Prevent cross-talk between MAP kinase cascades.
Crosstalk:
Signaling pathways often integrate inputs from RTKs and GPCRs.
Overview of the Cell Cycle
Purpose:
Duplicate parent cell contents and divide them into two child cells.
Stages of the Cell Cycle:
G1 (Gap 1): Growth and preparation for DNA replication.
S (Synthesis): DNA replication.
G2 (Gap 2): Preparation for mitosis.
M (Mitosis): Chromosome segregation and division into two cells.
Checkpoints:
Ensure conditions are favorable for progression (e.g., proper DNA replication and damage repair).
Cyclin-Dependent Kinases (Cdks) and Cyclins
Cdks:
Present throughout the cell cycle but inactive without cyclins.
Phosphorylate target proteins to drive cell cycle progression.
Cyclins:
Levels oscillate during the cell cycle.
Different cyclins activate specific Cdks at specific stages.
Cyclin-Cdk complexes ensure stage-specific activities:
Example: M-Cdk promotes mitosis by phosphorylating spindle assembly and nuclear envelope breakdown proteins.
Regulation of Cyclin-Cdk Activity
Cyclin Synthesis and Degradation:
Cyclin levels are controlled by synthesis and polyubiquitination for degradation.
Example: APC/C and Cdc20 ubiquitinate M-cyclin for proteasome-mediated degradation.
Phosphorylation and Dephosphorylation:
Activation: CAK (Cdk-activating kinase) phosphorylates Cdks to activate them.
Inhibition: Wee1 kinase adds inhibitory phosphate; Cdc25 phosphatase removes it.
Cyclin-Cdk Inhibitors (CKIs):
Bind Cyclin-Cdk complexes to inhibit activity (e.g., p21, p27).
Positive and Negative Feedback Loops
Positive Feedback:
Active M-Cdk activates Cdc25 and inhibits Wee1, ensuring robust activation.
Negative Feedback:
M-Cdk promotes degradation of its own cyclin, terminating its activity.
Molecular Inputs Regulating the Cell Cycle
Mitogens and Growth Factors:
Activate pathways (e.g., RTKs, Ras, MAP kinase cascade) that increase Myc expression.
Myc promotes G1-cyclin production, activating G1-Cdk, which inhibits Rb.
Retinoblastoma (Rb):
Binds E2F to inhibit S-phase gene expression.
Phosphorylation by G1-Cdk releases E2F, allowing S-phase progression.
DNA Damage Response:
Activates p53, which induces p21 CKI expression to inhibit G1/S-Cdk and S-Cdk.
Key Events in the Cell Cycle
G1/S Transition:
Controlled by G1/S-Cdk, promoting entry into S-phase.
S-Phase:
S-Cdk activates DNA helicases to initiate replication.
M-Phase:
M-Cdk triggers spindle assembly and nuclear envelope breakdown.
Cytokinesis:
Cytoplasmic division completes the cell cycle.
Overview of Programmed Cell Death (PCD)
Functions of PCD:
Shapes development (e.g., digit separation in embryos).
Removes damaged, abnormal, or excess cells.
Plays roles in metamorphosis (e.g., tadpole to frog) and immune system function.
Types of Cell Death:
Apoptosis: Controlled, non-inflammatory cell death.
Necrosis: Accidental, uncontrolled death causing inflammation.
Features of Apoptosis
Cellular Changes:
Cell shrinkage, cytoskeleton disassembly, and DNA fragmentation.
Altered cell adhesion and lipid distribution.
Removal via phagocytosis.
Caspases:
Proteases synthesized as inactive procaspases.
Activated by cleavage into active dimers.
Amplified via caspase cascades:
Initiator Caspases: Caspase-8, Caspase-9.
Executioner Caspases: Caspase-3, Caspase-6, Caspase-7.
Caspases cleave specific targets, triggering apoptosis.
Key Caspase Targets and Effects
Cell Shape Changes: Cleavage of adhesion proteins.
DNA Fragmentation:
Breakdown of nuclear lamins.
Activation of DNA endonucleases.
Cytoskeleton Disassembly: Cleavage of actin-regulating proteins.
Lipid Changes:
Flippase inactivation and scramblase activation disrupt lipid asymmetry.
Exposes "eat me" signals for engulfment.
Pathways of Apoptosis
Extrinsic Pathway (Receptor-Mediated):
Fas Receptor and Ligand:
Killer lymphocytes express Fas ligand, binding Fas receptors on target cells.
Death-inducing signaling complex (DISC) activates caspase-8.
Decoy Receptors:
Healthy cells express decoy receptors lacking death domains to block apoptosis.
Intrinsic Pathway (Mitochondrial):
Cytochrome c Release:
Bak and Bax proteins form channels in the mitochondrial membrane (MOMP).
Cytochrome c binds Apaf1, forming the apoptosome to activate caspase-9.
Regulation by Bcl2 Family Proteins:
Anti-apoptotic: Bcl2, BclxL inhibit Bak/Bax.
Pro-apoptotic: Bad inhibits Bcl2/BclxL, promoting MOMP.
Inhibitors of Apoptosis Proteins (IAPs):
XIAP blocks caspases.
Anti-IAPs (released with cytochrome c) neutralize XIAP, allowing apoptosis.
Regulation by Survival Factors
Increase Anti-apoptotic Proteins:
Activate signaling cascades to upregulate Bcl2/BclxL.
Inactivate Pro-apoptotic Proteins:
Akt kinase phosphorylates Bad, inactivating it.
Pathological Implications
Too Much Apoptosis: Can cause degenerative diseases.
Too Little Apoptosis: Enables cancer cells to survive despite damage or detachment from their environment.
Overview of Cancer
Cancer Statistics (Canada 2024):
40-45% of Canadians will receive a cancer diagnosis.
1 in 4 Canadians will die from cancer.
Advances in early detection and treatment are improving survival rates.
Clonal Origin of Cancer:
Cancer originates from a single cell acquiring a mutation that allows uncontrolled growth.
Tumor cells acquire additional mutations over time to enhance survival and proliferation.
Types of Cancer
Carcinomas: Originate from epithelial cells (most common due to high division rates).
Sarcomas: Arise from connective tissue or muscle.
Leukemias: Develop from blood cells.
Tumor Formation and Progression
Benign Tumors:
Localized and non-invasive.
Malignant Tumors:
Invade surrounding tissues and can metastasize.
Metastasis: Tumor cells spread via blood/lymphatic vessels to colonize new tissues.
Key Features of Cancer Cells
Genetic Instability:
Mutations in DNA repair pathways or checkpoints allow rapid mutation accumulation.
Large-scale chromosomal rearrangements and point mutations are common.
Altered Cell Behavior:
Decreased adhesion and increased invasion.
Escape from apoptosis.
Recruitment of other cells to form a supportive tumor microenvironment.
Causes of Cancer
Mutagens:
Chemicals, radiation (e.g., UV, X-rays), or biological agents (e.g., viruses).
Example: Aflatoxin B1 becomes mutagenic after liver metabolism.
Detection of Mutagens:
Ames Test: Detects potential chemical mutagens using bacterial mutation assays.
Viral Infections:
Viruses can integrate into the genome and disrupt normal cell regulation.
Cancer-Critical Genes
Oncogenes:
Mutated proto-oncogenes that promote excessive cell survival, growth, and division.
Features:
Dominant mutations (only one copy needs alteration).
Overactive signaling pathways.
Examples:
Ras: Point mutations inhibit GTPase activity, locking Ras in an active state.
Myc: Overexpression enhances cell cycle progression.
Bcr-Abl (Philadelphia Chromosome): Fusion protein with constitutive tyrosine kinase activity.
Tumor Suppressor Genes:
Normally inhibit cell survival, growth, and division.
Features:
Recessive mutations (both copies need alteration).
Loss promotes unchecked cell growth.
Examples:
Rb (Retinoblastoma): Loss leads to unregulated E2F activity and cell cycle progression.
p53: Responds to stress; loss prevents apoptosis and cell cycle arrest.
Cancer Development Requires Multiple Mutations
Progression of Colorectal Cancer:
Over-proliferation of cells.
Additional mutations in growth pathways.
Activation of oncogenes (e.g., Ras).
Loss of tumor suppressors.
Loss of p53.
Further mutations promote malignancy.
Cancer Prevention and Treatment
Targeting Oncogenes:
Example: Gleevec inhibits Bcr-Abl kinase in leukemia.
General Strategies:
Drugs targeting oncogenic signaling pathways or restoring tumor suppressor function.
Chemotherapy and radiation to target rapidly dividing cells.
PI3-Kinase Overview
Phosphoinositide 3-kinase (PI3-kinase) is crucial for cell signaling, influencing growth, proliferation, and survival.
Mechanism of Action
Catalyzes phosphorylation of phosphatidylinositol to form phosphatidylinositol(3,4,5)-trisphosphate (PIP3), a secondary messenger.
Activates Akt (Protein Kinase B) by recruiting it to the plasma membrane.
Key Functions
Cell Survival: Akt inhibits apoptosis and promotes cell growth.
Metabolism: Regulates glucose uptake and lipid synthesis.
Related Pathways
Activated by receptor tyrosine kinases (RTKs), influencing cross-talk with other signaling pathways like MAPK.
Clinical Significance
Cancer: Abnormal activation is linked to uncontrolled cell growth.
Therapy Target: PI3-kinase and Akt inhibitors are in development for cancer treatment.
Conclusion
PI3-kinase is vital for regulating cellular processes, with implications in cancer when dysregulated.
Diffusible extracellular signals are signaling molecules that can travel freely in the extracellular space to reach target cells. They include various types of molecules such as:
Hormones: Chemical messengers produced by glands, capable of triggering physiological responses in distant organs.
Cytokines: Small proteins important for cell signaling, especially in immune responses.
Growth Factors: Proteins that stimulate cell proliferation and differentiation.
Morphogens: Substances that determine the fate of cells in developing tissues based on their concentration gradients.
These signals can activate receptors on target cells, initiating a response that can influence growth, behavior, and differentiation of cells.
Extrinsic Pathway of Apoptosis:
Ligand Binding➜ Death receptors (e.g., Fas) on cell surface bind ligands.
Caspase Activation➜ Formation of the death-inducing signaling complex (DISC).➜ Activates initiator caspases (e.g., caspase-8).
Execution Phase➜ Initiator caspases activate executioner caspases (e.g., caspase-3).
Cell Death➜ Apoptotic changes: DNA fragmentation, cell shrinkage.
Intrinsic Pathway of Apoptosis:
Cellular Stress/Damage➜ Internal signals (e.g., DNA damage, oxidative stress).
Mitochondrial Outer Membrane Permeabilization (MOMP)➜ Pro-apoptotic proteins (e.g., Bak, Bax) form channels.➜ Release of cytochrome c into the cytosol.
Caspase Activation➜ Cytochrome c binds Apaf-1, forming the apoptosome.➜ Activates initiator caspase-9.
Execution Phase➜ Caspase-9 activates executioner caspases (e.g., caspase-3).
Cell Death➜ Apoptotic changes: DNA fragmentation, cell shrinkage.
Caspases are synthesized as inactive forms called procaspases, which become active during the apoptosis process. The process is as follows:
Gene Expression➜ Caspase genes are transcribed into mRNA.
Translation➜ mRNA is translated into inactive procaspase proteins in the cytoplasm.
Activation➜ During apoptosis, procaspases undergo proteolytic cleavage.
Formation of Active Caspases➜ Cleavage results in the formation of active caspase dimers.
Execution➜ Active caspases cleave specific substrates within the cell, leading to apoptosis.
Caspases trigger apoptosis through the following steps:
Caspase Activation➜ Initiator caspases (e.g., caspase-8 or caspase-9) are activated during the extrinsic or intrinsic pathways of apoptosis.
Activation of Executioner Caspases➜ Activated initiator caspases cleave and activate executioner caspases (e.g., caspase-3, caspase-6, or caspase-7).
Caspase Cascades➜ Executioner caspases propagate the apoptotic signal by cascading activation of more caspases, leading to an amplification of the apoptotic response.
Cleavage of Cellular Targets➜ Active executioner caspases cleave specific substrates:➜ - DNA fragmentation: Cleavage of nuclear lamins and activation of DNA endonucleases.➜ - Cytoskeleton disassembly: Cleavage of actin-regulating proteins.➜ - Cell adhesion changes: Cleavage of adhesion proteins.
Apoptotic Changes➜ Resulting cellular changes include cell shrinkage, membrane blebbing, and the exposure of "eat me" signals that lead to phagocytosis by neighboring cells.
Survival factors initiate signaling cascades that promote cell survival through several key steps. Here's a simplified explanation of the process using arrows to illustrate the flow of signals:
Survival Factor (e.g., Growth Factor) Binding
➜ Receptor Activation (e.g., Receptor Tyrosine Kinase, RTK)
Receptor Dimerization
➜ Autophosphorylation of the receptor
➜ Creation of Binding Sites for downstream signaling proteins (like Grb2 or PI3-kinase)
Recruitment of Adaptor Proteins (e.g., Grb2)
➜ Activation of RAS (through Ras-GEF)
➜ Ras-GTP Activation
MAPK Pathway Activation
➜ Raf Activation
➜ MEK Activation
➜ ERK Activation (MAPK pathway)
➜ Transcriptional Changes for survival genes
PI3-Kinase Pathway Activation
➜ PI3-Kinase phosphorylates PIP2 to PIP3
➜ Recruitment of Akt to membrane
➜ Akt Phosphorylation (by PDK1)
➜ Inhibition of Apoptosis (by inhibiting pro-apoptotic factors like Bad)
Cell Survival Response
➜ Regulates Growth and Proliferation
➜ Prevents Apoptosis
This signaling cascade is crucial for maintaining cellular health, preventing apoptosis, and ensuring proper responses to growth factors.