Molecular Cell Biology Lecture 20 Notes

Molecular Cell Biology: G Protein-Coupled Receptors I

Lecture Details

  • Instructor: Mitra Esfandiarei, Ph.D.

  • Contact: mesfan@midwestern.edu

  • Date: October 9, 2025

Learning Objectives

  • Components of G Protein-Coupled Receptor (GPCR) Signaling System

    • Understand the components involved in GPCR signaling.

  • Heterotrimeric G Protein Activation

    • Grasp the mechanism of activation of heterotrimeric G proteins.

  • Types of Gα Subunits

    • Identify different Gα subunits associated with GPCRs (e.g., Gsα, Gqα, Giα).

  • Pathway Tracing

    • Analyze GPCR pathways through given figures, identifying ligands, receptors, effector proteins, second messengers, and downstream effects.

  • Examples of Ligands

    • Provide examples of ligands that activate various GPCRs.

  • Signal Transduction Pathways

    • Trace the signal transduction pathways following GPCR activation.

  • GPCR Desensitization Mechanism

    • Explain GPCR desensitization methods, including degradation by endocytosis.

  • Roles of GRKs and Arrestin

    • Understand the roles of G-protein coupled receptor kinases (GRKs) and arrestin in GPCR desensitization.

  • Function of Gα Subunits

    • Differentiate the functions of Gsα, Gqα, Gtα, and Giα subunits.

  • Effectors of GPCRs

    • Discuss different effectors of GPCRs and their downstream pathways.

  • Signaling Pathways of Adrenaline and Glucagon

    • Explain the signaling pathways activated by adrenaline and glucagon, specifically regarding their respective receptors and locations.

  • PKA Regulation by cAMP

    • Describe the relationship between cyclic AMP (cAMP) and protein kinase A (PKA) and its regulation by adenyl cyclase.

  • PKA in Hepatocytes and Skeletal Muscle

    • Understand the role of PKA in hepatocytes vs. skeletal muscle, along with its cytoplasmic vs. nuclear functions.

  • PKA and CREB Relationship

    • Comprehend the relationship between PKA and the cAMP response element-binding protein (CREB).

  • Giα Subunit Role in Cardiac Cells

    • Investigate Giα subunit's role in acetylcholine response in cardiac cells.

  • PKA in Cardiac Pacemaker Cells

    • Explore PKA's role in controlling cardiac pacemaker cells.

  • Acetylcholine and Epinephrine Effects

    • Compare how acetylcholine and epinephrine exert opposite effects on cardiac muscle cells.

  • Gsα vs. Giα Activation Consequences

    • Explain the consequences of activating Gsα versus Giα and their interactions with adenyl cyclase.

Components of the G Protein-Coupled Receptor System

  • Three Main Components

    1. Receptors (GPCRs)

    2. Heterotrimeric G Proteins (comprised of α, β, γ subunits)

    3. Effector Proteins

G Protein-Coupled Receptors (GPCRs)

  • GPCRs are characterized by:

    • Seven transmembrane spanning segments.

    • A large G-protein binding domain on the cytoplasmic side for interaction with Gα subunit.

    • Interaction with a diverse array of extracellular signaling molecules.

    • Activation consequences depend on the specific Gα subunit involved.

    • Over 30% of currently available drugs target GPCRs for therapeutic effects.

Heterotrimeric G Proteins

  • Structure

    • Consists of three subunits: α, β, γ.

    • α and γ subunits are anchored to the plasma membrane via lipid chains.

  • Gα Subunits

    • Humans have 16 genes coding for Gα subunits grouped into five families:

    • Gsα

    • Gq/11α

    • Gtα

    • Giα

    • G12/13α

    • These subunits, while sharing a similar activation mechanism, activate different target proteins (effectors) leading to distinct physiological effects.

Mechanism of Activation of Heterotrimeric G Proteins

  • Gα Activation

    • Contains a conserved G domain that binds to GDP (inactive) or GTP (active).

    • When in the inactive state (bound to GDP), Gα associates with Gβ and Gγ to form a heterotrimer.

    • Upon activation (GTP binding), Gα dissociates from the Gβγ complex, leading to the activation of separate downstream signaling pathways.

  • Upstream Activators

    • GPCRs function as guanine nucleotide exchange factors (GEF) upon ligand binding, enabling activation of heterotrimeric G proteins.

Effector Proteins (Effectors)

  • General Functions

    • These are enzymes mainly located on the plasma membranes of target cells, serving as cellular effectors targeted by activated G proteins.

    • G proteins can modulate effector protein activity, either enhancing or inhibiting their function.

  • Examples of Effector Proteins

    • Adenylyl Cyclase: Converts ATP to cyclic AMP (cAMP) and pyrophosphate.

    • Phospholipase C (β1, β2, β3): Hydrolyzes inositol phospholipids to produce inositol 1,4,5-triphosphate (IP3) and diacylglycerol (DAG).

    • Cyclic GMP-Phosphodiesterase: Breaks phosphodiester bonds in cyclic GMP (cGMP) to form GMP.

Second Messengers

  • Definition

    • Intracellular signaling molecules produced by effector proteins that convey signals from receptors on the cell surface to target molecules within the cell.

  • Examples

    • Cyclic AMP (cAMP)

    • Cyclic GMP (cGMP)

    • Inositol 3-phosphate (IP3)

GPCR Desensitization

  • Process Overview

    • Activated GPCRs can activate intracellular kinases called GRKs (GPCR kinases), which phosphorylate GPCRs on their intracellular domains.

    • Phosphorylation serves as a binding site for another protein, arrestin, which targets GPCRs for endocytosis (internalization into the cell) leading to their degradation, thus shutting down downstream signaling.

Signaling Pathways of Adrenaline and Glucagon

  • Context of Fight-or-Flight Response

    • Adrenaline (Epinephrine)

    • Produced by: Adrenal medulla

    • Trigger: Stress, exercise, or fight-or-flight response

    • Target Cells: Hepatic and skeletal muscle cells

    • Receptors: Beta-2 adrenergic receptors (GPCR)

    • G protein: Gsα

    • Effector protein: Adenylyl cyclase

    • Outcomes: Glycogenolysis (breakdown of glycogen)

    • Glucagon

    • Produced by: Alpha cells in pancreas

    • Trigger: Low blood glucose levels

    • Target Cells: Hepatic cells (liver)

    • Receptors: Glucagon receptors (GPCR)

    • G protein: Gsα

    • Effector protein: Adenylyl cyclase

    • Outcomes: Glycogenolysis

Mechanism of Glycogenolysis Activation

Step I: Activation by Adrenaline

  • Mechanism:

    1. Adrenaline binds to GPCR, activating Gsα.

    2. Gsα activates adenylyl cyclase, increasing cytoplasmic cAMP.

    3. cAMP binds to regulatory subunits of PKA, releasing catalytic subunits and activating PKA.

Step II: Phosphorylation Events

  • Mechanism:

    1. Activated PKA phosphorylates phosphorylase kinase, which in turn activates glycogen phosphorylase.

    2. Glycogen phosphorylase breaks down glycogen to glucose-1-phosphate, contributing to increased blood glucose levels.

    3. Note: In skeletal muscle, glucose is utilized for energy, not released into the bloodstream.

Genomic Effects of PKA Activation

  • CREB Activation

    • Activated PKA can translocate into the nucleus and phosphorylate the transcription factor CREB (cAMP response element-binding protein).

    • Phosphorylated CREB binds to cAMP response elements on target gene promoters, regulating genes involved in glucose metabolism.

Cardiac Muscle Responses

  • Adrenaline Stimulation of Cardiac Contraction

    • Ligand: Adrenaline

    • Trigger: Stress, exercise, or fight-or-flight response

    • Target Cells: Cardiac muscle cells

    • Receptors: β₁-adrenergic receptors

    • G protein: Gsα

    • Effector protein: Adenylyl cyclase

    • Outcomes: Increased cardiac contraction.

Mechanism Overview

  • Adrenaline activates β₁ receptors on cardiac cells, which leads to the activation of Gsα, subsequently activating adenylyl cyclase and increasing cAMP levels.

  • Increased cAMP activates PKA, which leads to enhanced calcium ion entry via L-type calcium channels, ultimately increasing cardiac contraction strength and rate.

  • Acetylcholine Induction of Cardiac Relaxation

    • Ligand: Acetylcholine (ACh)

    • Produced by: Vagus nerve

    • Target Cells: Cardiac pacemaker cells

    • Receptors: M2 muscarinic receptors

    • G protein: Giα

    • Outcomes: Cardiac relaxation.

Mechanism Overview

  • Activation of M2 muscarinic receptors by acetylcholine leads to Giα activation, inhibiting adenylyl cyclase which decreases cAMP levels, thus reducing PKA activity. This results in decreased intracellular calcium concentration, resultant hyperpolarization of pacemaker cells, and ultimately lower heart rates.

Comparative Responses in Cardiac Muscle Cells

  • Differences in Signaling

    • Same Cell Type: Cardiac muscle cells

    • Different Ligands: Epinephrine vs. Acetylcholine

    • Different Receptors: β₁-adrenergic for epinephrine vs. M2 muscarinic for acetylcholine.

    • Different G Subunits: Gsα vs. Giα

    • Same Effector Protein: Adenylyl cyclase

    • Different Outcomes: Contraction (via epinephrine) vs. Relaxation (via acetylcholine).

Summary of Effects

  • Epinephrine

    • Increases heart contraction and rate (positive chronotropy and inotropy)

    • Mechanism: Activation of Gsα → Activation of adenylyl cyclase → Increase in cAMP → Activation of PKA → Enhanced calcium entry.

  • Acetylcholine

    • Decreases heart contraction rate (negative chronotropy)

    • Mechanism: Activation of Giα → Inhibition of adenylyl cyclase → Decrease in cAMP → Reduced PKA activity → Decreased calcium entry.

Conclusion

  • Understanding GPCR Mechanisms

    • Recognition of GPCR signaling importance in regulating physiological responses (e.g., stress, metabolic regulation, cardiac function).

    • Consideration of the broad therapeutic implications as many drugs target GPCRs.