Cell Signalling and Nuclear Receptors

BS2092 Molecular Cell Biology Notes

Introduction

  • Overview of cell signalling as a critical biological process.

  • Structure of the course includes:
      - Introduction to Cell Signalling
      - Phosphorylation in signalling (Parts 1 and 2)
      - Regulation of signalling output

Learning Objectives

  1. Understand the basic frameworks of cell signalling mechanisms.

  2. Recognize the conceptual role of receptors in signalling events.

  3. Gain insights into the molecular nature of nuclear receptors.

  4. Develop a habit of seeking supporting evidence for established concepts.

Cell Signalling Overview

  • Cell signalling is integral to diverse biological activities such as:
      - Cell Proliferation and Differentiation: Critical for growth and development.
      - Embryonic Development: Influences morphogenesis and organogenesis.
      - Hematopoietic Stem Cell Dynamics: Involves renewal and differentiation of blood cells.
      - Neural Signalling: Facilitates communication among neurons.
      - Example Applications: Tumorigenesis, sensory detection (sound, heat, light), and fertilisation (egg and sperm interactions).

Basic Framework of Cell Signalling

  • Extracellular Signals: These molecules (ligands) interact with receptors on the cell surface or inside the cell.

  • Signal Transduction: The process of converting the extracellular signal into a functional cellular response.
      - Altered Cellular Machinery: Leads to changes in cell behavior.

Components Involved in Signalling

  • Receptor: A protein that binds a specific ligand and initiates a cellular response.

  • Ligand: A molecule that interacts with the receptor.

  • Intracellular Signalling Molecules: Act as second messengers to propagate the signal, such as cyclic AMP, calcium ions, etc.

  • Regulatory Molecules: Modulate the signalling pathway's activity, either enhancing or inhibiting it.

  • Effector Molecules: Execute the actual response in the cell, which may include enzymes or structural proteins.

Mechanisms of Signalling Response

  • Fast Response (Seconds to Minutes): Signals that result in rapid cellular responses, such as enzyme activity changes.

  • Slow Response (Minutes to Hours): Involves gene expression modulation, leading to altered protein synthesis.

Receptors

  • Types of Receptors:
      - Cell-Surface Receptors: These are typically membrane-bound and interact with hydrophilic ligands (e.g., insulin).
      - Intracellular Receptors: Located in the cytoplasm or nucleus; interact with hydrophobic ligands (e.g., steroid hormones).

  • Ligands:
      - Can be classified based on their size and solubility (hydrophilic vs hydrophobic).

Examples of Ligands and Their Actions

  • Estradiol (steroid, hydrophobic) from the ovary stimulates metabolism in various cell types.

  • Thyroid Hormones (hydrophobic) regulate glucose metabolism and protein synthesis in liver and muscle cells.

  • Insulin (hydrophilic): A peptide hormone that stimulates glucose synthesis and lipid breakdown.

  • Glucagon (hydrophilic): Increases blood glucose levels by inducing glycogen breakdown.

  • Cortisol (steroid; adrenal gland): Modulates protein and lipid metabolism.

  • Adrenaline (hydrophilic): Increases heart rate and metabolism via cell surface receptors.

Intracellular Receptor Signalling

  • Nuclear Receptors: Ligand-activated transcription factors that regulate gene expression.

  • Mechanism:
      1. Ligand Binding: Ligand binds to the receptor, leading to a conformational change.
      2. Co-factor Interaction: Active receptors recruit coactivator proteins to enhance transcription.
      3. Transcription Activation: The active receptor-ligand complex binds to the DNA at specific sites to regulate transcription of target genes.

  • Structure of Nuclear Receptors: Consists of a DNA-binding domain, ligand-binding domain, and transcription-activating domain.

Research in Nuclear Receptors

  • History of significant discoveries in glucocorticoid receptors and their medical applications, particularly for anti-inflammatory treatments like asthma.

  • Advances in genetic and molecular techniques for identifying nuclear receptors and understanding their functions, including:
      - Obtaining DNA fragments encoding receptors for functional studies.
      - Biochemical assays to assess DNA binding and protein interactions.
      - Structural analysis through techniques like X-ray crystallography.

Generating Recombinant Proteins in Bacteria

  • Steps:
      1. Prepare Plasmids: Incorporate the cDNA of interest along with antibiotic resistance genes into bacterial plasmids.
      2. Transformation: Introduce plasmids into E. coli for protein expression.
      3. Induction of Expression: Use IPTG to induce protein expression in bacteria.
      4. Protein Purification: Harvest and lyse bacterial cells to extract and purify the recombinant protein.

  • Applications: This method allows for the study of protein function, localization, and interactions in living cells.

Conclusion

  • Understanding the intricate processes of cell signalling and the role of receptors is critical for advancing molecular biology, genetics, and biomedical applications.