Endoplasmic Reticulum and Disease L3 PDF
Document Details
Uploaded by WinningHoneysuckle
UCLan
2024
Dr Temba Mudariki
Tags
Related
- The Endoplasmic Reticulum PDF
- Roles of Cytosolic Hsp70 and Hsp40 Molecular Chaperones in Post-translational Translocation of Presecretory Proteins PDF
- Cell and Molecular Biology Lecture 13 PDF
- AQ-112 - Fundamentals of Biochemistry PDF
- Introduction to Cell and Molecular Biology PDF
- Molecular Biology of the Cell Chapter 12 - Intracellular Compartments and Protein Sorting PDF
Summary
This document is a presentation on the endoplasmic reticulum (ER). It discusses the ER's structure, function, role in protein synthesis, folding, and trafficking, and its connection to various diseases. The presentation also covers the process of protein import into the ER, modifications, quality control mechanisms, and examples of diseases related to ER dysfunction. The document also looks at specific therapeutic strategies.
Full Transcript
Presentation by: Dr Temba Mudariki Endoplasmic Reticulum Molecular Medicine XY3121 2024 Introduction Learning Outcomes Know the differences between the rough and smooth ER, their location and the appearance of these organelles on electron microscopy Desc...
Presentation by: Dr Temba Mudariki Endoplasmic Reticulum Molecular Medicine XY3121 2024 Introduction Learning Outcomes Know the differences between the rough and smooth ER, their location and the appearance of these organelles on electron microscopy Describe the functions of the smooth ER Describe the functions of the rough ER Understand the differences between co-translational translocation and post-translational translocation, and which types of proteins undergo which type of translocation Understand the processes of protein translocation to the lysosomes, endosomes and plasma membrane Understand the processes of protein secretion Explain protein import into the ER Explain what happens to proteins in the ER Explain what happens to misfolded proteins in the ER Explain the cellular pathogenesis and consequence of defective protein folding. Identify and understand medical conditions associated with defective transmembrane protein folding The Endoplasmic Reticulum: The Cellular Highway The endoplasmic reticulum (ER) serves as the cell's biosynthetic powerhouse and a highway for the transport of molecular materials. We will discuss: The ER's structure and function. The differentiation between rough and smooth ER. The critical role of the ER in protein and lipid synthesis. The consequences of ER dysfunction on cellular health. Understanding the ER The ER is a network of membranous tubules and flattened sacs that extend throughout the cytoplasm. It is contiguous with the outer nuclear membrane. The ER is categorized as rough (RER) or smooth (SER) based on its appearance under a microscope. Functions of the ER include: Protein synthesis (RER) Lipid metabolism (SER) Calcium storage (SER) Drug detoxification (SER) The Rough ER: A Ribosome-Studded Landscape The RER is characterized by the presence of ribosomes on its cytoplasmic surface. It is the site of synthesis for secretory, membrane-bound, and organelle-targeted proteins. Newly synthesized proteins enter the RER lumen where they undergo folding and post-translational modifications. The Smooth ER: Diverse Functionalities The SER lacks ribosomes and has a more tubular appearance. It is involved in lipid and steroid hormone synthesis, crucial for cell membrane formation and signalling. The SER also plays a role in detoxifying metabolic by-products and xenobiotics. It regulates intracellular Ca²⁺ levels, important for muscle contractions and other signalling pathways. Endoplasmic Reticulum (ER) Protein Manufacturing in the Rough ER Ribosomes translate mRNA into polypeptide chains that are co- translationally translocated into the RER. Inside the RER, proteins are folded with the help of chaperones. They undergo modifications such as glycosylation and disulphide bond formation. Properly folded proteins are then packaged into vesicles for transport to the Golgi apparatus. Membrane Protein Insertion Type I Membrane Protein Insertion into the ER Defining Characteristics: Single transmembrane domain N-terminus in the ER lumen, C-terminus in the cytosol Contains a cleavable N-terminal signal sequence Insertion Process: Begins with ribosome translation of the signal sequence Signal Recognition Particle (SRP) pauses translation and directs the ribosome to the ER Signal peptide is inserted into the translocon, and cleaved off as translation resumes The hydrophobic stop-transfer sequence halts translocation, anchoring the protein in the membrane The C-terminal continues to be synthesized into the cytosol Example Proteins: Glycophorin & CD4 Type II Membrane Protein Insertion into the ER Defining Characteristics: Single transmembrane domain N-terminus in the cytosol, C-terminus in the ER lumen Contains a signal-anchor sequence not cleaved Insertion Process: Begins with ribosome translation of internal signal-anchor sequence SRP directs the ribosome to the ER; signal-anchor sequence initiates insertion The orientation is dictated by positive charges flanking the signal-anchor sequence The polypeptide chain grows into the ER lumen, forming the C-terminus Example Proteins: Asialoglycoprotein receptor G protein-coupled receptors (some types) Type III Membrane Protein Insertion into the ER Defining Characteristics: Single transmembrane domain N-terminus in the ER lumen, C-terminus in the cytosol Signal-anchor sequence remains as a transmembrane segment Insertion Process: Translation begins with an internal signal-anchor sequence SRP binds to the signal-anchor and guides the complex to the ER The orientation is generally N-lumenal, often influenced by the distribution of positive charges Protein continues to elongate, inserting the C-terminal into the cytosol Example Proteins: Cytochrome P450 enzymes Flippase proteins Type IV Membrane Proteins: Multipass Insertion Defining Characteristics: Multiple transmembrane domains Complex orientation with both termini on the same or opposite sides of the ER membrane Contains several signal-anchor and stop-transfer sequences Insertion Process: Translation begins with a signal-anchor sequence that is not cleaved Multiple signal-anchor and stop-transfer sequences guide the ribosome in inserting the protein The protein loops in and out of the translocon, inserting multiple domains into the membrane Example Proteins: G protein-coupled receptors with multiple transmembrane domains Ion channels like the potassium channel Protein Trafficking Pathways Translocation Overview: Proteins synthesized in the ER are destined for various locations: lysosomes, endosomes, or the plasma membrane. Transport to these locations is highly regulated and critical for proper cellular function. Vesicle Transport: Vesicles are the primary mode of transport for proteins from the ER to their destinations. Bud from the ER or Golgi apparatus, vesicles ferry encapsulated proteins through the cytoplasm. Protein Trafficking Pathways Sorting Signals: Proteins contain specific amino acid sequences that act as postal codes, directing them to the correct cellular address. These signals are recognized by adapter proteins which mediate the sorting of proteins into vesicles. Interactive Activity: Let's map out the journey of a protein from its synthesis to its specific cellular location. Audience participation to identify the sorting signals and predict the destination of example proteins. Lysosomes receive Proteins and Cargo from Multiple Pathways The Secretory Pathways Secretory Pathways Overview: Post-translational pathway: Proteins are synthesized in the ER, processed in the Golgi, and then transported out of the cell. Secretory proteins are packed into vesicles that bud from the Golgi and migrate towards the plasma membrane. Constitutive Secretion: Continuous, non-selective process where secretory vesicles fuse with the plasma membrane to release their contents. Operates constantly in all cells, delivering proteins like extracellular matrix components. The Secretory Pathways Regulated Secretion: Selective, triggered process in response to specific signals or environmental cues. Common in cells that produce hormones, neurotransmitters, and digestive enzymes. Real Cell Examples: Constitutive: Collagen secretion by fibroblasts for extracellular matrix formation. Regulated: Insulin release from pancreatic β-cells in response to glucose levels. Protein Import into the ER Import Mechanism: Proteins destined for the ER have a signal sequence that directs them to the ER membrane. The signal recognition particle (SRP) binds to this sequence and pauses translation. The SRP-ribosome complex docks on the ER membrane, and the protein is threaded into the ER lumen through a translocon channel. Protein Folding: Once inside the ER, proteins must fold into their three-dimensional shapes to become functional. The ER provides an optimized environment for protein folding, with a unique set of enzymes and conditions. Protein Import into the ER Chaperones and Foldases: Molecular chaperones, such as BiP, assist in proper protein folding and prevent aggregation. Foldases, like protein disulphide isomerase (PDI), facilitate the formation of disulfide bonds between cysteines. Post-translational Modifications: Proteins in the ER are modified through processes like glycosylation, which attaches sugar molecules to specific amino acids. Other modifications include the formation of disulfide bonds and proper folding to achieve mature protein conformation. Ensuring Precision: Quality Control in the ER ER-Associated Degradation (ERAD): A surveillance system that identifies and disposes of misfolded or unassembled proteins. Misfolded proteins are retrotranslocated back into the cytosol, ubiquitinated, and targeted for degradation by the proteasome. Unfolded Protein Response (UPR): A cellular stress response triggered by the accumulation of unfolded proteins in the ER. UPR aims to restore normal function by halting protein translation, degrading misfolded proteins, and activating the signaling pathways that increase the production of molecular chaperones. UPR SIGNALING 3 Key UPR signal activator proteins Inositol requiring 1 (IRE1) PKR-like ER-kinase (PERK) Activating factor 6 (ATF6) Consist of 3 domains ER luminal domain (LD) Single pass membrane spanning domain Cytosolic domain Domain Organisation- LD UPR SIGNALING Adaptive unfolded protein response (UPR) signalling under acute endoplasmic reticulum (ER) stress. Accumulation of unfolded protein triggers UPR by activation of (B) PERK (C) Activating Transcription Factor 6. (A) inositol-requiring 1 (IRE1) This leads to upregulation of ER-associated degradation protein and folding chaperons to mitigate ER stress and maintain homeostasis. Ensuring Precision: Quality Control in the ER Clinical Implications: Persistent ER stress and an overwhelmed UPR can contribute to the development of diseases like neurodegeneration, diabetes, and cancer. Therapeutic strategies targeting ER stress pathways are being explored to treat these conditions. Clinical Correlations of Protein Folding Pathologies Defective Protein Folding: Misfolding can occur due to genetic mutations, environmental factors, or a combination of both, leading to loss of function or toxic gain of function. Misfolded proteins can aggregate, leading to cellular stress and activation of the UPR, which may result in apoptosis if homeostasis cannot be restored. Related Diseases: Cystic Fibrosis: Caused by mutations in the CFTR gene leading to misfolded CFTR protein, which results in faulty chloride ion transport. Alpha-1 Antitrypsin Deficiency: Due to mutations in the SERPINA1 gene, misfolded alpha-1 antitrypsin accumulates in the liver, impairing lung function due to unregulated elastase activity. Clinical Correlations of Protein Folding Pathologies Therapeutic Strategies: Pharmacological Chaperones: Small molecules that stabilize the native state of proteins, improving their folding and trafficking. Proteostasis Regulators: Compounds that modulate the UPR pathways, chaperone levels, and proteasomal degradation to alleviate stress. Gene Therapy: Strategies to replace defective genes or introduce correct copies to restore normal protein function. Clinical Case Study: Alpha-1 Antitrypsin Deficiency Background on Alpha-1 Antitrypsin (AAT): AAT is a protease inhibitor produced in the liver, functioning primarily to protect the lungs by inhibiting neutrophil elastase. It is synthesized as a single polypeptide chain that folds into a stable tertiary structure within the ER of hepatocytes. Pathogenesis of AAT Deficiency: Caused by mutations in the SERPINA1 gene, leading to the production of a misfolded variant of AAT called Z-AAT. Misfolded Z-AAT accumulates in the ER of hepatocytes, forming insoluble polymers that cause liver cell damage and ER stress. Clinical Case Study: Alpha-1 Antitrypsin Deficiency Disease Manifestation: Liver damage due to ER stress and apoptosis of hepatocytes. Reduced levels of functional AAT in the blood lead to unchecked neutrophil elastase activity, resulting in lung tissue damage and emphysema. Therapeutic Interventions: Augmentation Therapy: Infusion of purified AAT to restore its protective levels in the lungs. Small Molecule Correctors: Compounds that assist in proper folding and prevent polymerization of Z-AAT. Gene Therapy: Approaches to correct the underlying genetic defect, providing a source of functional AAT. Case History: Patient with Alpha-1 Antitrypsin Deficiency Patient Details: Age: 42 years old Sex: Male Medical History: Non-smoker, moderate alcohol use, no significant past medical history Family History: Father died at 55 from liver cirrhosis, mother has chronic obstructive pulmonary disease (COPD) Presenting Complaints: Shortness of breath with exertion for the past 6 months Chronic productive cough Recent episodes of wheezing Case History: Patient with Alpha-1 Antitrypsin Deficiency Diagnosis Clinical Workup: Physical Examination: Reduced breath sounds, wheezing upon auscultation, no jaundice or other signs of liver failure Pulmonary Function Tests (PFTs): Showed reduced FEV1/FVC ratio indicative of obstructive lung disease Liver Function Tests (LFTs): Mildly elevated AST and ALT, normal bilirubin levels Imaging: Chest X-ray revealed hyperinflated lungs; CT scan confirmed the presence of emphysema Genetic Testing: Performed due to the combination of early-onset emphysema and family history; confirmed homozygosity for the Z allele of the SERPINA1 gene Diagnosis: Alpha-1 Antitrypsin Deficiency: The patient's symptoms, imaging, and genetic profile are consistent with this diagnosis, with manifestations including early-onset pulmonary emphysema and liver involvement. Treatment and Management Immediate Management: o Smoking cessation counselling (despite patient being a non-smoker, counseling is standard due to the risk of exacerbating lung disease) o Bronchodilators to manage wheezing and improve breathability o Vaccinations to prevent respiratory infections (influenza and pneumococcal vaccines) Ongoing Treatment: o Augmentation Therapy: Initiation of intravenous AAT replacement therapy to increase circulating levels of functional AAT and slow the progression of lung disease o Lifestyle Modifications: Patient education on avoiding environmental pollutants and maintaining a healthy weight to reduce stress on the lungs and liver Treatment and Management Long-Term Management: o Regular monitoring of pulmonary function to assess progression of lung disease o Annual liver function tests and consideration of liver imaging to monitor for signs of liver disease progression o Patient education on recognizing signs of liver disease (jaundice, ascites, easy bruising) o Consideration for liver transplantation in the event of liver failure Special Considerations: o Referral to a support group for individuals with Alpha-1 Antitrypsin Deficiency o Genetic counselling for the patient and family members given the hereditary nature of the condition o Monitoring for potential side effects of augmentation therapy, such as transfusion reactions Follow-Up and Prognosis Follow-Up Care: The patient will have follow-up visits with a pulmonologist and hepatologist every 3 months initially, then annually or as clinically indicated. Monitoring will include PFTs, LFTs, imaging studies, and assessment of symptoms. Prognosis: With early diagnosis and appropriate management, the patient's lung function decline can be slowed, and liver disease can be monitored and managed. The prognosis is variable and depends on the level of lung and liver involvement and the patient's adherence to the treatment regimen. Conclusion and Key Takeaways Recap of Key Points: The ER is essential for protein synthesis, folding, and trafficking, ensuring proteins reach their destinations functional and intact. Misfolded proteins and ER stress can lead to diseases like Cystic Fibrosis and Alpha-1 Antitrypsin Deficiency. The UPR and ERAD are critical for managing ER stress and maintaining cellular homeostasis. Therapeutic interventions, including pharmacological chaperones, proteostasis regulators, and gene therapy, offer hope for treating protein misfolding diseases. Conclusion and Key Takeaways ER’s Role: The ER's multifaceted roles underscore its importance in cell biology, from acting as a gatekeeper of protein quality to a sensor of cellular stress. Dysfunctions in the ER's processes can have wide-ranging impacts, from isolated cellular issues to systemic diseases. Final Thoughts: Ongoing research into the ER's functions and stress responses is vital not only for understanding the fundamental biology but also for developing new treatments for complex diseases. The future of medicine lies in our ability to innovate and translate basic research on the ER into clinical applications that improve patient outcomes. End of Session