PCL481H1 Lecture Notes - Lecture 7: Autophagy, Ripk3, Ripk1
26 Feb 2017
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Autophagy and Autophagic cell death
• Autophagy: Normal intracellular process where stress-damaged organelles along with some cytoplasm-
o Are sequestered in vesicles for delivery to lysosomes for degradation
o Re-establishes homeostasis and promotes cell survival
o Regulated set of pathways to allow the cell to remain intact and remove damaged organelles.
• Autophagic cell death: an excess level of the above will result in cell death-
o Autophagy taken to an unfortunate extreme
• Autophagy followed by the cell surviving and will replace the organelles
o But too much will lead to autophagic cell death. ****
• Drug inibits vps4 inhibits both→ necrosis
Cross-talk between cell death modes
Picture
Relationship between toxicant dose and cell response
• Some chemicals may produce autophagy or apoptosis at low doses, and necrosis at higher doses
• Some chemicals may exhibit multiple modes of cell death
• Some chemicals may produce different cell death mechanism in different cell types
• If both targets the mitochondrian and membrane→ both modes
Induction of mitochondrial autophagy by resveratrol in ovarian cancer cells
• Microscopy- visualize single cells, use fluorescently labeled (antibody) biomarkers to check for toxicity
o Targets autophagy marker microtubule-associated protein:
▪ 1A/1B-light chain 3 (LC3)
▪ Mitochondrial leakage marker cytochrome C (Cyct C)
• Orange colour indicates that the two markers co-localize in the cells
• Resveratrol (a chemical present in red wine) induces mitochondrial autophagy in these cells
• Red wine→ can produce autophagy at higher doses
• Orange = foci
Necroptosis
• Regulated or programmed necrosis
• Allows regulated death in cells that cannot undergo apoptosis
• Caspase-independent
• Simply because there are particular pathways that specially lead to this caspase independent pathway→
o Distinct from apoptosis.
• Can be initiated by binding of ligands such as TNFa to cell surface death receptors
o Binding recruits adaptor proteins that produce an activating interaction between:
▪ Kinases RIPK1 and RIPK3
• Activation→ phosphorylation & oligomerization of mixed lineage kinase domain-like PRO. (MLKL)→
o Which attacks the plasma membrane causing cell death→
o In terms of simple biomarkers it looks like normal necrosis.
• MLKL is able to permeablize the plasma membrane- regulated
find more resources at oneclass.com
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Document Summary
Induction of mitochondrial autophagy by resveratrol in ovarian cancer cells: microscopy- visualize single cells, use fluorescently labeled (antibody) biomarkers to check for toxicity, targets autophagy marker microtubule-associated protein: In terms of simple biomarkers it looks like normal necrosis: mlkl is able to permeablize the plasma membrane- regulated. Mitochondiral function: oxidative phosphorylation- generation fo atp and citric acid and the etc, heat generation, control of apoptotic cell death, signaling via mitochondrial ros, calcium storage and signaling, regulation of membrane potential, steroid synthesis, heme synthesis. Mitochondrial dna: 37 genes, 13 for proteins in oxidative phosphorylation, 22 for trnas, 2 for rrna, inherited from mom, many mitochondrial function used imported gene products, mitochondrion is not self-sufficient. And the 4 comlexes: atp synthase, of adp to atp requires transport proteins and carrier proteins / enzymes. The electron transport chain: series of 4 complexes+ 1.
