2009 Lecture 17

From CellBiology

Cell Death 1

recycling

Introduction

We have discussed the cell lifespan and how cell's are born. These next two lectures will discuss how cells die in response to stress, pathological conditions and by programmed cell death. This first lecture will also introduce the concept of cellular recycling and the response to cellular stress. The next lecture Cell Death 2 will focus on programmed cell death by apoptosis.

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Lecture 17: Cell Death 1

Archive

MH - note that content listed below will not match exactly current lecture structure but has been selected as having similar content

Cell Lifespan

  • Neutrophil - 6-7 hours circulating, 4 days in tissue
  • Red blood cell - 120 days
  • Brain neuron - 50 - 100 years

Cell Recycling

There are a number of different cellular mechanisms and processes for reusing cellular components or removing abnormal products. These pathways can also be utilized during periods when the cell is placed under specific or limited growth conditions. Autophagy is an important part of this process, but will be covered under the Cell Death section of this current lecture.

Proteasome

Figure 6-88. Two general ways of inducing the degradation of a specific protein Figure 6-86. The proteasome

Ubiquitin-mediated protein degradation
  • Protein complex that degrades cellular proteins by proteolysis
  • located in nucleus and cytoplasm

Functions

  • regulate protein levels and degrade misfolded proteins

Structure

  • 26S proteasome Mr 2000 kDa
    • two 19S regulatory caps -ATPase active sites and ubiquitin binding sites
    • one 20S core hollow structure - catalytic core


Ubiquitin

Ubiquitin

Calpain

Function

  • proteolysis in embryogenesis, regulation of key enzymes, structural proteins and in proinflammatory responses
  • mediates MMP2 expression and cell migration (fibroblasts and leukemic cells)
  • contribute to cell death in neurons by cleaving essential cytoskeletal proteins
  • tissue damage in response to pathological events (myocardial infarcts, stroke, atherosclerosis, and brain trauma)
  • Deregulated calpain activity following loss of Ca2+ homeostasis
  • intracellular free calcium concentration (Ca2+i)regulator in some late apoptotic signaling


MH - calcium will be covered again in signaling.

Structure

  • 13 genes in mammals
  • heterodimer large 80 kDa catalytic and a small 30 kDa regulatory subunit

Links: PMID: 17608959 | Physiol. Rev - The Calpain System

Cell Stress

The term "cell stress" can cover many different issues. It is used here to describe two specific circumstances: when normal cellular processes may function abnormally (protein misfolding), or under specific limiting growth conditions (starvation). Under these conditions the cellular response is graded, initially to correct the problem (Unfolded Protein Response) or reuse existing resources (Autophagy).

Endoplasmic Reticulum Stress

  • Endoplasmic Reticulum functions
    • protein synthesis
    • lipid metabolism
    • calcium regulation (Ca2+) storage, release, signaling

Abnormal protein folding can lead to the Unfolded Protein Response (UPR)

  • accumulation of misfolded proteins
  • aggregate in the ER lumen
  • causes ER stress

Unfolded Protein Response (UPR)

MBoC - Figure 12-56. The unfolded protein response in yeast

  • decrease in the arrival of new proteins into the ER
    • preventing additional protein misfolding and overloading of the organelle
  • increase in the amount of ER chaperones
    • increasing the folding capacity of the ER to deal with misfolded proteins
  • increase in the extrusion of irreversibly misfolded proteins from the ER
    • subsequently degradation of these proteins in the proteasome

If all the above UPR steps fail, cell death by apoptosis is triggered.

Classification of Cell Death

Classification of cell death: recommendations of the Nomenclature Committee on Cell Death 2009 Cell Death and Differentiation (2009) 16, 3–11; doi:10.1038/cdd.2008.150; published online 10 October 2008 PMID: 18846107

Classified by cell death mode and morphological features (modified from Table 2)

Cornification

  • Elimination of cytosolic organelles
  • Modifications of plasma membrane
  • Accumulation of lipids in keratohyalin granules in stratum granulosum
  • Extrusion of lipids in the extracellular space
  • Desquamation (loss of corneocytes) by protease activation

Cornified envelope - formation or ‘keratinization’ is specific of the skin to create a barrier function. Although apoptosis can be induced by injury in the basal epidermal layer (e.g., UV irradiation), cornification is exclusive of the upper layers (granular layer and stratum corneum).

Autophagy

  • Lack of chromatin condensation
  • Massive vacuolization of the cytoplasm
  • Accumulation of (double-membraned) autophagic vacuoles
  • Little or no uptake by phagocytic cells, in vivo

Autophagic cell death - defines cell death occurring with autophagy, though it may misleadingly suggest a form of death occurring by autophagy as this process often promotes cell survival.

Necrosis

  • Cytoplasmic swelling (oncosis)
  • Rupture of plasma membrane
  • Swelling of cytoplasmic organelles
  • Moderate chromatin condensation

Necrosis - identifies, in a negative fashion, cell death lacking the features of apoptosis or autophagy. Note that necrosis can occur in a regulated fashion, involving a precise sequence of signals.

Apoptosis

  • Rounding-up of the cell
  • Retraction of pseudopodes
  • Reduction of cellular and nuclear volume (pyknosis)
  • Nuclear fragmentation (karyorrhexis)
  • Minor modification of cytoplasmic organelles
  • Plasma membrane blebbing
  • Engulfment by resident phagocytes, in vivo

Apoptosis - is the original term introduced by Kerr et al. to define a type of cell death with specific morphological features. Apoptosis is NOT a synonym of programmed cell death or caspase activation.

Cornification

Epidermis cartoonCornified Epithelium

Caspase-14 and epithelial cornification

Occurs in the skin epithelium and occurs in the upper layers (granular layer and stratum corneum)

  • Elimination of cytosolic organelles
  • Modifications of plasma membrane
  • Accumulation of lipids in keratohyalin granules in stratum granulosum
    • F-granules (histidine-rich) are large, irregularly shaped granules
    • L-granules (sulphur-rich)
  • Extrusion of lipids in the extracellular space
  • Desquamation (loss of corneocytes) by protease activation
  • Apoptosis can also be induced by injury to the basal epidermal layer
    • UVB irradiation, chemicals, cytotoxic cytokines

Caspase 14

  • enzyme, cysteinyl aspartate–specific proteinase
  • recently identified as having a role in this process
  • activation correlates with cornification
  • not clear yet exact role in filaggrin processing, there are 2 theories
    • cleave filaggrin fragments - leading to further degradation into free amino acids by another endo- and/or exopeptidase
    • directly or indirectly activate an endo- and/or exopeptidase - further processes smaller filaggrin fragments


Autophagy

Autophagy types

The Cell - Lysosomes in phagocytosis and autophagy

Autophagosome formation Movies PMID: 18725538

See also Lecture - Endocytosis

Autophagy Processes

There are several different classifications of autophagy this is probably the simplest.

  1. macroautophagy (also called autophagy)
  2. microautophagy
  3. chaperone-mediated autophagy
  • to remove abnormal cytoplasmic organelles and components, it is also a stress response
    • cellular self-catabolic process "eating oneself”
  • initial sequestered in a structure called a phagophore
  • which then closes into a double membrane vesicle the autophagosome
    • some autophagosomes formed in a PI3P-enriched (phosphatidylinositol 3-phosphate) membrane compartment dynamically connected to the endoplasmic reticulum
  • an autophagosome fuses with a lysosome
  • Regulated process of the degradation and recycling of organelles and cellular components
  • Resulting in organelle turnover and in the bioenergetics of starvation
  • Could result in cell death
    • through excessive self-digestion and degradation of essential cellular constituents

Is autophagy a cell survival or a cell death pathway?

Necrosis

Neuron necrosis apoptosis.jpg

Necrosis ethidium homodimer marker
  • Greek, nekros = corpse
  • pathological cell death from extrinsic injury
    • tissue damage
  • autoimmune insulin-dependent diabetes?
  • irreversible

Induction

  • tumor necrosis factor, double-stranded RNA, viral infection or bacterial toxins
  • does not shut down of protein synthesis
    • occurs in apoptosis due to caspase-dependent breakdown of eukaryotic translation initiation factor (eIF) 4G, activation of the double-stranded RNA-activated protein kinase PKR, and phosphorylation of its substrate eIF2-

Early stages

  • cell and organelles (mitochondria) swell (oncosis)
    • (Greek, onkos = 'swelling') previously described as a separate form of cell death
  • due to disruption of plasma membrane
  • cell contents leak out leading to inflammation and necrosis

Late stage

  • loss of cell membrane integrity
  • finally cell disintegration
  • cell lysis can also trigger an inflammatory response
    • leading to further inflammation and damage
    • triggering a cycle of death

Bacteria - S. aureus

  • pore-forming secreted toxins can induce necrosis

Prothymosin-alpha 1 (ProT)

  • inhibits necrosis
  • switches cells from necrosis to apoptosis

Programmed Cell Death?

Until very recently this has been exclusively about apoptosis, covered in Cell Death 2.

There is a theory though that there is no such thing as "unregulated" cell death and that even necrosis may involve a regulated program. In addition, a novel cell death pathway has been identified in neutrophils in the fight against pathogens.

Neutrophil Extracellular Traps

File:JCB200606027.V2.mov

Neutrophil extracellular traps

Neutrophils in circulation are targeted by cytokines to migrate into infected tissues, where they activate, and engulf pathogens into a phagosome.

A second defense mechanism has recently been described Neutrophil Extracellular Traps (NETs)

  • composed of chromatin decorated with cytoplasmic proteins in the extracellular space
    • bind Gram-positive and -negative bacteria, as well as fungi

Pathway

  • requires generation of reactive oxygen species (ROS) by NADPH oxidase
  • lobulated nuclear morphology lost
  • euchromatin and heterochromatin distinction lost
  • all the internal membranes disappear
    • allowing NET components to mix
  • NETs emerge from the cell as the cytoplasmic membrane is ruptured
    • process distinct from necrosis or apoptosis

Chronic Granulomatous Disease (CGD)

  • mutations in NADPH oxidase
  • cannot activate this cell-death pathway or make NETs
  • these individuals are susceptible to infections

Links: Medline Plus - CGD | OMIM 233700

References

Textbooks

Molecular Biology of the Cell

Alberts, Bruce; Johnson, Alexander; Lewis, Julian; Raff, Martin; Roberts, Keith; Walter, Peter New York and London: Garland Science; c2002

Molecular Cell Biology

Lodish, Harvey; Berk, Arnold; Zipursky, S. Lawrence; Matsudaira, Paul; Baltimore, David; Darnell, James E. New York: W. H. Freeman & Co.; c1999

The Cell- A Molecular Approach

Cooper, Geoffrey M. Sunderland (MA): Sinauer Associates, Inc.; c2000

Madame Curie Bioscience Database

Chapters taken from the Madame Curie Bioscience Database (formerly, Eurekah Bioscience Database) Eurekah.com and Landes Bioscience and Springer Science+Business Media; c2009

Search Online Textbooks

Journals

"Apoptosis is an international peer-reviewed journal devoted to the rapid publication of innovative basic and clinically-oriented investigations into programmed cell death. It aims to stimulate research on the basis of mechanisms of apoptosis and on its role in various human disease processes including: cancer, autoimmune disease, viral infection, AIDS, cardiovascular disease, neurodegenerative disorders, osteoporosis and ageing. The editors intend to encourage the development of clinical therapies against apoptosis-related diseases."
"Autophagy, is a unique peer-reviewed journal with an international audience that covers the following topics: Macroautophagy, microautophagy, specific organelle degradation (e.g., pexophagy) and additional autophagic processes including chaperone-mediated autophagy; The molecular mechanism of autophagy including the characterization of structural proteins and structure/function relationships; Signaling and autophagic regulation; The role of autophagy in diseases including cancer, neurodegeneration and myopathies; Developmental roles of autophagy and its connection with aging; Autophagy in microbial invasion and the immune response; and Autophagy in cell death."

PubMed

Reviews

  • Taatjes DJ, Sobel BE, Budd RC. Morphological and cytochemical determination of cell death by apoptosis. Histochem Cell Biol. 2008 Jan;129(1):33-43. Epub 2007 Nov 14. Review. PMID: 18000678
  • Denecker G, Ovaere P, Vandenabeele P, Declercq W. Caspase-14 reveals its secrets. J Cell Biol. 2008 Feb 11;180(3):451-8. Epub 2008 Feb 4. Review. PMID: 18250198
  • Artal-Sanz M, Tavernarakis N. Proteolytic mechanisms in necrotic cell death and neurodegeneration. FEBS Lett. 2005 Jun 13;579(15):3287-96. Epub 2005 Apr 2. Review. PMID: 15943973

Articles

  • Kroemer G, Galluzzi L, Vandenabeele P, Abrams J, Alnemri ES, Baehrecke EH, Blagosklonny MV, El-Deiry WS, Golstein P, Green DR, Hengartner M, Knight RA, Kumar S, Lipton SA, Malorni W, Nuñez G, Peter ME, Tschopp J, Yuan J, Piacentini M, Zhivotovsky B, Melino G; Nomenclature Committee on Cell Death 2009. Classification of cell death: recommendations of the Nomenclature Committee on Cell Death 2009. Cell Death Differ. 2009 Jan;16(1):3-11. Epub 2008 Oct 10. PMID: 18846107
  • Axe EL, Walker SA, Manifava M, Chandra P, Roderick HL, Habermann A, Griffiths G, Ktistakis NT. Autophagosome formation from membrane compartments enriched in phosphatidylinositol 3-phosphate and dynamically connected to the endoplasmic reticulum. J Cell Biol. 2008 Aug 25;182(4):685-701. PMID: 18725538
  • Ueda H, Fujita R, Yoshida A, Matsunaga H, Ueda M. Identification of prothymosin-alpha1, the necrosis-apoptosis switch molecule in cortical neuronal cultures. J Cell Biol. 2007 Mar 12;176(6):853-62. PMID: 17353361
  • Fuchs TA, Abed U, Goosmann C, Hurwitz R, Schulze I, Wahn V, Weinrauch Y, Brinkmann V, Zychlinsky A. Novel cell death program leads to neutrophil extracellular traps. J Cell Biol. 2007 Jan 15;176(2):231-41. Epub 2007 Jan 8. PMID: 17210947 PMC JCB - NETs
  • Brinkmann V, Zychlinsky A. Beneficial suicide: why neutrophils die to make NETs. Nat Rev Microbiol. 2007 Aug;5(8):577-82. Review. PMID: 17632569
  • Steer SA, Scarim AL, Chambers KT, Corbett JA. Interleukin-1 stimulates beta-cell necrosis and release of the immunological adjuvant HMGB1. PLoS Med. 2006 Feb;3(2):e17. Epub 2005 Dec 20. PMID: 16354107
  • Saelens X, Festjens N, Parthoens E, Vanoverberghe I, Kalai M, van Kuppeveld F, Vandenabeele P. Protein synthesis persists during necrotic cell death. J Cell Biol. 2005 Feb 14;168(4):545-51. Epub 2005 Feb 7. PMID: 15699214
  • Denecker G, Hoste E, Gilbert B, Hochepied T, Ovaere P, Lippens S, Van den Broecke C, Van Damme P, D'Herde K, Hachem JP, Borgonie G, Presland RB, Schoonjans L, Libert C, Vandekerckhove J, Gevaert K, Vandenabeele P, Declercq W. Caspase-14 protects against epidermal UVB photodamage and water loss. Nat Cell Biol. 2007 Jun;9(6):666-74. Epub 2007 May 21. PMID: 17515931


  • Kerr, J. F., Wyllie, A. H. & Currie, A. R. Apoptosis: a basic biological phenomenon with wide-ranging implications in tissue kinetics. Br. J. Cancer 26, 239-257 (1972). PMID: 4561027

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2009 Course Content

Lectures

Cell Biology Introduction | Cells Eukaryotes and Prokaryotes | Cell Membranes and Compartments | Cell Nucleus | Cell Export - Exocytosis | Cell Import - Endocytosis | Cell Mitochondria | Cell Junctions | Cytoskeleton Introduction | Cytoskeleton 1 Intermediate Filaments | Cytoskeleton 2 Microtubules | Cytoskeleton 3 Microfilaments | Extracellular Matrix 1 | Extracellular Matrix 2 | Cell Cycle | Cell Division | Cell Death 1 | Cell Death 2 | Signal 1 | Signal 2 | Stem Cells | Stem Cells | Development | Revision

Laboratories

Introduction to Lab | Microscopy Methods | Preparation/Fixation | Immunochemistry | Cell Knockout Methods | Cytoskeleton Exercise | Confocal Microscopy | Tissue Culture 1 | Tissue Culture 2 | Microarray Lab visit

Dr Mark Hill 2015, UNSW Cell Biology - UNSW CRICOS Provider Code No. 00098G