Lecture V. Cell Birth and Death Bio 3411 Wednesday September 15, 2010 1 Lecture V. Cell Birth &...

Lecture V. Cell Birth and Death

Bio 3411 Wednesday

September 15, 2010

September 15, 2010 1Lecture V. Cell Birth & Death

Readings

NEUROSCIENCE: 4th ed, pp 596-609 (end of Chapter 23: Construction of Neural Circuits)References posted:• †Abbott, A. (2009). Neuroscience: One hundred years of Rita. Nature, 458(7238), 564-567.

• †Berry, D. (2006). Apoptosis and signal transduction. http://www.wehi.edu.au/education/wehi-tv/?page=2 .

• †Cohen, S. (1987). Autobiography. http://nobelprize.org/nobel_prizes/medicine/laureates/1986/cohen-autobio.html

• †Hengartner, M. O. (2000). The biochemistry of apoptosis. Nature, 407(6805), 770-776.

• †Hollyday, M. (2001). Viktor Hamburger (1900-2001): A rememberance. Developmental Biology, 236(1), 1-2.

• †Raff, M. (1998). Cell suicide for beginners. Nature, 396(6707), 119-122.

______________________

†(pdfs on course websites: [http://artsci.wustl.edu/~bio3411/] & [http://www.nslc.wustl.edu/courses/Bio3411/bio3411.html])

2Lecture V. Cell Birth & DeathSeptember 15, 2010

http://www.wehi.edu.au/education/wehi-tv/?page=2

http://nobelprize.org/nobel_prizes/medicine/laureates/1986/cohen-autobio.html

Cited A

• Acehan, D., Jiang, X., Morgan, D. G., Heuser, J. E., Wang, X., & Akey, C. W. (2002). Three-dimensional structure of the apoptosome: implications for assembly, procaspase-9 binding, and activation. Mol Cell, 9(2), 423-432.

• Brenner, S. (1973). The genetics of behaviour. Br Med Bull, 29(3), 269-271.• Cleary, M. L., Smith, S. D., & Sklar, J. (1986). Cloning and structural analysis of cDNAs for bcl-2 and a

hybrid bcl-2/immunoglobulin transcript resulting from the t(14;18) translocation. Cell, 47(1), 19-28.• Cohen, S., Levi-Montalcini, R., & Hamburger, V. (1954). A Nerve Growth-Stimulating Factor Isolated from

Sarcom as 37 and 180. Proc Natl Acad Sci U S A, 40(10), 1014-1018.• Cowan, W. M., & Wenger, E. (1967). Cell loss in the trochlear nucleus of the chick during normal

development and after radical extirpation of the optic vesicle. J Exp Zool, 164(2), 267-280.• Ellis, R. E., & Horvitz, H. R. (1991). Two C. elegans genes control the programmed deaths of specific cells

in the pharynx. Development, 112(2), 591-603.• Ellis, R. E., Yuan, J. Y., & Horvitz, H. R. (1991). Mechanisms and functions of cell death. Annu Rev Cell Biol,

7, 663-698.• Fesik, S. W. (2000). Insights into programmed cell death through structural biology. Cell, 103(2), 273-282.• Gross, A., McDonnell, J. M., & Korsmeyer, S. J. (1999). BCL-2 family members and the mitochondria in

apoptosis. Genes Dev, 13(15), 1899-1911.• Haldar, S., Reed, J. C., Beatty, C., & Croce, C. M. (1990). Role of bcl-2 in growth factor triggered signal

transduction. Cancer Res, 50(22), 7399-7401.

September 15, 2010 Lecture V. Cell Birth & Death 3

• Hamburger, V. (1958). Regression versus peripheral control of differentiation in motor hypoplasia. Am J Anat, 102(3), 365-409.

• Hamburger, V. (1975). Cell death in the development of the lateral motor column of the chick embryo. J Comp Neurol, 160(4), 535-546.

• Hamburger, V. (1977). The developmental history of the motor neuron. Neurosci Res Program Bull, 15 Suppl, iii-37.

• Kerr, J. F., Wyllie, A. H., & Currie, A. R. (1972). Apoptosis: a basic biological phenomenon with wide-ranging implications in tissue kinetics. Br J Cancer, 26(4), 239-257.

• Korsmeyer, S. J. (1992). Bcl-2: an antidote to programmed cell death. Cancer Surv, 15, 105-118.• Landmesser, L., & Pilar, G. (1974). Synaptic transmission and cell death during normal ganglionic

development. J Physiol, 241(3), 737-749.• Levi-Montalcini, R., & Hamburger, V. (1951). Selective growth stimulating effects of mouse sarcoma on

the sensory and sympathetic nervous system of the chick embryo. J Exp Zool, 116(2), 321-361.• Levi-Montalcini, R., Meyer, H., & Hamburger, V. (1954). In vitro experiments on the effects of mouse

sarcomas 180 and 37 on the spinal and sympathetic ganglia of the chick embryo. Cancer Res, 14(1), 49-57.

• Schlesinger, P. H., Ferdani, R., Liu, J., Pajewska, J., Pajewski, R., Saito, M., Shabany, H., & Gokel, G. W. (2002). SCMTR: a chloride-selective, membrane-anchored peptide channel that exhibits voltage gating. J Am Chem Soc, 124(9), 1848-1849.

• Sulston, J. E., & Horvitz, H. R. (1977). Post-embryonic cell lineages of the nematode, Caenorhabditis elegans. Dev Biol, 56(1), 110-156.


Cited B

• Sulston, J. E., Schierenberg, E., White, J. G., & Thomson, J. N. (1983). The embryonic cell lineage of the nematode Caenorhabditis elegans. Dev Biol, 100(1), 64-119.

• Vaux, D. L., Cory, S., & Adams, J. M. (1988). Bcl-2 gene promotes haemopoietic cell survival and cooperates with c-myc to immortalize pre-B cells. Nature, 335(6189), 440-442.

• White, J. G., Horvitz, H. R., & Sulston, J. E. (1982). Neurone differentiation in cell lineage mutants of Caenorhabditis elegans. Nature, 297(5867), 584-587.

• Wiesmann, C., Ultsch, M. H., Bass, S. H., & de Vos, A. M. (1999). Crystal structure of nerve growth factor in complex with the ligand-binding domain of the TrkA receptor. Nature, 401(6749), 184-188.

• Xue, D., & Horvitz, H. R. (1997). Caenorhabditis elegans CED-9 protein is a bifunctional cell-death inhibitor. Nature, 390(6657), 305-308.

• Xue, D., Shaham, S., & Horvitz, H. R. (1996). The Caenorhabditis elegans cell-death protein CED-3 is a cysteine protease with substrate specificities similar to those of the human CPP32 protease. Genes Dev, 10(9), 1073-1083.

• Yang, X., Chang, H. Y., & Baltimore, D. (1998). Essential role of CED-4 oligomerization in CED-3 activation and apoptosis. Science, 281(5381), 1355-1357.

• Yuan, J. Y., & Horvitz, H. R. (1990). The Caenorhabditis elegans genes ced-3 and ced-4 act cell autonomously to cause programmed cell death. Dev Biol, 138(1), 33-41.


Cited C

What the last Lecture was about

• Mesoderm induces neuroectoderm in overlying ectoderm that gives rise to neuronal or epidermal cells.

• The “default” state of neuroectodermal cells is neuronal.

• Neuroectoderm secretes Bone Morphogenic Protein-4 (BMP-4), a signaling molecule that blocks the neuronal fate in neighboring neuroectodermal cells.

• Mesoderm secretes proteins - Chordin, Noggin, Follistatin - that block BMP-4 and neuroectodermal cells continue as neuronal progenitors.

• This inductive mechanism is conserved between vertebrates and invertebrates.

• These, and other similar, signaling mechanisms are used by the developing nervous system to control other events later in development.

• BMP-4 is a member of the Transforming Growth Factor-beta (TGF-β) family of signaling molecules.


What this Lecture is about

• Cell Death – Necrosis vs Apoptosis

• Promoting growth and survival – “trophism”

• Inhibition of the “death mechanism”

• Broader implications: neuroembryology; cancer

• Different critters - Same genes

• Molecular models

• Connection of Trophic Factors to cell death


from Greek“apo” meaning “separation”

&“ptosis” for “falling off”

Apoptosis

Kerr, J. F., et al.,(1972)


Types of Cell Death

Apoptosis (Programmed)

• Cell-Autonomous• Stereotypic• Rapid• “Clean” (dead cells eaten)

Necrosis (Provoked)

• Not Self-Initiated• Not Stereotypic• Can Be Slow• “Messy” (injury can spread)


Ellis, R. E., et al., (1991)

Removing a neuron’s targets, leads to its death

Deprived NormalHamburger, V. (1958,1977)


Neuronal death is central for normal NS development

Hamburger, (1975);Landmesser & Pilar, (1974);

Cowan & Wenger, (1967)


Neuron survival correlateswith target innervation

TargetInnervation

Neuronal Loss

Motor neuronsTarget

Muscles

Not all neuronsinnervate targets


DevelopmentProgresses

Axon Outgrowth

Target innervation determines which neurons survive

More targets(more neurons)

Fewer targets(fewer neurons)


DevelopmentProgresses


Levi-Montalcini, R., & Hamburger, V. (1951)

Mouse tumor (sarcoma) transplanted next to developing chick spinal cord causes axon sprouting consistent with a diffusible

factor - a nerve growth factor

ViktorHamburger

RitaLevi-Montalcini

StanleyCohen


Levi-Montalcini, et al. (1954);Cohen, S., et al. (1954)

A quantitative functional assay for Nerve Growth Factor (NGF) activity, using explanted cultures of sensory ganglia

Hollyday, M. (2001); Abbott, A. (2009).Cohen, S. (1987)

NGF is the founding member of a large gene familyof Neurotrophins (NTs), distantly related to insulin

NGF bindsas a dimer

to its receptor


Wiesmann, C., et al. (1999)

NGF/Neurotrophins Signal through Trk (tyrosine kinase) Receptors

NGF/NT

Multiple SignalingPathways

Via kinases and scaffolding proteins

Apoptosispathway

(PLC/PKC kinase)

Intracellular Ca+2

release, modulation of ion channels

Gene Activation/Repression

(PIK3/AKT kinase)

(Ras/MAP kinase)


Trk Receptors (TrkA, TrkB, TrkC, p75)

C. elegans is the model organism for molecular genetic studies

SydneyBrenner

JohnSulston

H. RobertHorvitz

Hypoderm

Bodywall Muscle

Cuticular CellsNeurons

PharynxIntestineVulva

Germ CellsGonad

Muscle

deve

lopm

enta

l tim

e

Neuronal Cell Death Lineages


Brenner, S. (1973) ; Sulston, J. E., & Horvitz, H. R. (1977); Sulston, J. E., et al. (1983); White, J. G., et al. (1982)

Programmed Cell Death of single identifiedneurons can be followed in live worms

P11aap


Sulston, J. E., & Horvitz, H. R. (1977)

2 Classes of C. elegans Cell Death Mutants

(pro-survival genes + pro-apoptosis genes)

(normal number of cells)


(fewer cells)


(extra cells)September 15, 2010 20Lecture V. Cell Birth & Death

WT

Mutant class I

Mutant class II

ced-3

ced-4 CellDeath

ced-9(pro-survival)

gene (pro-apoptosis)genes

Genetic analysis of cell death genes in C. elegans defines a genetic pathway

ced-9(lf) excessive cell death (fewer cells) animals dieas embryos

ced-3(lf) reduced cell death (extra cells) viableced-4(lf) reduced cell death (extra cells) viable

ced-9(lf) ced-3(lf) reduced cell death (extra cells) viableced-9(lf) ced-4(lf) reduced cell death (extra cells) viable

Ellis, R. E., et al., (1991); Ellis, R. E., & Horvitz, H. R. (1991)


XX

XX

t(14;18) Chromosomal Translocation Causes Human B-Cell Leukemia by Overexpression of Bcl-2

Stanley Korsmeyer

Bcl-2

Chromosome 18Ig Heavy Chain

Chromosome 14

Bcl-2

Chromosome 18

Ig Heavy Chain

Chromosome 14

t(14;18) Chromosomal Translocation

Bcl-2


Cleary, M. L., et al., (1986); Haldar, S., et al. (1990); Korsmeyer, S. J. (1992); Vaux, D. L., et al. (1988)

The “core” Cell Death genes found in C. elegans are conserved as multigene families in vertebrates

ced-9 / Bcl-2:Bcl-2: B-Cell Leukemia.• “Pro-survival” protein.• Inhibits release of cytochrome C from mitochondria (vertebrates).• Sequesters CED-4 from cytoplasm (worms).

ced-4 / Apaf:

Apaf: Apoptosis activity factor.• “Adaptor” or “scaffold” protein.• Aggregates inactive procaspase, causing auto-activation by proximity.• Requires cytochrome C, and ATP for multimerization (vertebrates).

ced-3 / Caspase: Caspase: Cysteineactive-site, aspartate cleavage-site, Protease.• “Terminator” protein.• Protease activity when activated by proteolysis.


Yang, X., (1998)

Xue, D., & Horvitz, H. R. (1997)

Xue, D., et al. (1996);Yuan, J. Y., & Horvitz, H. R. (1990)

Molecular Model for Apoptosis

activatedcaspase

(cascade)

(BH3 domains)

Apaf aggregation

single BH3domain protein

(egl-1)

mitochondria

Bcl-2(ced-9)

Apaf(ced-4)

caspase(ced-3)

---

Death

InactiveProcaspase

recruited

(procaspase recruitment) ---

---

Cytochrome C

(*Catalysis of the removal of

auto-inhibitory caspase domain*)


Gross, A., et al., (1999)

NGF is only one of multiple pathways to the “core” death mechanism, through many single-BH3 proteins

Single BH3domain protein



BCL-2

“Initiator”caspase-8

NGF

Initiation of apoptosis byextracellular ligands (FAS, TNF)

*

*

*

**

“Core” apoptotic components


Apaf/Cytochrome C Aggregate intoa 7-Spoke Apoptosome Complex (“Wheel of Death”)

+procaspase-9 (x7?)

procaspase-9

Acehan, D., et al. (2002)

Apaf gene

Cytochrome CCARD(caspase activationand recruitment domain)

ApafWD-40

WD-40

Single-particleElectron Microscope

Analysis


Pro-deathPro-survival

Single-BH3 domain molecules integrate multiple signals that trigger apoptosis.

Mitochondria integrate “Pro-survival” and “Pro-death”signals from a family of Bcl-2-like genes.

(BH3)(BH3)

“Pro-death” Single-BH3 domain proteins

complex with Bcl-2 to release cytochrome C

from mitochondria through “giant”

mitochondrial ionic channels.

pA

Diptheria Toxin(pore forming)

Bcl-xL(Bcl-2 like)

BH3


Schlesinger, P. H., et al., (2002), Fesik, (2000)

Molecular Animation of Cell Death Mediated by the FAS pathway

Berry, D. (2006) http://www.wehi.edu.au/education/wehi-tv/?page=2.


http://www.wehi.edu.au/education/wehi-tv/?page=2

What this Lecture was about

• Programmed cell death (apoptosis) is a physiological mechanism distinct from necrotic cell death.

• Apoptosis occurs widely during normal development of the nervous system.

• Isolation of specific molecules involved in promoting growth and survival – “trophism,” e.g., Nerve Growth Factor (NGF).

• What is the “death mechanism” that NGF (and other neruotrophins) inhibit?

• Broader implications: controlled cell death in neuroembryology vs uncontrolled cell growth of cancer.

• Gene homologies between organisms - humans and worms (nematodes)

• Molecular models for apoptosis

• How do trophic factors connect to this cell death pathway(s)?


END


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