Clash Of The Titans! Convergence Of Tau And Alpha-Synuclein

Amyloid In Neurodegenerative Diseases

John Q. Trojanowski, M.D., Ph.D.Institute on Aging

Center for Neurodegenerative Disease Research Department of Pathology and Laboratory Medicine

University of PennsylvaniaPhiladelphia, PA

Neurodegenerative Diseases Characterized by Brain Amyloidosis

Disease Lesions Components

Parkinson’s Disease LBs -Synuclein

Dementia with Lewy Bodies LBs -Synuclein

Multiple System Atrophy GCIs -Synuclein

Alzheimer’s Disease SPs Aβ(Most common synucleinopathy!) NFTs Tau

LBs α-Synuclein

Prion diseases SPs Prions

Tauopathies NFTs Tau

Trinucleotide Inclusions Expanded Repeat Expansion PolyQ tracts

0

5

10

15

20

25

1950 1960 1970 1980 1990 2000 2010 2020 2030 2040 2050

High

Median

Low

Mill

ion

s

Year

US Population Demographics 85 Years of Age or Older: 1950-2050

Synucleinopathies• Parkinson’s disease - familial and sporadic*• Dementia with Lewy bodies • Multiple system atrophy• Neurodegeneration with brain iron

accumulation-1 (formerly H-SD) • Pure autonomic failure• REM sleep behavior disorder• Down syndrome*• Alzheimer’s disease* (The most common α-

synuclein brain amyloidosis!)• * These disorders are “triple” brain amyloidoses

with tau, Aβ and α-synuclein amyloid deposits

Normal Alpha Synuclein

• An abundant low Mr synaptic protein, present to a lesser extent in perikarya and axons, but also in oligodendroglia • Other members of the synuclein family

of synaptic proteins include beta- and gamma-synuclein • Function is unknown but may play roles

in synaptic transmission• Is a phosphoprotein, but role of alpha-

synuclein phosphorylation in its normal function is unknown

Alpha-Synuclein Mutations Cause Familial Parkinson’s Disease

H2N COOH

Hydrophobic Middle Negatively Charged

NAC peptide (aa 61-95)A30P

mutationA53T

mutation

= 6 imperfect repeats of 11amino acids with the conservedcore KTKEGV

Carboxy-terminusSection

Mutations promote protein aggregation and filament formation

Pathological Alpha-Synuclein

• Forms insoluble filamentous aggregates with the properties of amyloid• Amino acids 71-82 in the NAC domain are

the minimal, essential sequences required for fibrilization• Filamentous alpha-synuclein inclusions

form in neuronal perikarya, processes and in gliaI cells• Is abnormally phosphorylated, nitrated

and ubiquitinated

Alpha-Synuclein Pathological Inclusions

Lewy bodiesElecton microscopy

Lewy neuritesLewy bodies Glial cytoplasmic inclusions

Glial cytoplasmic inclusionsElectron microscopy

Environmental and/or Genetic

Risk Factors

Synuclein Dysfunction and/or Aggregation

Genetic Factors-Synuclein

Mutations or Duplications

APP, PS1, PS2 Mutations, DS

Neurodegeneration

?

Alpha-synuclein Dysfunction And Aggregation Play Central Roles in

Mechanisms of Neurodegenerative Disease

But hold on! Tau and Alpha-Synuclein Pathology Commonly Co-occurr in Neurodegenerative

Diseases – So what does this mean? • Diseases with tau, α-synuclein and abundant A deposits

– Lewy body variant of Alzheimer’s disease– Alzheimer’s disease– Familial Alzheimer’s disease– Down’s syndrome

• Diseases with tau, α-synuclein and scant/no A deposits

– Parkinson’s disease/Dementia with Lewy bodies

– Multiple system atrophy

– Guam ALS/PDC

– Neurodegeneration with brain iron accumulation type 1– Some cases of PSP, CBD, and Pick’s disease

Normal Tau Proteins

• Abundant low molecular weight microtubule (MT) associated proteins localized mainly in axons

• Promote MT polymerization, bind to MTs and stabilize MTs in the polymerized state

• Normally is phosphorylated at a range of Ser and Thr residues

• Phosphorylation negatively regulates binding of tau to MTs

4R2N

4R1N

4R0N

3R2N

3R1N

3R0N

1 441E2

R1 R2 R3 R4

R1 R2 R3 R41 412

R2R1

R2

R3 R41 383

E3

R3 R41 410

R1 R3 R41 381

R1 R3 R41 352

R1

E10SDS-PAGE

67kDa

62kDa

59kDa

54kDa

52kDa

48kDa

Six Human Brain Tau Isoforms Are Generated By Alternative Splicing

PHF-Tau Proteins• Insoluble

• Form filamentous amyloid deposits in neuronal cell bodies/processes and glia

• Aberrantly hyperphosphorylated at Ser/Thr; ubiquitinated

• Unable to bind to MTs unless dephosphorylated in vitro

AAUAAGAAGCUGGAUCUU----------CCGGGAGGCGGCAG-U

4411

Exon 10

N27

9K2

80K

Intron 9 g u

u

g a

g

a

c

c

u

L284

L

P30

1L/S

S30

5N/S

V33

7M

R40

6W

G C U AAAG

cc a

u

N279K 280K P301L

S305N S305SL284L

a

u

acgu

Intron 10

+3

+14

u/t

+16

g

+13

C

G27

2V

K25

7TI2

60V

G38

9R

t

+12

TP301S

Mutations Impairing Tau Protein Function

-G272V-280K -P301L-P301S -V337M-G389R-R406W

Mutations AlteringExon 10 Splicing

-N279K-280K -L284L-S305N -S305S

-Intron 10 mutationsE

342V

Tau Mutations Cause FTDP-17 By Different Mechanisms And Aggregation Of Tau Disrupts Axonal Transport Leading to Neurodegeneration

Mutations Promote Tau Aggregation-G272V -P301L –P301S-V337M

Tau Positive Inclusions in Neurons

DEMPEG

CBD

GUAMAD

PSP

FTDP17

Pick’s disease

Paired Helical Filaments (PHFs) Formed By Tau Are Building Blocks Of NFTs In AD Brain

• Two twisting strands with an apparent periodicity of 80nm and an alternating width between 8 and 20nm

• Molecular composition tau

From Lee et al. Science (1991) 251, 675-8

2N4R

2N3R

1N4R

1N3R

0N4R

0N3R

72kDa68kDa

64kDa

60kDa

Dephos. + + +- - -

67kDa

62kDa

59kDa

54kDa

52kDa

48kDa

• AD• ALS/PDC• Down’s syndrome• FTDP-17 (G272V, V337M,etc.)• GSS • Nieman-Pick disease type C

• FTDP-17 (K280)• Pick’s disease

• CBD• FTDP-17 (mutations

in I10, L284L, etc.)• PSP

Sarkosyl-insoluble Tau Bands Before and After Dephosphorylation

Tau Dysfunction And The Pathogenesis Of AD And Related Neurodegenerative Tauopathies

Genetic Factors

Tau Mutations

APP, PS1, PS2Mutations

Environmental Factors

Perturbation of 4R/3R Ratio

Loss of Tau Function

Gain of Toxic Function

Hyperphosphorylation

DeP-TauKinases

PhophotasesP-Tau

Tau Dysfunction

Tau Aggregation/ MT Loss

Impaired Transport & Neurodegeneration

? ?

Tau Pathology in Patients with the A53T Alpha-Synuclein Mutation

-syn Tau

Summary of -Synuclein Assembly Studies

-Synuclein readily assembles into 10 nm diameter filaments

• A53T mutation facilitates -synuclein assembly

-Synuclein is incapable of fibrillogenesis

• Residue 71-82 is required for -synuclein filament assembly

HSP70 PROTECTS AGAINST ALPHA- SYNUCLEIN INDUCED DOPAMINERGIC NEURON DEGENERTATION(Auluck et al., Science 2002)

Characterization of -Synuclein Transgenic Mice(Giasson et al., Neuron, 2002)

Abundant -synuclein Inclusions in A53T -synuclein Transgenic Mice

SNL-4 SNL-4 SNL-4Syn 303

Syn 303 Syn 505 Syn 505

Syn 506

Syn 303

Syn 303 Syn 505 Syn 505

Spinal cord Spinal cord Spinal cord Spinal cord

Raphe pons pons midbrain

pons pons locus ceruleus cerebellum

Accumulation of Detergent Insoluble -synuclein in the Spinal Cord of A53T Tg Mice

α-Synuclein Filaments can be Isolated from A53T Transgenic Mice

a-syn

a-syn tau

tau tau tau

tau overlay

Tau Pathology in the Spinal Cord and Midbrain of A53T a-Syn Tg Mice

Lessons From Nicoll, et al. (Nat. Med., 2003) For Therapy Of Brain Amyloidoses • Subtraction of Aβ from the brains of patients with

diseases having abundant tau, alpha-synuclein & A pathologies– Lewy body variant of Alzheimer’s disease– Sporadic/Familial AD– Down’s syndrome

• Will convert them to phenocopies of diseases with abundant tau and/or alpha-synuclein, but scant/no A pathologies– Parkinson’s disease/Dementia with Lewy bodies– Guam Marianna dementia/PDC

Model of Fibrilization in Neurodegenerative Disease

= -helix

= random coil= -pleated sheet

^ ^^

mRNA

Ribo-somes

N

N

chaperone

C

N N

C

Misfolded Smalloligomers

Inclusions

CellDeath

N

C

CellDeath

Proteasome

The Deleterious Consequences

Of Protein Misfolding

-synucleintauvesicles

Improper traffickingIncreased abundanceTau hyperphosphorylation

Microtubules

Pathologicalaggregates

More Deleterious Consequences Of Protein Misfolding

Five Year Delay in AD Onset Halves Prevalence & Incidence

It Takes Great A Team!

• Mark Forman• Benoit Giasson• Makoto Higuchi• Hiro Uryu• Bin Zhang• CNDR Members• Virginia Lee

• N. Bonini, J. Duda• J. Galvin, D. Galasko• L. Golbe, M. Grossman• H. Hurtig, T. Iwatsubo• C. Lippa, B. Miller• D. Murphy, M. Stern• Nat’l AD Coordinating

Center• NIA Alzheimer Disease

Centers• Penn Head Injury Center

Supported by Grants from the NIA, the Alzheimer’s Association, Michael J. Fox Foundation and the Families

of our Patients

DISCLAIMER

If you attended to the content of this lecture, you may have participated in effortful mental activity, and this may reduce your risk for dementia. * However, this lecture is not intended as a therapeutic intervention and there is no guarantee that this lecture has therapeutic benefit.

* Wilson et al. JAMA 287:742-748, 2003; Verghese et al. NEJM, 34:2508-2516, 2003