Download Telomerase and the consequences of telomere dysfunction Carol W. Greider

Telomerase and the consequences
of telomere dysfunction
Carol W. Greider
Setting the stage: telomere function is
conserved
Ori
Marker
Linear plasmid
Add Tetrahymena telomeres
T
Transform
f
yeastt
Telomere
Ori
Marker
Telomere
Cell May 1982
Setting the stage: telomere elongation in
trypanosomes
Nature June 1983
Setting the stage: Yeast sequences added
onto Tetrahymena telomeres
Telomere
Ori
Marker
Maintain linear
plasm id in yeast
Remove right telomere
Add yeast genomic DNA
Yeast telom eres stabilizes
linear plasm id
Nature July 1984
Two different models for elongation
RECOMBINATION
3’
5
5’
DE NOVO ADDITION
3’
5’
5’
5
Strand invasion
Elongation
3’
Extension of 3’ end
3
Repair synthesis
Resolution &
p
synthesis
y
repair
Net telomere
elongation
Net telomere
elongation
The hunt for
telomerase
Telomerase activity
identified
Greider & Blackburn Cell 1985
Identification of the RNA component
Greider & Blackburn Cell 1987
Telomerase uses RNA template to add
TTGGGG repeats
Greider & Blackburn Nature 1989
What happens without telomerase?
Greider &
Harley 1988
Telomere shortening in cellular
senescence
Nature May 1990
Telomeres are shorter in cancer cells
Mol Cell Biol.
1990
Nature 1990
Telomerase protein component identified
Science 1997
TERT transfection extends the lifespan
off human
h
cells
ll
Science 1998
Using genetics to examine the role of
telomerase in mammals
Maria Blasco
Telomerase knockout mouse
m TR+ / -
mTR+ / +
m TR+ / -
mTR+ / -
mTR / mTR-
G1
G1
G2
G2
G3
G3
G4
G4
G5
Cell death
or
Senescence
G5
G6
G6
Blasco et al. Cell 1997
Quantiative analysis of telomere length
Telomeres shorten progressively in
telomerase-null mice
m TR+ / -
mTR+ / -
m TRTR / - (G1)
G1
G2
G2
G3
G3
G4
G4
G5
G5
G6
Blasco et al. Cell 1997
Loss of telomere sequence leads to loss
of telomere function
WT
KO-G2
KO-G4
KO-G6
The consequences of short telomeres
m TR+ / -
mTR+ / +
m TR+ / -
mTR+ / -
mTR / mTR-
G1
G1
G2
G2
G3
G3
G4
G4
G5
Cell death
or
Senescence
G5
G6
G6
Mike Hemann
Mice with 1/2 short and 1/2 long telomeres
show telomere dysfunction
m TR+ / -
m TR- / - G6
m TR
TR- / - (iG7)
m TR+ / - (iG7)
Hemann et al. Cell 2001
The shortest telomere triggers cell death
100
90
80
70
mTR+/-
60
50
40
MEAN 44
30
20
10
0
100
mTR-/G6
90
80
70
60
50
40
MEAN 24
30
20
10
0
140
mTR /
mTR-/(iG7)
120
100
80
60
MEAN 32
40
20
0
140
mTR+/(iG7)
120
100
80
60
MEAN 33
40
20
0
Hemann et al. Cell 2001
Short telomeres induce a DNA
damage response
Double strand break
Telomere shortening
DNA
Telomere
p53
Apoptosis
Senescence
Telomerase is required for cells that
must divide many times
CANCER CELLS
STEM CELLS
Short telomeres supress tumorigenesis
Cancer model
Effect
ff
off short
telomeres
Reference
Squameous cell
S
ll carcinoma
i
(DMBA)
D
Decrease
Gonzalez-Suarez
G
l
S
ett al.
l Nat
N t
Gen. 2000
Colon cancer
(APCmin)
Decrease
(apoptosis)
Rudolph et al. Nat Gen.
2001
Lymphoma& fibrosarcoma
(INK4a/Arf -/-)
Decrease
Greenberg et al. Cell 1999
T cell lymphoma
(ATM-/-)
Decrease
(apoptosis)
Qi et al
al. Can.
Can Res. 2003
Wong et al. Nature 2003
Hepatocellular carcinoma
(Alb-uPA-/-)
(Alb
uPA / )
Decrease
Farazi et al. Can. Res. 2003
Soft tissue sarcoma
(p53-/-)
Increase
(loss of apoptosis)
Artandi et al. Nature 2000
David Feldser
Eµ Myc induced B-cell
Eµ-Myc
B cell lymphoma growth
can be blocked by short telomeres
Myc+ m TR+ / -
mTR+ / -
Myc+ m TR- / - G1
Myc+ G1
G1
Myc+ G2
G2
Myc+ G3
G3
Myc+ G4
G4
Myc+ G5
Myc
y + G6
G5
G6
Feldser & Greider Cancer Cell
2007
Short telomeres protect against
B-cell lymphoma
Percen
nt alive
e
Eμ-myc
mTR-/- G6
Eμ-myc
mTR+/+ mice
Eμ-myc - mTR-/G1
D
Days
Feldser & Greider Cancer Cell
2007
Both apoptosis and senescence block
tumor growth
Short
telomere
p53
Bcl‐2
Senescence
Apoptosis
Feldser & Greider Cancer Cell
2007
Nature 2001
Clinical features of dyskeratosis congentia
• Dermatologic criteria
– Skin hyperpigmentation
– Oral leukoplakia
– Nail dystrophy
• Mortality
–Aplastic anemia
–Pulmonary fibrosis
–Cancer
Julian Chen
Telomerase RNA secondary structure
CR4/CR5
TERT
Human Telomerase
RNA
Pseudoknot/Template
TERT
SnoRNA
Nhp2
Nop10
Dyskerin
Mitchell et al. MCB
1999
Chen et al. Cell 2000
TERT mutations cause autosomal
dominant inheritance of dyskeratosis
congentia
Hopkins Family 1
I1



2
Genetic anticipation.
anticipation
Affected people are
heterozygous for hTR or
hTERT mutations.
II 1
2
3
4
5
6
7
?
III 1
2
3
4
Dominant negative or
h l i
haploinsufficiency?
ffi i
?
Armanios et al. PNAS
(2005)
Cast/Ei mice have short telomeres
similar to humans

Bred mTR null
allele onto Cast/Ei
background
Hemann & Greider NAR
2000
Ling Yang Hao
Ling-Yang
Progressive telomere shortening during
mTR+/- breeding
m TR+ / -
mTR+ / -
HG1
HG1
HG2
HG2
HG3
HG3
HG4
HG4
HG5
Short telomeres in CAST/EiJ mTR+/mTR /
mice
Hao et al. Cell 2005
Inheritance of short telomeres decreases
survival
m TR+ / -
m TR+ / -
HG1
HG1
HG2
HG2
HG3
KO(G2)
HG3
HG4
HG4
HG5
KO(G3)
KO(G4)
KO(G5)
Margaret
Strong
Perce
ent surviv
val
Decreased survival of mTR
mTR-// from later
generation heterozygous crosses
100
WT
KO from HG3 parents
75
KO from HG6 parents
KO from HG8 parents
50
25
0
0
100
200
300
400
D ays
Inheritance of short telomeres decreases survival.
genetic anticipation
p
in dyskeratosis
y
congenita.
g
Resembles g
Telomere shortening during mTR+/mTR /
breeding affects wildtype offspring
m TR+ / -
m TR+ / -
HG1
HG1
HG2
HG2
HG3
HG3
HG4
WT5*
WT5
HG4
HG5
KO(G5)
Short telomeres in Wt5*
Wt5 mice
HG4
KO
HG4
HG5
WT5*
Hao et al. Cell 2005
Short telomeres cause phenotypes in
Wt* mice
Testis Apoptosis
Genetic disease in the absence of telomerase
genetic disease))
mutation ((Occult g
Mary Armanios
Telomere shortening leads to genetic
anticipation
Hopkins Family 1
I1

Genetic anticipation
off pulmonary
l
fibrosis.
II 1
2
2
3
4
5
6
7
?
III 1
2
3
4
Armanios et al. PNAS
(2005)
NEJM 357 p1317
(2007)
Table 2 Spectrum
S ectru of bone marrow,
arro
lung, and liver disease seen in
y
of
individuals with syndromes
telomere shortening
Annu. Rev. Genomics Hum. Genet. 2009.
10:45-61
Many phenotypes from telomere
shortening syndromes share features of
age related disease

Bone marrow failure

Liver disease

Immune senescence


Chemotherapy
intolerance
Increased cancer
incidence

Pulmonary fibrosis
•Implies short telomeres may play a role in disease
without mutation.
Wildtype
yp mice with short telomeres show a
decreased number of B and T lymphocytes
B Cells
T Cells
Armanios et al. AJHG 2009
Decreased CD4/CD8 ratio in mice with
short telomeres is a hallmark of immune
senescence
*
**
wt9*1
HG9
20
2.0
CD4/CD8
RATIO
1.5
1.0
0.5
0.0
wt
Armanios et al. AJHG 2009
Extensive telomere length heterogeneity
in the human population
Valdes et al, 2005 Lancet
(2005)
•Combined effect of initial telomere length and environmental history may
contribute to age related disease.
Summary
y and implications
p

Telomerase is essential for telomere maintenance.

Telomere shortening leads to cell death or
senescence after many cell divisions.

Short telomeres inhibit tumor growth through
apoptosis or senescence.

Telomerase inhibitors may be effective in cancer
therapy.

Haploinsufficiency for telomerase causes telomere
shortening.

Short telomeres limit cell g
growth even in the
presence of telomerase.

Short telomeres may limit long term stem cell division.
Summary
y and implications
p

IInheritance
h it
off short
h t ttelomeres
l
causes phenotypes
h
t
even in wildtype animals (Wt* or occult genetic
disease).

Short telomeres may cause loss of tissue renewal in
normal aging population.

Telomere
T
l
llength
th may predict
di t onsett off certain
t i age
related diseases.
Telomere shortening during mTR+/mTR /
breeding affects wildtype offspring
m TR+ / -
m TR+ / -
HG1
HG1
HG2
HG2
HG3
HG3
HG4
WT5*
WT5
HG4
HG5
KO(G5)
Decreased survival of mTR+/mTR / from later
generation heterozygous crosses
Inheritance of short telomeres decreases survival.
genetic anticipation
p
in dyskeratosis
y
congenita.
g
Resembles g
Armanios et al. AJHG 2009
Telomere length equlibrium is
reestablished after five generations
Armanios et al. AJHG (2009)
Wt mice reestablish telomere
Wt*
equilibrium after several generations
mTR+ / - HG4
mTR+ / - HG4
mTR+ / + WT5*1
m TR+ / + WT5*1
m TR+ / + WT5*2
mTR+ / + WT5*2
mTR+
TR / + WT5*3
mTR+ / + WT5*4
mTR+
TR / + WT5*3
mTR+ / + WT5*4
Armanios et al. AJHG 2009