Download Mechanisms of p53 Regulated Apoptosis

p53: To Be or Not to Be…
Ashim Malhotra
Bio 722
Instructor: Dr. Lockshin
March, 24, 2003
What is p53?
• Tumor suppressor phosphoprotein with 4 domains; Conserved in
vertebrates
• Gene name: Human: TP53, 17p13.1; Mouse : Trp53
• Gene structure: 19.21 kb; 11 exons
• Subcellular Location: Nuclear; in a punctate granular/vesicular
patter
• Evolution: Xenopus (68% homology)
• Sequence comparison: shows 5 highly conserved regions, coinciding
with mutation clusters found in p53 in human cancers
Why talk of p53?
• 27, 458
• Molecule of the Year, Science, 1993
• Guardian of the Genome; Cellular Gatekeeper for
Growth & Division (Levine, 1997 review)
• Emergency brake in the cell cycle
• Bio 722! Involved in PCD
What does it do?
• Suppresses progression through the cell cycle in
response to DNA damage
• Initiates apoptosis if the damage to the cell is
severe
• Often as a tumor suppressor
• A potent transcription factor
Involvement in diseases; putative
applications
• p53 gene mutations are the most frequently observed
genetic lesions in human cancers
• p53 and radiation sensitivity: Problems in radiotherapy
• Mutant p53: problems in tumor hypoxia
• Gene therapy with p53; Nature 1996
• Clinical Phase trials: (I) Adenovirus vectors expressing
p53 in tumors casues regression of mouse lymphomas
Come into the mad world of
p53…
Discovery of p53
The viral approach
The serological approach
Courtesy:
Courtesy:
The IARC Human p53 Mutation
Database
Current release has nearly 7000 entries detailing:
•
•
•
•
•
Location of point mutations
Description of mutation
Resultant tumor type
Exposure information
Pointer to original reference article
http://www.iarc.fr/p53
Protein structure & DNA binding
• Domain: Specific interactions with bcl-2 require
ankyrin & SH3 domains
• p53 binds as a tetramer to PBS & activates expression
of downstream genes inhibiting growth &/or invasion
Mechanisms of p53 inactivation
1.
2.
3.
4.
5.
p53 binding to PBS. Deletion of one or both p53 alleles=
expression of tetramers & expression of the growth inhibitory
genes.
Nonsense or splice site mutations result in protein-truncation
preventing oligomerization reducing tetramers: lung &
esophagus.
Missense mutations resulting in dominant-negative effects &
even greater reduction of functionally active tetramers: colon,
brain, lung, breast, skin & bladder.
Expression of the HPV - E6 gene causes functional inactivation
of p53 through binding and degradation.
The p53 pathway may also be disrupted by alteration of a
cellular gene, MDM2.
p53 Polymorphisms
• 2 p53 variants, 1 containing arginine & the other
containing proline.
• Arginine containing p53 kills cancer cell more
effeciently than the one containing proline.
• The proline form has an enhanced frequency in
African Americans & darker skinned people living
closer to the equator!!
• Arginine form travels out of the nucleus and into the
Mt better than the proline form.
Dumont, Leu & Pietra, Nature Genetics, March 2003
What does it do?
•Suppresses progression through the cell cycle in
response to DNA damage
• Initiates apoptosis if the damage to the cell is
severe
• Often as a tumor suppressor
• A potent transcription factor
Convergence of distinct stimuli at p53
de Stanchina et al, Genes and Development, 1998
Pathways! Pathways!
P on S-15
UV
DNA- damage
P on S-20
Cytostasis
Mdm2
p53
p53
P53 induced cell cycle arrest
I
N
H
p53
I
p21
B
I
T
I
Cyclin-Cdk2
complex
Hyp-Rb/E2F complex
O
N
Cytostasis
S-20 phosphorylation in response to DNA damage
Cheha , Malikzay, Stavridi & Halazonetis. Cell Biology, 1999
Summary of p53 stabilization pathways after
DNA damage
Giaccia, and Kastan, Genes and Development, 1998
p53 / Rb connection
Sherr, Science, 1996
What does it do?
•Suppresses progression through the cell cycle in
response to DNA damage
• Initiates apoptosis if the damage to the cell is
severe
• Often as a tumor suppressor
• A potent transcription factor
p53 associated apoptosis
•
Of 7,202 transcripts identified, only 14 (0.19%) are markedly
increased in p53-expressing cells.
•
Many of these genes encode proteins that generate or respond
to oxidative stress.
•
These observations suggest that p53 causes apoptosis through a
three-step process:
1. Transcriptional induction of redox-related genes.
2. Formation of reactive oxygen species and.
3. Oxidative degradation.
Polyak, Xia, Zweier, Kinzler & Vogelstein, Nature, 1997.
Possible steps in p53 mediated apoptosis:
step I
UV
DNA
PK
DNA damage
p53
DNA PKcs mediates apoptosis but not cytostasis
via p53
Wang, Guo ,Ouyang , Li,Cardo ,Kurimasa,Chen,Fuks, Ling , & Li, Cell Biology, 2000
P53! Anoint thee wench!
Peg3/PW1
Transcriptional
Bax
p53
Activation
PUMA
Bax sublocalization with Peg3
Yibin & Wu, Cell Biology, 2000
Peg3/PW1 causes translocation of Bax to
Mitochondria
Yibin & Wu, Cell Biology, 2000
Scheme for PUMA induced Mt translocation of
Bax
PUMA
p53
Mt
Bcl-Xl Translocation Bax
What does it do?
•Suppresses progression through the cell cycle in
response to DNA damage
• Initiates apoptosis if the damage to the cell is
severe
• Often as a tumor suppressor
• A potent transcription factor
To P or not to P:
c-Myc & p53
p53
C-Myc
Miz
p53
Inhibits
p21/WAF1
Summary
• p53, a tumor suppressor, exists as a polymorph & this
may be responsible for cancer susceptibility across
differing ethnicities.
• At a molecular level, it switches cells from cytostasis
to apoptosis.
• p53 mediated apoptosis occurs via a number of
molecular processes: transcriptional upregulation of
Bax, hence a change in Bcl2/Bax ratio; of PUMA &
Peg3/PW1 & an increase in translocation of Bax to Mt.
• p53 cytostasis occurs through the p21/WAF1, Rb
pathway.
Summary
• p53 is being employed for ablation of tumors
using gene therapy.
• p53 interacts with a number of other proteins, such
as JNKs, etc and is involved in a variety of
pathways.
• P53 offers a profound insight into the working of
cancer cells & an opportunity for future
therapeutic treatments.
Me! Running against time to finish the presentation!