Other examples of oncogenic retroviruses

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Other examples of oncogenic retroviruses
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The v-myc oncogene has a cellular origin
Burkitts lymphoma/plamacytoma
neuroblastoma/lung CA
Amplification
Translocation
c-myc locus
also N-myc, L-myc
Roussel et a. 1979. Nature 281, 452 others
Retroviral transduction
Retroviral insertion
lymphomas/carcinomas
lymphomas
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Myc Gene Rearrangements in Cancer
Burkitts lymphoma/plamacytoma
neuroblastoma/lung CA
Amplification
Translocation
c-myc locus
also N-myc, L-myc
Retroviral transduction
Retroviral insertion
lymphomas/carcinomas
lymphomas
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Ras
Oncogenes co-operate in Malignant cell
transformation
in vitro
Ras Myc
Land et al. 1983 Nature 304, 596.
Myc
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and in vivo
Sinn et al. Cell. 198749465. MMTV/v-Ha-ras and
MMTV/c-myc genes in transgenic mice
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Myc is an immediate-early serum-response geneand
is continously expressed in proliferating cells
c-myc
c-fos
FCS
-FCS
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Growth factors Cytokines Cell adhesion
... Contact-inhibition TGFb Differentiation
...
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Growth factors Cytokines Cell adhesion
... Contact-inhibition TGFb Differentiation
...
Cellular Transformation
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Growth factors Cytokines Cell adhesion
... Contact-inhibition TGFb Differentiation
...
Normal Function ?
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Myc is an intracellular sensor and transducer of
extracellular stimuli
Growth factors Cytokines Cell adhesion
... Contact-inhibition TGFb Differentiation
...
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Functional capabilities tumor cells have to aquire
Hanahan and Weinberg 2000 The hallmarks of
cancer. Cell 100, 57-70.
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Functional capabilities tumor cells have to aquire
Hanahan and Weinberg 2000 The hallmarks of
cancer. Cell 100, 57-70.


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Structure of Myc lt 1987
An anonymous nuclear protein of 439 a.a.
439 a.a.
320-328 Nuclear Localization Sequence Dang
and Lee 1988. Mol Cell Biol. 8, 4048
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Structure of Myc lt 1987
An anonymous nuclear protein of 439 a.a.
439 a.a.
320-328 Nuclear Localization Sequence Dang
and Lee 1988. Mol Cell Biol. 8, 4048
studied by structure-function assays in cells
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Structure of Myc 1987
320-328 NLS
Stone et al. 1987. Mol Cell Biol. 7, 1697
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1989 recognition of the bHLH-LZ domain
bHLH (Murre at al. 1989. Cell 11, 537)
Leucine zipper
bHLH-LZ domains are highly modular and
versatile- Functional when fused either N- or
C-terminal of epitopes or bulky proteins.-
Fusion proteins function both in vitro and in
vivo (e.g. yeast two-hybrid).- Specificity is
usually retained in indicidual domains.-
Depending on proteins, HLH alone, LZ alone or
HLHLZ sufficient for dimerization in solution.
- Myc did NOT form homodimers and did not bind
other known HLH proteins.
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Structure of Myc 1989
DIMERIZATION PARTNER ? DNA BINDING ?
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Transcription factors are modular proteins
Transcription through GAL UAS
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Structure of Myc 1990
DIMERIZATION PARTNER ? DNA BINDING ?
TRANSACTIVATION DOMAIN
Kato GJ, Barrett J, Villa-Garcia M, Dang CV. Mol
Cell Biol. 1990 Nov10(11)5914-20. An
amino-terminal c-myc domain required for
neoplastic transformation activates transcription.
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106
Bacteriophage plaques
Bacterial lawn
Blackwood EM, Eisenman RN. Science. 1991 Mar
8251(4998)1211-7. Max a helix-loop-helix
zipper protein that forms a sequence-specific
DNA-binding complex with Myc.
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Structure of Myc, Max 1991
TRANSCRIPTIONAL ACTIVATION
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Structure of Myc, Max 1991
TRANSCRIPTIONAL ACTIVATION
1992 The Myc/Max dimer functions as a
transcription factor in vivo Kretzner L,
Blackwood EM, Eisenman RN. Nature 359,
426-9. Myc and Max proteins possess distinct
transcriptional activities. Amati B, Dalton S,
Brooks MW, Littlewood TD, Evan GI, Land H.
Nature 359, 423-6. Transcriptional activation by
the human c-Myc oncoprotein in yeast requires
interaction with Max.
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Structure of Myc, Max 1991
TRANSCRIPTIONAL ACTIVATION
1992 The Myc/Max dimer functions as a
transcription factor in vivo

but does it
transform cells?
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O'Shea EK, Klemm JD, Kim PS, Alber T. Science.
1991 Oct 25254(5031)539-44. X-ray structure of
the GCN4 leucine zipper, a two-stranded, parallel
coiled coil.
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Point mutations in the Myc and Max Leucine
Zippers are sufficient to eliminate dimerization
in vivo and abrogate oncogenic activity
Dimerization
yes
no
no
yes
Amati et al. 1993 Cell 72, 233
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Co-immunoprecipitation of in vitro-translated
proteins
Myc wt
Myc EG
T
T
Y
Y
Max wt
Y
Y
T
T
Y
Y
Max EG
Y
Y
Amati et al. 1993 Cell 72, 233
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Amati et al. 1993 Cell 72, 233
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Oncogenic activity of Myc depends upon
dimerization with Max, DNA binding and
transcriptional activation
wild-type Myc
dimerization mutant
complementary Myc/Max mutants
DNA-binding mutant
transactivation mutant
Myc overexpressed by transfection
Endogenous cellular Max
Oncogenic activity
-

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THUS Interfering with Myc/Max dimerization and
DNA binding eliminates Myc function
IS THIS ACHIEVABLE IN HUMAN TUMORS?
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Structure of Myc, Max, Mad 1993
TRANSCRIPTIONAL ACTIVATION
Ayer et al. 1993 Cell 72, 211
Zervos et al 1993 Cell 72, 223
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Structure of Myc, Max, Mad 1993
TRANSCRIPTIONAL ACTIVATION
TRANSCRIPTIONAL REPRESSION
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The Myc - Max - Mad network
GROWTH STIMULI
ARREST DIFF. SIGNALS
Mad / Mxi
Max
Max
Max
Max
Myc
TRANSCRIPTIONAL ACTIVATION
TRANSCRIPTIONAL REPRESSION
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The Max Network now
Mnt
Normal functions vs. Roles in Neoplasia
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Crystal structure of the DNA-bound Max bHLH-LZ
domain homodimer
C
L-Zipper
Dimerization
Helix II
Loop
Helix I
DNA-binding
basic
N
Ferre-dAmare et al. 1993 Nature 363, 38-45
The Myc/Max heterodimer has a similar
structure (S.K. Nair S.K. Burley 2003. Cell).
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C
C
C
N
N
N
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Myc can function as an activator or repressor of
different target genes
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c-Myc regulates genes that are involved in many
different aspect of the control of cell behavior
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c-Myc regulates genes that are involved in many
different aspect of the control of cell behavior
Myc binds THOUSANDS of genes !!!
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Functional capabilities tumor cells have to aquire
Hanahan and Weinberg 2000 The hallmarks of
cancer. Cell 100, 57-70.


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Myc



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A little bit of pRB history
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pRB and the cell cycle
M
G2
G1
S
Buchkovich et al. 1989, Cell 58 DeCaprio et al.
1989, Cell 58 Chen et al. 1989, Cell 58 Goodrich
et al. 1991, Cell 67
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Model for pRB function
Activated
Kovesdi et al. 1986, Cell 45 Chellappan et al.
1991, Cell 65 Bagchi et al. 1991, Cell
65 Chittenden et al. 1991, Cell 65 Bandara et al.
1991, Nature 351
Adenovirus
Cyclin/CDK
E2F
E2F
TTTCGCGC
GCGCGAAA
Cloning of E2F Helin et al. 1992, Cell 70 Kaelin
et al. 1992, Cell 70
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DNA tumor viruses are smart
Adenovirus
E1B
Human Papilloma Virus
E6
Simian Virus 40
pRB
T
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Regulation of pRB activity
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E2F and Rb families
Activators, repressors Cell cycle and apoptosis
E2F1
E2F3
pRB
E2F2
pRB
p130
E2F4
Repressors Cell cycle, differentiation
E2F5
p107
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1-5
1,2
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• Alevizopoulos K, Vlach J, Hennecke S, Amati B.,
  1997
• EMBO J. 16, 5322.
• Cyclin E and c-Myc promote cell proliferation in
  the presence of p16INK4a
• and of hypophosphorylated retinoblastoma family
  proteins.

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