1. Cytoplasmic Signaling Circuitry Programs Many of the Traits of Cancer
Chapter 6
Cytoplasmic Signaling Circuitry Programs
Many of the Traits of Cancer
5.9 -
6.2 ~ 6.9 -
Apr 3 & 12, 2007

2. 5.9 The Ras protein functions as a G protein
- The ras oncogene triggers many of the same
changes in cells which are transformed by
erbB (truncated EGF-R) or sis (PDGF-B).
- Could Ras be found somewhere downstream
of erbB and sis ?
- Do the signals emitted by EGF-R and PDGF-R
converge on some common molecule ?

3. growth factor receptor
Ras
GRB2:
growth factor receptor-
bound protein 2
SH2, SH3: Src homolog 2 or 3
Sos: son of sevenless
(guanosine diphosphate)
(guanosine triphosphate)

4. Regulation of Ras (a GTPase) activity
(mitogenic signals)
Guanine nucleotide
Exchange Factor
e.g., Sos, which catalyzes conversion of inactive GDP-bound Ras to the active GTP-bound form.
GTPase-
Activating
(or Accelerating)
Protein

5. Regulation of Ras (a GTPase) activity
(mitogenic signals)
Guanine nucleotide
Exchange Factor
e.g., Sos, which catalyzes conversion of inactive GDP-bound Ras to the active GTP-bound form.
GTPase-
Activating
(or Accelerating)
Protein
(e.g., H-Ras)

6. The structure of the Ras protein
Two most frequently
altered a.a. residues
found in human
oncoproteins are
glycine 12 and
glutamine 61, which
are closed associated
with the γ-phosphate
of GTP.
Figure The Biology of Cancer (© Garland Science 2007)

7. EGF/EGFR-mediated Ras activation

8. Two classes of GTPase switch proteins 1. trimeric (large) G proteins,
(guanine nucleotide-binding proteins) :
1. trimeric (large) G proteins,
which directly bind to and
activated by receptors,
e.g., Smoothened, Frizzled.

9. 2. monomeric (small) G proteins, such as Ras and various Ras-
like proteins. Ras doesn’t bind to receptors. It indirectly links
to receptors via adapter proteins (such as GRB2) and GEF
proteins (such as Sos).

10. 6.2 The Ras protein stands in the middle of a
complex signaling cascade
The story of
son of sevenless (sos)
the genetics of eye
development in Drosophila
In the absence of sevenless, the 7th cell in each ommatidium fail to form.
Figure 6.5 The Biology of Cancer (© Garland Science 2007)

11. 6.3 Tyrosine phosphorylation controls the
location and thereby the actions of many
cytoplasmic signaling proteins
Why is phosphorylation critical?
- protein relocalization.

12. Domain structure of Src protein
substrate recognition
- proline rich
(SH3 domain)
- p-tyrosine (SH2 domain)
catalytic SH1 domain
Figure 6.7a The Biology of Cancer (© Garland Science 2007)

13. Structure and function of SH2 domain
act as a modular plug
Figure 6.8a The Biology of Cancer (© Garland Science 2007)

14. Signal-transducing proteins attach to phosphorylated receptors
a phosphotyrosine phosphatase (PTP)
Figure 6.9 The Biology of Cancer (© Garland Science 2007)

15. Signaling molecules with SH2 and SH3 domains
Figure 6.10a The Biology of Cancer (© Garland Science 2007)

16. Molecular “ligand domains” and “receptor domains” in order to facilitate intermolecular interactions
Figure 6.10b The Biology of Cancer (© Garland Science 2007)

17. Table 6.2 The Biology of Cancer (© Garland Science 2007)

18. 6.4 SH2 groups explain how growth factors activate Ras and acquire signaling
Figure The Biology of Cancer (© Garland Science 2007)

19. Mammalian Ras proteins have been studied in great detail because mutants Ras proteins are associated with many types of human cancer. These mutant proteins, which bind but cannot hydrolyze GTP, are permanently in the “on” state and contribute to neoplastic transformation.
Most oncogenic, constitutively active Ras protein contain a mutation at position 12. Replacement of the normal glycine-12 with other amino acid blocks the functional binding of GAP, and in essence “lock” Ras in the active GTP-bound state.

20. Sidebar 6.3 The SH2 domain of Src has two alternative functions
inactive dephosphorylation of Y phosphorylation of Y416
SH2/SH3 bind to p-receptor exposure of the catalytic cleft
Figure The Biology of Cancer (© Garland Science 2007)

21. 6.5 A cascade of kinases forms one of three important
signaling pathways downstream of Ras
Figure The Biology of Cancer (© Garland Science 2007)

22. The Ras effector loop can bind Raf, PI3K and Ral-GEF
Figure The Biology of Cancer (© Garland Science 2007)

23. Ras → Raf → MAP (Mitogen-activated protein) kinase pathway
Raf, MEK and Erk1/2
are serine/threonine
kinases
phosphotyrosines attract signaling partners → relocalization of partners
phosphorylation of serines/threonines → shift in structure
→ functional activation
MEK: MAP/Erk kinase
Erk: extracellular signal-
regulated kinases 1 and 2
(Erk1/2)
Figure The Biology of Cancer (© Garland Science 2007)

24. Ras/Raf/MAP Kinase Pathway
dimer
Figure 6.10b The Biology of Cancer (© Garland Science 2007)

25. Activated MAPK induces gene transcription
transcription factor TCF:
ternary complex factor
SRE:
serum response element
transcription factor SRF:
serum response factor
(early response gene) → enter the cell cycle

26. The Ras-PI3 kinase (PI3K) pathway
(cytoplasm)
Figure The Biology of Cancer (© Garland Science 2007)

27. Generation of the 2nd messengers – DAG and IP3

28. PI3K converts PIP2 into PIP3
PI3K can bind to phosphotyrosine and Ras.
Figure 6.16b The Biology of Cancer (© Garland Science 2007)

29. Docking of PH domains of Akt/PKB to PIP3
PTEN:
phosphatase and
tensin homolog
PH domain: pleckstrin homology
Figure 6.19a The Biology of Cancer (© Garland Science 2007)

30. Table 6.3 The Biology of Cancer (© Garland Science 2007)

31. AKT/PKB enhances the growth of cells
Normal islet cells
Islet cells expressing constitutively active Akt/PKB. The cells are fourfold larger than normal.
Figure The Biology of Cancer (© Garland Science 2007)

32. Table 6.4 The Biology of Cancer (© Garland Science 2007)

33. Sidebar 6.4 Ras is the prototype of a large family of similar proteins
Small G proteins – 35 similarly structured proteins
e.g., Ras, Ral, Rac, Ran, Rho…
- All small G proteins operate like a binary switch,
using a GTP-GDP-GTP cycle to flip back and forth
between an on and an off state.
- Each small G protein has its own specialized guanine
nucleotide exchange factor (GEF) to activate it and
its own GTPase-activating proteins (GAP)

34. 6.7 A third Ras-regulated pathway acts through
Ral, a distinct cousin of Ras
(Ral guanine nucleotide exchange factors)
(Ral-A and Ral-B are Ras-like proteins)
(Rho family proteins)
(control of cytoskeleton)
play key roles in the motility that enables
cancer cells to invade and metastasize
Figure The Biology of Cancer (© Garland Science 2007)

35. 6.8 The Jak-STAT pathway allows signals to be transmitted
from the plasma membrane directly to the nucleus
cytokines
Figure The Biology of Cancer (© Garland Science 2007)

36. 6.9 Cell adhesion receptors emit signals that converge
with those released by growth factor receptors
Figure 6.24a The Biology of Cancer (© Garland Science 2007)

37. Figure 6.24b The Biology of Cancer (© Garland Science 2007)