1. Cytoskeleton Journal Club June 13, 2012
Presented by Suk Namgoong

2. Adenomanous Polypolis Coli (APC)
• Main component of Wnt Signaling Pathway
• In the absence of bounded Wnt on Wnt receptor, it forms ‘destruction
complex’ for β-catanin,
- mediates proteolytic degradation of β-catenin
• By the bound of wnt into wnt receptor Frizzled, APC release β-catenin and β-
catenin was translocated into nucleus and ctivated various Wnt target genes
(Development, Morphogenesis, Cancers..)
• Tumor Suppressor Gene
- Mutations on APC gene cause a hereditary cancer syndrome called
Familiar Adenomanous Polypolis (FAP)
- Loss of APC functions by mutations has been found frequently in colorectal
cancers

3. APC directly nucleates actin assembly via C-terminal
Basic Domain
© 2010 Okada et al. Okada K et al. J Cell Biol 2010;189:1087-1096

4. ANS : Actin nucleation sequences
No Sequence Homology with known actin binding domain/motifs
It is dimer in solutio

5. mDia1 and APC-B synergize to promote actin assembly in
the combined presence of profilin and capping protein.
Okada K et al. J Cell Biol 2010;189:1087-1096
© 2010 Okada et al.

6. Two different binding mode of Actin nucleators
Arp2/3 complex stay on the ‘pointed end’ (slow growing end) of actin filaments
Formins remains processively bound on ‘barbed end’ (fast growing end)
How about APC? From which side of actin filaments it initiate actin nucleation?

7. Synergistic actin nucleations by APC and mDia1
(Cappuccino) Yeast Bnr1 – bud6
During the nucleation, Formins and other nucleation
factors interact each other and synergize nucleations.
But what will happen after nucleation?
Do formins and other nucleators dissociate after
nucleation?
or formins carry all other stuffs along with elongation?
(not likely, but we don‟t know yet)

8. Fluorescence Probe Labeling with SNAP-tag (O6-alkylguanine-DNA alkyltransferase)
54
9
SNAP FH1 FH2
SNAP APC (Basic)
64
7
Oregon-Green
Labeled Actin
64
7
PEG A
Laser excitation

9. Immobilized SNAP-APC Stepwise Photobleaching Addition of LatB
Can generate actin filaments Inhibit new filament generation
(APC is stay bound on pointed end)

10. Bulk Pyrene Actin Assay Stepwise photobleaching for the determination of
Oligomeric state in single molecules

12. Complex Formations of mDia1, APC-C, OregonGreen-Actin (LatB treated)

15. Interaction via C-
terminal region of
mDia1

17. Namgoong et al., Nature Struct Mol. Biol. 2011
I feel De ja vu….. 

18. APC VopL
APC stays bound on the pointed
end of actin Filaments VopL is dissociated from the filament end
and „spit out‟ new filaments continuously
vs
“Rocket Launcher” “Machine Gun”