Recruitment of adenomatous polyposis coli and beta-catenin to axin-puncta.
The adenomatous polyposis coli (APC) tumour suppressor is a multifunctional protein involved in the regulation of Wnt signalling and cytoskeletal dynamics. Little is known about how APC controls these disparate functions. In this study, we have used APC- and axin-fluorescent fusion proteins to examine the interactions between these proteins and show that the functionally distinct populations of APC are also spatially separate. Axin-RFP forms cytoplasmic punctate structures, similar to endogenous axin puncta. Axin-RFP recruits beta-catenin destruction complex proteins, including APC, beta-catenin, glycogen synthase kinase-3-beta (GSK3-beta) and casein kinase-1-alpha (CK1-alpha). Recruitment into axin-RFP puncta sequesters APC from clusters at cell extensions and this prevents its microtubule-associated functions. The interaction between APC-GFP and axin-RFP within the cytoplasmic puncta is direct and dramatically alters the dynamic properties of APC-GFP. However, recruitment of APC to axin puncta is not absolutely required for beta-catenin degradation. Instead, formation of axin puncta, mediated by the DIX domain, is required for beta-catenin degradation. An axinDeltaDIX mutant did not form puncta, but still mediated recruitment of destruction complex proteins and phosphorylation of beta-catenin. We conclude that there are distinct pools of APC and that the formation of axin puncta, rather than the axin/APC complex, is essential for beta-catenin destruction.
Faux
M C
MC
Ludwig Institute for Cancer Research, Melbourne Tumour Biology Branch, Parkville, Victoria, Australia. maree.faux@ludwig.edu.au
Coates
J L
JL
Catimel
B
B
Cody
S
S
Clayton
A H A
AH
Layton
M J
MJ
Burgess
A W
AW
D025601:Adenomatous Polyposis Coli Protein / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D060466:Axin Protein; D002460:Cell Line; D003593:Cytoplasm / Q000378:metabolism / Q000648:ultrastructure; D004285:Dogs; D031541:Fluorescence Resonance Energy Transfer; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D051379:Mice; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012097:Repressor Proteins / Q000235:genetics / Q000378:metabolism*; D051176:beta Catenin / Q000378:metabolism*
D025601:Adenomatous Polyposis Coli Protein; D060466:Axin Protein; D011993:Recombinant Fusion Proteins; D012097:Repressor Proteins; D051176:beta Catenin; D049452:Green Fluorescent Proteins