Regulation and function of the interaction between the APC tumour suppressor protein and EB1

The interaction between the adenomatous polyposis coli (A PC) tumour suppre ssor and the microtubule-associated protein EB1 was examined, Immunoprecipi tation suggested that APC and EB1 were not associated in cultures of HCT116 cells arrested in mitosis, The C-terminal 170 amino acids of APC, purified as a bacterial fusion protein, precipitated EB1 from cell extracts, signif icantly refining the location of the EB1 interaction domain in APC, In vitr o phosphorylation of this fusion protein by either protein kinase A or p34( cdc2) reduced its ability to bind to EB1, Expression of GFP fusions to C-te rminal APC sequences Lacking or including the APC basic domain but encompas sing the EB1 binding region in SW480 cells revealed a microtubule tip assoc iation which co-localized with that of EB1, Expression of the basic domain alone revealed a non-specific microtubule localization. In vitro interactio n studies confirmed that the APC basic domain did not contribute to EB1 bin ding. These findings strongly suggest that the interaction between APC and EB1 targets APC to microtubule tips, and that the interaction between the t wo proteins is down-regulated during mitosis by the previously described mi totic phosphorylation of APC.