title: APC functions at the centrosome to stimulate microtubule growth.
abstract: The adenomatous polyposis coli (APC) tumor suppressor is multi-functional. APC is known to localize at the centrosome, and in mitotic cells contributes to formation of the mitotic spindle. To test whether APC contributes to nascent microtubule (MT) growth at interphase centrosomes, we employed MT regrowth assays in U2OS cells to measure MT assembly before and after nocodazole treatment and release. We showed that siRNA knockdown of full-length APC delayed both initial MT aster formation and MT elongation/regrowth. In contrast, APC-mutant SW480 cancer cells displayed a defect in MT regrowth that was unaffected by APC knockdown, but which was rescued by reconstitution of full-length APC. Our findings identify APC as a positive regulator of centrosome MT initial assembly and suggest that this process is disrupted by cancer mutations. We confirmed that full-length APC associates with the MT-nucleation factor γ-tubulin, and found that the APC cancer-truncated form (1-1309) also bound to γ-tubulin through APC amino acids 1-453. While binding to γ-tubulin may help target APC to the site of MT nucleation complexes, additional C-terminal sequences of APC are required to stimulate and stabilize MT growth.
authors.affiliation: Centre for Cancer Research, The Westmead Institute for Medical Research, The University of Sydney, Westmead, NSW 2145, Australia.
authors.forename: Christina
authors.initials: C
authors.lastname: Lui
authors.affiliation: Centre for Cancer Research, The Westmead Institute for Medical Research, The University of Sydney, Westmead, NSW 2145, Australia.
authors.forename: Cahora
authors.initials: C
authors.lastname: Ashton
authors.affiliation: Centre for Cancer Research, The Westmead Institute for Medical Research, The University of Sydney, Westmead, NSW 2145, Australia.
authors.forename: Manisha
authors.initials: M
authors.lastname: Sharma
authors.affiliation: Centre for Cancer Research, The Westmead Institute for Medical Research, The University of Sydney, Westmead, NSW 2145, Australia.
authors.forename: Mariana G
authors.initials: MG
authors.lastname: Brocardo
authors.affiliation: Centre for Cancer Research, The Westmead Institute for Medical Research, The University of Sydney, Westmead, NSW 2145, Australia. Electronic address: beric.henderson@sydney.edu.au.
authors.forename: Beric R
authors.initials: BR
authors.lastname: Henderson
mesh_terms: D025601:Adenomatous Polyposis Coli Protein / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism; D001665:Binding Sites; D045744:Cell Line, Tumor; D018385:Centrosome / Q000187:drug effects / Q000378:metabolism* / Q000648:ultrastructure; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism* / Q000648:ultrastructure; D005786:Gene Expression Regulation; D017930:Genes, Reporter; D005816:Genetic Complementation Test; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D007399:Interphase / Q000187:drug effects; D008870:Microtubules / Q000187:drug effects / Q000378:metabolism* / Q000648:ultrastructure; D008938:Mitosis / Q000187:drug effects; D009154:Mutation; D015739:Nocodazole / Q000494:pharmacology; D011485:Protein Binding; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction; D014404:Tubulin / Q000235:genetics / Q000378:metabolism*; D050257:Tubulin Modulators / Q000494:pharmacology
chemical_list: C555337:APC protein, human; D025601:Adenomatous Polyposis Coli Protein; D034741:RNA, Small Interfering; D014404:Tubulin; D050257:Tubulin Modulators; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; D015739:Nocodazole
keywords: APC; Centrosome; Gamma-tubulin; Microtubule elongation; Microtubule nucleation