Open AccessSpatio‐temporal composition of the mitotic Chromosomal Passenger Complex detected using in situ proximity ligation assay
Author(s) -
Vuoriluoto Mariaana,
Laine Leena J.,
Saviranta Petri,
Pouwels Jeroen,
Kallio Marko J.
Publication year - 2011
Publication title -
molecular oncology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.332
H-Index - 88
eISSN - 1878-0261
pISSN - 1574-7891
DOI - 10.1016/j.molonc.2010.10.002
Subject(s) - aurora b kinase , mitosis , microbiology and biotechnology , centromere , anaphase , biology , plk1 , proximity ligation assay , spindle apparatus , kinetochore , polo like kinase , aurora kinase , cell division , cell cycle , cell , chromosome , genetics , receptor , gene
Cell division is orchestrated by a complex protein network that aims to maintenance of genomic stability. Visualisation of mitotic protein–protein associations in space and time has been limited due to the lack of proper biochemical and easy‐to‐use imaging tools. Here we report adaptation of the in situ proximity ligation assay (is‐PLA) to study mitotic protein interactions with spatio‐temporal resolution. We examined the composition of the Chromosomal Passenger Complex (CPC) at various mitotic phases and after chemical treatments using is‐PLA with antibodies against the core CPC subunits Aurora B, INCENP, Survivin and Borealin. Our results support the notion that the core CPC functions as a single structural unit at centromeres in early mitosis and at central spindle after the onset of anaphase. Treatment of cells with the Aurora B inhibitor ZM447439 diminished the is‐PLA signals at centromeres suggesting that Aurora B activity contributes to structural maintenance and/or proper subcellular localization of the core CPC. Is‐PLA‐based analysis of interaction between INCENP and Polo‐like kinase 1 (Plk1) proposes that the kinase co‐travels with CPC during late mitosis. The data illustrates both the strengths and limitations of the is‐PLA in the analysis of mitotic macromolecule associations at sub‐organelle level.