Functional characterization of MCAK/Kif2C cancer mutations using high-throughput microscopic analysis.

ABSTRACT: The microtubule (MT)-depolymerizing activity of MCAK/Kif2C can be quantified by expressing the motor in cultured cells and measuring tubulin fluorescence levels after enough hours have passed to allow tubulin autoregulation to proceed. This method allows us to score the impact of point mutations within the motor domain. We found that, despite their distinctly different activities, many mutations that impact transport kinesins also impair MCAK/Kif2C's depolymerizing activity. We improved our workflow using CellProfiler to significantly speed up the imaging and analysis of transfected cells. This allowed us to rapidly interrogate a number of MCAK/Kif2C motor domain mutations documented in the cancer database cBioPortal. We found that a large proportion of these mutations adversely impact the

SUBMITTER: Wagenbach M 

PROVIDER: S-EPMC7202071 | biostudies-literature | 2020 Mar

REPOSITORIES: biostudies-literature