Interplay between BRCA1 and RHAMM Regulates Epithelial Apicobasal Polarization and May Influence Risk of Breast Cancer
Figure 6
RHAMM depletion alters TPX2 localization and AURKA activity.
(A) Depletion of RHAMM, but not BRCA1, results in re-localization of TPX2 from the nucleus to the nuclear envelope and cytoplasm (arrows). With RHAMM depletion, microtubule organization is less focused and radial. Scale bar represents 20 µm. (B) RHAMM depletion alters AURKA-TPX2 association. In triplicate experiments, MCF10A were untreated or depleted of BRCA1 or RHAMM, and lysates were immunoprecipitated with AURKA, TPX2, or control IgG antibodies. Compared to untreated or BRCA1-depleted samples, RHAMM depletion resulted in an increase of TPX2 co-precipitated with AURKA. Short and long Western blot exposures are shown. (C) RHAMM depletion alters AURKA activity. Immunoprecipitation beads from triplicate experiments were analyzed for kinase activity using luminescent detection of ATP. Luminescence values were normalized to those obtained for beads precipitated with control IgG. Beads from untreated lysates precipitated with AURKA but not TPX2 antibodies demonstrated modest kinase activity. Depletion of RHAMM led to a significant increase in kinase activity with both AURKA and TPX2 precipitation (asterisks indicate one-sided t test p<0.05). Graph shows means and standard errors from triplicate experiments.