Journal of Biological Chemistry
Volume 285, Issue 34, 20 August 2010, Pages 26461-26474
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Cell Biology
RHAMM Promotes Interphase Microtubule Instability and Mitotic Spindle Integrity through MEK1/ERK1/2 Activity*

https://doi.org/10.1074/jbc.M110.121491Get rights and content
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An oncogenic form of RHAMM (receptor for hyaluronan-mediated motility, mouse, amino acids 163–794 termed RHAMMΔ163) is a cell surface hyaluronan receptor and mitotic spindle protein that is highly expressed in aggressive human cancers. Its regulation of mitotic spindle integrity is thought to contribute to tumor progression, but the molecular mechanisms underlying this function have not previously been defined. Here, we report that intracellular RHAMMΔ163 modifies the stability of interphase and mitotic spindle microtubules through ERK1/2 activity. RHAMM−/− mouse embryonic fibroblasts exhibit strongly acetylated interphase microtubules, multi-pole mitotic spindles, aberrant chromosome segregation, and inappropriate cytokinesis during mitosis. These defects are rescued by either expression of RHAMM or mutant active MEK1. Mutational analyses show that RHAMMΔ163 binds to α- and β-tubulin protein via a carboxyl-terminal leucine zipper, but in vitro analyses indicate this interaction does not directly contribute to tubulin polymerization/stability. Co-immunoprecipitation and pulldown assays reveal complexes of RHAMMΔ163, ERK1/2-MEK1, and α- and β-tubulin and demonstrate direct binding of RHAMMΔ163 to ERK1 via a D-site motif. In vitro kinase analyses, expression of mutant RHAMMΔ163 defective in ERK1 binding in mouse embryonic fibroblasts, and blocking MEK1 activity collectively confirm that the effect of RHAMMΔ163 on interphase and mitotic spindle microtubules is mediated by ERK1/2 activity. Our results suggest a model wherein intracellular RHAMMΔ163 functions as an adaptor protein to control microtubule polymerization during interphase and mitosis as a result of localizing ERK1/2-MEK1 complexes to their tubulin-associated substrates.

ERK
Fibroblast
Microtubules
Mitotic Spindle
Oncogene
Hyaluronan Receptor
Microtubule Dynamics
Unconventional Proteins

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*

This work was supported, in whole or in part, by National Institutes of Health Grant 5R0119092 from NCI (to J. B. M. and E. A. T.). This work was also supported by a grant from the Cancer Research Society, Montreal, Canada (to E. A. T.), and by the Canadian Breast Cancer Society (partial salary to E. A. T.).

The on-line version of this article (available at http://www.jbc.org) contains supplemental Figs. 1–6.

1

Supported by a Translational Breast Cancer Studentship from the London Regional Cancer Program.