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Microtubule association of EML proteins and the EML4-ALK variant 3 oncoprotein require an N-terminal trimerization domain.

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journal contribution
posted on 11.03.2015, 12:18 by Mark W. Richards, Laura O'Regan, D. Roth, Jessica M. Montgomery, A. Straube, Andrew M. Fry, R. Bayliss
Proteins of the echinoderm microtubule associated protein-like (EML) family contribute to formation of the mitotic spindle and interphase microtubule (MT) network. EML1-4 consist of WD40 repeats and an N-terminal region containing a putative coiled-coil. Recurrent gene rearrangements in non-small cell lung cancer (NSCLC) fuse EML4 to anaplastic lymphoma kinase (ALK) causing expression of several oncogenic fusion variants. The fusions have constitutive ALK activity due to self-association through the EML4 coiled-coil. We have determined crystal structures of the coiled-coils from EML2 and EML4, which describe the structural basis of both EML self-association and oncogenic EML4-ALK activation. The structures reveal a trimeric oligomerization state directed by a conserved pattern of hydrophobic residues and salt bridges. We show that the trimerization domain (TD) of EML1 is necessary and sufficient for self-association. The TD is also essential for MT binding, however this property requires an adjacent basic region. These observations prompted us to investigate MT association of EML4-ALK and EML1-ABL1 fusions in which variable portions of the EML component are present. Uniquely, EML4-ALK variant 3, which includes the TD and basic region of EML4 but none of the WD40 repeats, was localized to MTs, both when expressed recombinantly and in a patient-derived NSCLC cell line (H2228). This raises the question of whether the mislocalization of ALK activity to MTs might influence downstream signalling and malignant properties of cells. Furthermore, the structure of EML4 TD may enable the development of protein-protein interaction inhibitors targeting the trimerization interface, providing a possible avenue towards therapeutic intervention in EML4-ALK NSCLC.

Funding

This work was supported by a Royal Society University Research Fellowship and Cancer Research UK programme funding to R.B. [C24461/A12772], a Marie Curie Cancer Care programme grant and Lister Institute Research Prize to A.S. and funding from Worldwide Cancer Research and the Wellcome Trust to A.M.F.

History

Citation

Biochemical Journal, 2015

Author affiliation

/Organisation/COLLEGE OF MEDICINE, BIOLOGICAL SCIENCES AND PSYCHOLOGY/School of Biological Sciences/Department of Biochemistry

Version

AM (Accepted Manuscript)

Published in

Biochemical Journal

Publisher

Portland Press for Biochemical Society

issn

0264-6021

eissn

1470-8728

Copyright date

2015

Available date

05/09/2015

Publisher version

http://www.biochemj.org/bj/imps/abs/BJ20150039.htm

Notes

We thank the beamline support staff at DLS I02 and I04 for assistance with data collection and the Centre for Core Biotechnology Services (University of Leicester) for imaging support.

Language

en

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