Microtubule association of TRIM3 revealed by differential extraction proteomics

Glover, H. L. , Mendes, M., Gomes-Neto, J., Rusilowicz-Jones, E. V., Rigden, D. J., Dittmar, G., Urbé, S. and Clague, M. J. (2024) Microtubule association of TRIM3 revealed by differential extraction proteomics. Journal of Cell Science, 137(2), jcs261522. (doi: 10.1242/jcs.261522) (PMID:38149663)

Abstract

The microtubule network is formed from polymerised tubulin subunits and associating proteins, which govern microtubule dynamics and a diverse array of functions. To identify novel microtubule-binding proteins, we have developed an unbiased biochemical assay, which relies on the selective extraction of cytosolic proteins from U2OS cells, while leaving behind the microtubule network. Candidate proteins are linked to microtubules by their sensitivities to the depolymerising drug nocodazole or the microtubule-stabilising drug taxol, which is quantitated by mass spectrometry. Our approach is benchmarked by co-segregation of tubulin and previously established microtubule-binding proteins. We then identify several novel candidate microtubule-binding proteins, from which we have selected the ubiquitin E3 ligase tripartite motif-containing protein 3 (TRIM3) for further characterisation. We map TRIM3 microtubule binding to its C-terminal NHL-repeat region. We show that TRIM3 is required for the accumulation of acetylated tubulin, following treatment with taxol. Furthermore, loss of TRIM3 partially recapitulates the reduction in nocodazole-resistant microtubules characteristic of α-tubulin acetyltransferase 1 (ATAT1) depletion. These results can be explained by a decrease in ATAT1 following depletion of TRIM3 that is independent of transcription.

• Item Type: Articles
• Additional Information: Funding: H.G. and J.G.N. are recipients of PhD studentships from the Wellcome Trust and North West Cancer Research, respectively. M.C. is a recipient of a Royal Society Industry Fellowship INF\R2\212031. Open Access funding provided by University of Liverpool. Deposited in PMC for immediate release.
• Status: Published
• Refereed: Yes
• Glasgow Author(s) Enlighten ID: Glover, Ms Hannah

Creator Roles

Glover, H.: Conceptualization, Methodology, Formal analysis, Investigation, Writing – original draft, Writing – review and editing, Visualization

• Authors: Glover, H. L., Mendes, M., Gomes-Neto, J., Rusilowicz-Jones, E. V., Rigden, D. J., Dittmar, G., Urbé, S., and Clague, M. J.
• College/School: College of Medical Veterinary and Life Sciences > School of Cancer Sciences
• Journal Name: Journal of Cell Science
• Publisher: Company of Biologists
• ISSN: 0021-9533
• ISSN (Online): 1477-9137
• Copyright Holders: Copyright: © 2024. Published by The Company of Biologists Ltd.
• First Published: First published in Journal of Cell Science 137(2): jcs261522
• Publisher Policy: Reproduced under a Creative Commons licence