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First published online 11 November 2008
doi: 10.1242/jcs.038414


Journal of Cell Science 121, 3971-3980 (2008)
Published by The Company of Biologists 2008
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Research Article

Two distinct regions of Mto1 are required for normal microtubule nucleation and efficient association with the {gamma}-tubulin complex in vivo

Itaru Samejima, Victoria J. Miller, Lynda M. Groocock and Kenneth E. Sawin*

Wellcome Trust Centre for Cell Biology, University of Edinburgh, Swann Building, Mayfield Road, Edinburgh EH9 3JR, UK

* Author for correspondence (e-mail: ken.sawin{at}ed.ac.uk)

Accepted 8 September 2008

Cytoplasmic microtubule nucleation in the fission yeast Schizosaccharomyces pombe involves the interacting proteins Mto1 and Mto2, which are thought to recruit the {gamma}-tubulin complex ({gamma}-TuC) to prospective microtubule organizing centres. Mto1 contains a short amino-terminal region (CM1) that is conserved in higher eukaryotic proteins implicated in microtubule organization, centrosome function and/or brain development. Here we show that mutations in the Mto1 CM1 region generate mutant proteins that are functionally null for cytoplasmic microtubule nucleation and interaction with the {gamma}-TuC (phenocopying mto1{Delta}), even though the Mto1-mutant proteins localize normally in cells and can bind Mto2. Interestingly, the CM1 region is not sufficient for efficient interaction with the {gamma}-TuC. Mutation within a different region of Mto1, outside CM1, abrogates Mto2 binding and also impairs cytoplasmic microtubule nucleation and Mto1 association with the {gamma}-TuC. However, this mutation allows limited microtubule nucleation in vivo, phenocopying mto2{Delta} rather than mto1{Delta}. Further experiments suggest that Mto1 and Mto2 form a complex (Mto1/2 complex) independent of the {gamma}-TuC and that Mto1 and Mto2 can each associate with the {gamma}-TuC in the absence of the other, albeit extremely weakly compared to when both Mto1 and Mto2 are present. We propose that Mto2 acts cooperatively with Mto1 to promote association of the Mto1/2 complex with the {gamma}-TuC.

Key words: Fission yeast, {gamma}-Tubulin complex, Centrosomin-related proteins


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