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Human ASPM participates in spindle organisation, spindle orientation and cytokinesis

Higgins, Julie and Midgley, Carol and Bergh, Anna-Maria and Bell, Sandra M. and Askham, Jonathan M. and Roberts, Emma and Binns, Ruth K. and Sharif, Saghira M. and Bennett, Christopher and Glover, David M. and Woods, C. Geoffrey and Morrison, Ewan E. and Bond, Jacquelyn (2010) Human ASPM participates in spindle organisation, spindle orientation and cytokinesis. BMC Cell Biology, 11 . Art. No. 85. ISSN 1471-2121. PMCID PMC2988714. https://resolver.caltech.edu/CaltechAUTHORS:20201005-153617371

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PDF (Additional file 3: . HeLa, COS-7, U2OS, SH-SY5Y and HDF cells were fixed and stained with the N-terminal ASPM antibody 216-1 (green), anti-α-tubulin (red) and DAPI (blue) to identify nuclei. Scale bar = 10 μm) - Supplemental Material
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Abstract

Background. Mutations in the A bnormal Sp indle M icrocephaly related gene (ASPM) are the commonest cause of autosomal recessive primary microcephaly (MCPH) a disorder characterised by a small brain and associated mental retardation. ASPM encodes a mitotic spindle pole associated protein. It is suggested that the MCPH phenotype arises from proliferation defects in neural progenitor cells (NPC). Results. We show that ASPM is a microtubule minus end-associated protein that is recruited in a microtubule-dependent manner to the pericentriolar matrix (PCM) at the spindle poles during mitosis. ASPM siRNA reduces ASPM protein at the spindle poles in cultured U2OS cells and severely perturbs a number of aspects of mitosis, including the orientation of the mitotic spindle, the main determinant of developmental asymmetrical cell division. The majority of ASPM depleted mitotic cells fail to complete cytokinesis. In MCPH patient fibroblasts we show that a pathogenic ASPM splice site mutation results in the expression of a novel variant protein lacking a tripeptide motif, a minimal alteration that correlates with a dramatic decrease in ASPM spindle pole localisation. Moreover, expression of dominant-negative ASPM C-terminal fragments cause severe spindle assembly defects and cytokinesis failure in cultured cells. Conclusions. These observations indicate that ASPM participates in spindle organisation, spindle positioning and cytokinesis in all dividing cells and that the extreme C-terminus of the protein is required for ASPM localisation and function. Our data supports the hypothesis that the MCPH phenotype caused by ASPM mutation is a consequence of mitotic aberrations during neurogenesis. We propose the effects of ASPM mutation are tolerated in somatic cells but have profound consequences for the symmetrical division of NPCs, due to the unusual morphology of these cells. This antagonises the early expansion of the progenitor pool that underpins cortical neurogenesis, causing the MCPH phenotype.


Item Type:Article
Related URLs:
URLURL TypeDescription
https://doi.org/10.1186/1471-2121-11-85DOIArticle
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2988714/PubMed CentralArticle
ORCID:
AuthorORCID
Glover, David M.0000-0003-0956-0103
Additional Information:© 2010 Higgins et al; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Received: 10 February 2010. Accepted: 2 November 2010. Published: 2 November 2010. Normal control and MCPH Human dermal fibroblast cultures were kindly prepared by the Yorkshire Regional Cytogenetics Laboratory, St James's University Hospital, Leeds, UK. We acknowledge the input from Adam Davison and the LIMM Core Flow Cytometry Facility. Statistical guidance was provided by Mohamed Saleh, LIMM. This research was supported by The Wellcome Trust (JB, JH and CGW). DMG and A-MB were funded by Cancer Research UK, and SMB and EEM by Yorkshire Cancer Research. CAM was funded by the Open University. SMS and CB were funded by the Leeds Teaching Hospitals NHS Trust. Authors' contributions: Project conceived and experiments designed by JB, EEM, CGW, CAM, SMB and DMG. Experiments carried out by JB, JH, CAM, A-MB, RKB, SMB, and JMA. Patients diagnosed, recruited and tissue biopsies obtained by CGW, SMS and CB. Manuscript prepared by JB, EEM, CGW, CAM and DMG. All authors have read and approved the final manuscript. Julie Higgins, Carol Midgley contributed equally to this work.
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Funding AgencyGrant Number
Wellcome TrustUNSPECIFIED
Cancer Research UKUNSPECIFIED
Yorkshire Cancer ResearchUNSPECIFIED
Open UniversityUNSPECIFIED
Leeds Teaching Hospitals NHS TrustUNSPECIFIED
PubMed Central ID:PMC2988714
Record Number:CaltechAUTHORS:20201005-153617371
Persistent URL:https://resolver.caltech.edu/CaltechAUTHORS:20201005-153617371
Official Citation:Higgins, J., Midgley, C., Bergh, A. et al. Human ASPM participates in spindle organisation, spindle orientation and cytokinesis. BMC Cell Biol 11, 85 (2010). https://doi.org/10.1186/1471-2121-11-85
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:105827
Collection:CaltechAUTHORS
Deposited By: George Porter
Deposited On:06 Oct 2020 14:09
Last Modified:06 Oct 2020 14:09

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