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Mitotic spindle assembly and γ-tubulin localisation depend on the integral nuclear membrane protein, Samp1
Stockholm University, Faculty of Science, Department of Neurochemistry.ORCID iD: 0000-0001-5835-503X
Stockholm University, Faculty of Science, Department of Neurochemistry.
Stockholm University, Faculty of Science, Department of Neurochemistry.
Stockholm University, Faculty of Science, Department of Neurochemistry.
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2018 (English)In: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 131, no 8, article id jcs211664Article in journal (Refereed) Published
Abstract [en]

We have investigated a possible role of the inner nuclear membrane protein, Samp1, in the mitotic machinery. Live cell imaging showed that Samp1aYFP distributed as filamentous structures in the mitotic spindle, partially co-localising with ß-tubulin. Samp1 depletion resulted in an increased frequency of cells with signs of chromosomal mis-segregation and prolonged metaphase, indicating problems with spindle assembly and/or chromosomal alignment. Consistently, mitotic spindles in Samp1 depleted cells contained significantly lower levels of ß-tubulin and γ-tubulin, phenotypes which were rescued by overexpression of Samp1aYFP. We found that Samp1 can bind directly to γ-tubulin and that Samp1 co-precipitated with γ-tubulin and HAUS6 of the Augmin complex in live cells. The levels of Haus6, in the mitotic spindle also decreased after Samp1 depletion. We show that Samp1 is involved in the recruitment of Haus6 and γ-tubulin to the mitotic spindle. Samp1 is the first inner nuclear membrane protein shown to have a function in mitotic spindle assembly.

Place, publisher, year, edition, pages
2018. Vol. 131, no 8, article id jcs211664
Keywords [en]
Samp1, nuclear membrane, mitotic spindle, gamma tubulin, Augmin, Cancer
National Category
Cell Biology
Research subject
Neurochemistry with Molecular Neurobiology
Identifiers
URN: urn:nbn:se:su:diva-154091DOI: 10.1242/jcs.211664ISI: 000440536400010OAI: oai:DiVA.org:su-154091DiVA, id: diva2:1190455
Funder
Swedish Research Council, 621-2010-448Swedish Cancer Society, 110590Stiftelsen Olle Engkvist ByggmästareAvailable from: 2018-03-14 Created: 2018-03-14 Last updated: 2018-08-20Bibliographically approved
In thesis
1. Characterization of the inner nuclear membrane protein Samp1, during interphase and mitosis
Open this publication in new window or tab >>Characterization of the inner nuclear membrane protein Samp1, during interphase and mitosis
2018 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

The nucleus, a hallmark in eukaryotic cells, contains the genome separating it from molecules in the cytoplasm. The nucleus is surrounded by a nuclear envelope consisting of two concentric membranes, the outer nuclear membrane and the inner nuclear membrane, the nuclear lamina and nuclear pore complexes. The cytoskeleton is physically connected with the nucleoskeleton by the LINC complexes, spanning the nuclear envelope. In this way, the cell surface is linked directly to chromatin. There are hundreds of unique inner nuclear membrane proteins, but today we only know the functions of a handful. The best characterized inner nuclear membrane proteins are involved in chromatin organization and gene regulation.

This thesis focuses on Samp1, an integral membrane protein that localizes to the inner nuclear membrane during interphase. During mitosis, a fraction localizes to the mitotic spindle, which is responsible for accurate segregation of chromosomes.

It is difficult to investigate inner nuclear membrane protein-protein interactions, because transmembrane proteins are often associated with the “hard-to-solubilize” nuclear lamina. MCLIP was developed as a method to detect interactions between proteins of the nuclear envelope in live cells. MCLIP has been valuable in identifying interaction partners of Samp1. In interphase, Samp1 distributes in distinct micro-domains of the inner nuclear membrane and interacts with the nuclear lamina, emerin and the LINC complex protein SUN1, suggesting that Samp1 might have a functional role associated with both the nucleoskeleton and cytoskeleton.

In mitosis Samp1 distributes in filamentous membrane structures partially overlapping with kinetochore microtubules of the mitotic spindle. Samp1 binds directly to γ-tubulin and recruits γ-tubulin and Haus6 to the mitotic spindle and thus contributes to spindle assembly. Samp1 also interacts with Aurora B, a kinase important for k-fiber error correction at the kinetochores. Depletion of Samp1 caused an increased activation and distribution of Aurora B at the metaphase plate, decreased formation of stable k-fibers, metaphase prolongation and increased chromosome mis-segregation. Samp1 is the first transmembrane protein found to be involved in mitotic spindle assembly and stability, important for correct segregation of chromosomes.

 

 

Place, publisher, year, edition, pages
Stockholm: Department of Biochemistry and Biophysics, Stockholm University, 2018. p. 64
Keywords
Samp1, LINC complex, nuclear lamina, MCLIP, nuclear envelope, spindle assembly, spindle stability, k-fiber, gamma tubulin, Aurora B, Haus6, Augmin
National Category
Biochemistry and Molecular Biology Cell Biology
Research subject
Neurochemistry with Molecular Neurobiology
Identifiers
urn:nbn:se:su:diva-154093 (URN)978-91-7797-199-3 (ISBN)978-91-7797-200-6 (ISBN)
Public defence
2018-05-03, Magnélisalen, Kemiska övningslaboratoriet, Svante Arrhenius väg 16 B, Stockholm, 10:00 (English)
Opponent
Supervisors
Note

At the time of the doctoral defense, the following paper was unpublished and had a status as follows: Paper 4: Manuscript.

Available from: 2018-04-10 Created: 2018-03-14 Last updated: 2018-04-04Bibliographically approved

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