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Identification and characterization of nuclear envelope protein interactions
Stockholm University, Faculty of Science, Department of Neurochemistry.ORCID iD: 0000-0002-3481-1106
2015 (English)Licentiate thesis, comprehensive summary (Other academic)
Abstract [en]

The Nuclear envelope which surrounds the chromatin of eukaryotic cells contains more than a hundred transmembrane proteins. Mutations in some genes encoding nuclear envelope proteins give rise to human diseases including neurological disorders. The function of many nuclear envelope proteins is not well established. This is partly because nuclear envelope proteins and their interactions are difficult to study due to the inherent resistance to extraction of nuclear envelope proteins. We have developed a novel method called MCLIP, to identify interacting partners of nuclear envelope proteins in live cells. Using MCLIP, we found three new binding partners of the inner nuclear membrane protein Samp1: the intermediate filament protein Lamin B1, the LINC complex protein Sun1 and the G-protein Ran. Furthermore, using in vitro studies, we show that Samp1 binds both Emerin and Ran directly. We have also studied the interaction between Samp1 and Ran in detail.

Place, publisher, year, edition, pages
Stockholm: Department of Neurochemistry, Stockholm University , 2015. , 48 p.
Keyword [en]
Samp1, MCLIP, Nuclear envelope, Ran, Emerin
National Category
Chemical Sciences Biological Sciences Cell and Molecular Biology
Research subject
Neurochemistry with Molecular Neurobiology
Identifiers
URN: urn:nbn:se:su:diva-122052ISBN: 978-91-7649-289-5 (print)OAI: oai:DiVA.org:su-122052DiVA: diva2:862387
Presentation
2015-11-04, Heilbronnsalen, C458, Svante Arrhenius väg 16 B, Stockholm, 13:00 (English)
Opponent
Supervisors
Funder
Swedish Research Council, 621-2010-448Swedish Cancer Society, 110590Stiftelsen Olle Engkvist Byggmästare
Available from: 2015-10-23 Created: 2015-10-21 Last updated: 2015-10-23Bibliographically approved
List of papers
1. MCLIP, an effective method to detect interactions of transmembrane proteins of the nuclear envelope in live cells
Open this publication in new window or tab >>MCLIP, an effective method to detect interactions of transmembrane proteins of the nuclear envelope in live cells
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2014 (English)In: Biochimica et Biophysica Acta - Biomembranes, ISSN 0005-2736, E-ISSN 1879-2642, Vol. 1838, no 10, 2399-2403 p.Article in journal (Refereed) Published
Abstract [en]

Investigating interactions of proteins in the nuclear envelope (NE) using co-immunoprecipitation (Co-IP) has previously been difficult or even impossible due to their inherent resistance to extraction. We have developed a novel method, MCLIP (Membrane protein Cross-Link ImmunoPrecipitation), which takes advantage of a cell permeable crosslinker to enable effective detection and analysis of specific interactions of NE proteins in live cells using Western blot. Using MCLIP we show that, in U2OS cells, the integral inner nuclear membrane protein Samp1 interacts with Lamin B1, the LINC (Linker of nucleoskeleton and cytoskeleton) complex protein, Sun1 and the soluble small GTPase Ran. The results show that the previously detected in vitro interaction between Samp1 and Emerin also takes place in live cells. In vitro pull down experiments show, that the nucleoplasmic domains of Samp1 and Emerin can bind directly to each other. We also, show that MCLIP is suitable to coprecipitate protein interactions in different stages of the cell cycle.

Keyword
Samp1, Nuclear envelope, Nuclear membrane, Crosslinking, CoIP, Protein–protein interaction
National Category
Chemical Sciences Biochemistry and Molecular Biology
Research subject
Neurochemistry with Molecular Neurobiology
Identifiers
urn:nbn:se:su:diva-109181 (URN)10.1016/j.bbamem.2014.06.008 (DOI)000340975600005 ()
Funder
Swedish Research Council, 621-2010-448Swedish Cancer Society, 110590
Available from: 2014-11-14 Created: 2014-11-14 Last updated: 2017-10-24Bibliographically approved
2. The nucleoplasmically exposed N-terminal domain of the inner nuclear membrane protein, Samp1, directly binds to the small monomeric GTPase, Ran
Open this publication in new window or tab >>The nucleoplasmically exposed N-terminal domain of the inner nuclear membrane protein, Samp1, directly binds to the small monomeric GTPase, Ran
(English)Manuscript (preprint) (Other academic)
Keyword
Samp1, Ran, protein-ptotein interaction, Nuclear envelope
National Category
Cell and Molecular Biology
Research subject
Neurochemistry and Neurotoxicology
Identifiers
urn:nbn:se:su:diva-121617 (URN)
Funder
Swedish Cancer SocietySwedish Research Council
Available from: 2015-10-12 Created: 2015-10-12 Last updated: 2016-01-29Bibliographically approved

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