| 1. |
- Bergqvist, Cecilia, et al.
(author)
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An inner nuclear membrane protein induces rapid differentiation of human induced pluripotent stem cells
- 2017
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In: Stem Cell Research. - : Elsevier BV. - 1873-5061 .- 1876-7753. ; 23, s. 33-38
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Journal article (peer-reviewed)abstract
- The ability of iPSCs (induced pluripotent stem cells) to generate any cell type in the body makes them valuable tools for cell replacement therapies. However, differentiation of iPSCs can be demanding, slowand variable. During differentiation chromatin is re-organized and silent dense heterochromatin becomes tethered to the nuclear periphery by processes involving the nuclear lamina and proteins of the INM(inner nuclearmembrane). The INM protein, Samp1 (Spindle AssociatedMembrane Protein 1) interacts with Lamin A/C and the INMprotein Emerin, which has a chromatin binding LEM(Lap2-Emerin-Man1)-domain. In this paperweinvestigate if Samp1 can play a role in the differentiation of iPSCs. Samp1 levels increased as differentiating iPSCs started to express Lamin A/C. Interestingly, even under pluripotent culturing conditions, ectopic expression of Samp1 induced a rapid differentiation of iPSCs, ofwhich some expressed the neuronal marker beta III-tubulin already after 6 days. This suggests that Samp1 is involved in early differentiation of iPSCs and could potentially be explored as a tool to promote progression of the differentiation process.
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| 2. |
- Buch, Charlotta, et al.
(author)
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An integral protein of the inner nuclear membrane localizes to the mitotic spindle in mammalian cells
- 2009
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In: Journal of Cell Science. - : The Company of Biologists. - 0021-9533 .- 1477-9137. ; 122:12, s. 2100-2107
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Journal article (peer-reviewed)abstract
- Here, we characterize a transmembrane protein of the nuclear envelope that we name spindle-associated membrane protein 1 (Samp1). The protein is conserved in metazoa and fission yeast and is homologous to Net5 in rat and Ima1 in Schizosaccharomyces pombe. We show that, in human cells, the protein is a membrane-spanning polypeptide with an apparent molecular mass of 43 kDa. This is consistent with a predicted polypeptide of 392 amino acids that has five transmembrane segments and its C-terminus exposed to the nucleoplasm. During interphase, Samp1 was specifically distributed in the inner nuclear membrane. Post-transcriptional silencing of Samp1 expression resulted in separation of centrosomes from the nuclear envelope, indicating that it is functionally connected to the cytoskeleton. At the onset of mitosis, most of the protein dispersed out into the ER, as expected. However, during mitosis, a significant fraction of the protein specifically localized to the polar regions of the mitotic spindle. We demonstrate for the first time, in human cells, the existence of a membranous structure overlapping with the mitotic spindle. Interestingly, another integral inner nuclear membrane protein, emerin, was absent from the spindle-associated membranes. Thus, Samp1 defines a specific membrane domain associated with the mitotic spindle.
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| 3. |
- Figueroa, Ricardo A., et al.
(author)
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Microtubule-associated nuclear envelope proteins in interphase and mitosis
- 2011
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In: Biochemical Society Transactions. - 0300-5127 .- 1470-8752. ; 39, s. 1786-1789
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Journal article (peer-reviewed)abstract
- The LINC (linker of nucleoskeleton and cytoskeleton) complex forms a transcisternal bridge across the NE (nuclear envelope) that connects the cytoskeleton with the nuclear interior. This enables some proteins of the NE to communicate with the centrosome and the microtubule cytoskeleton. The position of the centrosome relative to the NE is of vital importance for many cell functions, such as cell migration and division, and centrosomal dislocation is a frequent phenotype in laminopathic disorders. Also in mitosis, a small group of transmembrane NE proteins associate with microtubules when they concentrate in a specific membrane domain associated with the mitotic spindle. The present review discusses structural and functional aspects of microtubule association with NE proteins and how this association may be maintained over the cell cycle.
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| 4. |
- Figueroa, Ricardo, et al.
(author)
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A transmembrane inner nuclear membrane protein in the mitotic spindle
- 2010
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In: Nucleus (Austin). - 1949-1042. ; 1:3, s. 249-253
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Journal article (peer-reviewed)abstract
- We have recently characterized a novel transmembrane protein of the inner nuclear membrane of mammalian cells. The protein has two very interesting features. First, despite being an integral membrane protein it is able to concentrate in the membranes colocalizing with the mitotic spindle in metaphase and anaphase. Hence, the protein was named Samp1, Spindle associated membrane protein 1. Secondly, it displays a functional connection to centrosomes. This article discusses various aspects of Samp1 in relation to possible cellular function(s).
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| 5. |
- Gudise, Santhosh Kumar
(author)
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The functional organization of nuclear envelope proteins
- 2014
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Doctoral thesis (other academic/artistic)abstract
- In eukaryotic cells, the nucleus is enclosed by a double lipid membrane, termed the nuclear envelope (NE). The NE consists of the outer nuclear membrane (ONM), the inner nuclear membrane (INM), the nuclear pore complexes (NPCs) and the nuclear lamina. Recently it has been realized that the NE proteins not only serve structural functions but are also involved in a diverse group of genetic diseases collectively termed laminopathies or envelopathies. So far, only a few NE proteins have been characterized in detail. Here, we have identified and investigated a novel transmembrane protein from the NE, which is highly conserved in evolution. We termed the protein, Spindle associated membrane protein 1 (Samp1). During mitosis, a subpopulation of Samp1 is concentrated in the mitotic spindle. Samp1 has four transmembrane domains and is specifically localized to the INM. The N-terminal half of Samp1 contains a Zinc finger domain and is exposed in the nucleoplasm. Over expression of Zinc finger mutants of Samp1 gave an abnormal phenotype characterized by disruption of the localization of endogenous Samp1 and a specific set of NE proteins, suggesting that Samp1 is functionally associated with LINC complex and A-type lamina network proteins. After posttranscriptional silencing of Samp1 expression we showed that Samp1 is required for correct localization of Emerin to the NE. We also showed that Samp1 interacts with Emerin in live cells and that this interaction can occur by direct binding. The fact that the interaction between Emerin and Samp1 depended on Zinc, supports the idea that Samp1 has functional Zinc finger(s). Posttranscriptional silencing of Samp1 gave rise to an increase in the distance between the centrosome and the NE, suggesting that Samp1 is functionally associated with the microtubule cytoskeleton, most likely mediated via the LINC complexes. Using high-resolution fluorescence microscopy we showed that Samp1 is distributed in a distinct pattern in the NE and partially colocalized with the LINC complex protein, Sun1. We also showed that the Samp1 can interact with Sun1 in live cells. We developed a novel method, Membrane protein Cross-Link ImmunoPrecipitation (MCLIP) that enables detection of specific interactions of NE proteins in live cells. Using MCLIP we identified specific interaction partners of Samp1 in U2OS cells. Human induced pluripotent stem cells (hiPSCs) displayed increased expression of Samp1 during differentiation. Over expression of YFP-Samp1 induced a rapid differentiation of hiPSCs into neurons. The medium from
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| 6. |
- Gudise, Santhosh, et al.
(author)
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Samp1 is functionally associated with the LINC complex and A-type lamina networks
- 2011
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In: Journal of Cell Science. - : The Company of Biologists. - 0021-9533 .- 1477-9137. ; 124, s. 2077-2085
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Journal article (peer-reviewed)abstract
- The transmembrane inner nuclear membrane (INM) protein Samp1 is required for anchoring centrosomes near the nuclei. Using high-resolution fluorescence microscopy we show that Samp1 is distributed in a distinct and characteristic pattern in the nuclear envelope (NE), where it partially colocalizes with the LINC complex protein Sun1. By studying the localization of Samp1 deletion mutants and fusion proteins, we conclude that the cysteine-rich N-terminal half of Samp1 is nucleoplasmically exposed and is responsible for targeting to the INM. It contains four conserved CxxC motifs with the potential to form zinc fingers. The distribution of cysteine-to-alanine substitution mutants, designed to prevent zinc finger formation, showed that NE localization of Samp1 depends on intact CxxC motifs. Overexpression of Samp1 zinc finger mutants produced an abnormal dominant phenotype characterized by disrupted organization of a selective subset NE proteins, including emerin, Sun1, endogenous Samp1 and, in some cases, lamin A/C, but not lamin B, Sun2 or nucleoporins. Silencing of Samp1 expression showed that emerin depends on Samp1 for its correct localization in the NE. Our results demonstrate that Samp1 is functionally associated with the LINC complex protein Sun1 and proteins of the A-type lamina network.
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| 7. |
- Jafferali, Mohammed Hakim, et al.
(author)
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MCLIP, an effective method to detect interactions of transmembrane proteins of the nuclear envelope in live cells
- 2014
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In: Biochimica et Biophysica Acta - Biomembranes. - : Elsevier BV. - 0005-2736 .- 1879-2642 .- 0006-3002. ; 1838:10, s. 2399-2403
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Journal article (peer-reviewed)abstract
- 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.
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