| 1. |
- Boban, Mirta, et al.
(författare)
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A nuclear ubiquitin-proteasome pathway targets the inner nuclear membrane protein Asi2 for degradation
- 2014
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Ingår i: Journal of Cell Science. - : The Company of Biologists. - 0021-9533 .- 1477-9137. ; 127:16, s. 3603-3613
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Tidskriftsartikel (refereegranskat)abstract
- The nuclear envelope consists of inner and outer nuclear membranes. Whereas the outer membrane is an extension of the endoplasmic reticulum, the inner nuclear membrane (INM) represents a unique membranous environment containing specific proteins. The mechanisms of integral INM protein degradation are unknown. Here, we investigated the turnover of Asi2, an integral INM protein in Saccharomyces cerevisiae. We report that Asi2 is degraded by the proteasome independently of the vacuole and that it exhibited a half-life of similar to 45 min. Asi2 exhibits enhanced stability in mutants lacking the E2 ubiquitin conjugating enzymes Ubc6 or Ubc7, or the E3 ubiquitin ligase Doa10. Consistent with these data, Asi2 is post-translationally modified by poly-ubiquitylation in a Ubc7- and Doa10-dependent manner. Importantly Asi2 degradation is significantly reduced in a sts1-2 mutant that fails to accumulate proteasomes in the nucleus, indicating that Asi2 is degraded in the nucleus. Our results reveal a molecular pathway that affects the stability of integral proteins of the inner nuclear membrane and indicate that Asi2 is subject to protein quality control in the nucleus.
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| 2. |
- Boban, Mirta, et al.
(författare)
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Atypical Ubiquitylation in Yeast Targets Lysine-less Asi2 for Proteasomal Degradation
- 2015
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Ingår i: Journal of Biological Chemistry. - 0021-9258 .- 1083-351X. ; 290:4, s. 2489-2495
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Tidskriftsartikel (refereegranskat)abstract
- Proteins are typically targeted for proteasomal degradation by the attachment of a polyubiquitin chain to epsilon-amino groups of lysine residues. Non-lysine ubiquitylation of proteasomal substrates has been considered an atypical and rare event limited to complex eukaryotes. Here we report that a fully functional lysine-less mutant of an inner nuclear membrane protein in yeast, Asi2, is polyubiquitylated and targeted for proteasomal degradation. Efficient degradation of lysine-free Asi2 requires E3-ligase Doa10 and E2 enzymes Ubc6 and Ubc7, components of the endoplasmic reticulum-associated degradation pathway. Together, our data suggest that non-lysine ubiquitylation may be more prevalent than currently considered.
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| 3. |
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| 4. |
- Lu, Wenshu, et al.
(författare)
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Sun1 forms immobile macromolecular assemblies at the nuclear envelope
- 2008
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Ingår i: Biochimica et Biophysica Acta. Molecular Cell Research. - : Elsevier BV. - 0167-4889 .- 1879-2596. ; 1783:12, s. 2415-2426
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Tidskriftsartikel (refereegranskat)abstract
- SUN-domain proteins form a novel and conserved family of inner nuclear membrane (INM) proteins, which establish physical connections between the nucleoplasm and the cytoskeleton. In the current study, we provide evidence that within the nuclear envelope (NE) Stint proteins form highly immobile oligomeric complexes in interphase cells. By performing inverse fluorescence recovery after photobleaching analysis, we demonstrate in vivo that both perinuclear and nucleoplasmic Sun I segments are essential for maintenance of Sun I immobility at the NE. Our data in Particular underline the self-association properties of the C-terminal coiled-coil Sun I segment, the ability of which to form dimers and tetramers is demonstrated. Furthermore, the Sun1 tertiary Structure involves interchain disulfide bonds that might contribute to higher homo-oligomer formation, although the overall dynamics of the Sun1 C-terminus remains unaffected when the cysteins involved are mutated. While a major Sun1 pool colocalizes with nuclear pore complex proteins, a large fraction of the Sun1 protein assemblies colocalize with immunoreactive foci of Sun2, another SUN-domain paralogue at the NE. We demonstrate that the Sun1 coiled-coil domain permits these heterophilic associations with Sun2. Sun1 therefore provides a non-dynamic platform for the formation of different macromolecular assemblies at the INM Our data support a model in which SUN-protein-containing multivariate complexes may provide versatile outer nuclear membrane attachment sites for cytoskeletal filaments. Crown
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| 5. |
- Pantazopoulou, Marina, et al.
(författare)
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Cdc48 and Ubx1 participate in a pathway associated with the inner nuclear membrane that governs Asi1 degradation
- 2016
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Ingår i: Journal of Cell Science. - : The Company of Biologists. - 0021-9533 .- 1477-9137. ; 129:20, s. 3770-3780
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Tidskriftsartikel (refereegranskat)abstract
- The nuclear envelope is a barrier comprising outer and inner membranes that separate the cytoplasm from the nucleoplasm. The two membranes have different physical characteristics and protein compositions. The processes governing the stability of inner nuclear membrane (INM) proteins are not well characterized. In Saccharomyces cerevisiae, the INM Asi1-Asi3 complex, principally composed of integral membrane proteins Asi1 and Asi3, is an E3 ubiquitin ligase. In addition to its well-documented function in endoplasmic reticulum (ER)-associated degradation, the Doa10 E3 ubiquitin ligase complex partially localizes to the INM. The Asi1-Asi3 and Doa10 complexes define independent INM-associated degradation (INMAD) pathways that target discrete sets of nuclear substrates for proteasomal degradation. Here, we report that Asi1 is rapidly turned over (t(1/2)<= 30 min). Its turnover depends on ubiquitin-mediated degradation by nucleus-localized proteasomes, exhibiting a clear requirement for the E2 ubiquitin-conjugating enzyme Ubc7, Cue1 and the AAA ATPase Cdc48 and co-factor Ubx1. Asi1 turnover occurs largely independently of the Asi1-Asi3 or Doa10 complexes, indicating that it is subject to quality control at the INM in a manner distinct from that of the characterized INMAD pathways.
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| 6. |
- Thanisch, Katharina, et al.
(författare)
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Nuclear envelope localization of LEMD2 is developmentally dynamic and lamin A/C dependent yet insufficient for heterochromatin tethering
- 2017
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Ingår i: Differentiation. - : Elsevier BV. - 0301-4681 .- 1432-0436. ; 94, s. 58-70
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Tidskriftsartikel (refereegranskat)abstract
- Peripheral heterochromatin in mammalian nuclei is tethered to the nuclear envelope by at least two mechanisms here referred to as the A- and B-tethers. The A-tether includes lamins A/C and additional unknown components presumably INM protein(s) interacting with both lamins A/C and chromatin. The B tether includes the inner nuclear membrane (INM) protein Laurin B-receptor, which binds B-type lamins and chromatin. Generally, at least one of the tethers is always present in the nuclear envelope of mammalian cells. Deletion of both causes the loss of peripheral heterochromatin and consequently inversion of the entire nuclear architecture, with this occurring naturally in rod photoreceptors of nocturnal mammals. The tethers are differentially utilized during development, regulate gene expression in opposite manners, and play an important role during cell differentiation. Here we aimed to identify the unknown chromatin binding component(s) of the A-tether. We analyzed 10 mouse tissues by immunostaining with antibodies against 7 INM proteins and found that every cell type has specific, although differentially and developmentally regulated, sets of these proteins. In particular, we found that INM protein LEMD2 is concomitantly expressed with A-type lamins in various cell types but is lacking in inverted nuclei of rod cells. Truncation or deletion of Lmna resulted in the downregulation and mislocalization of LEMD2, suggesting that the two proteins interact and pointing at LEMD2 as a potential chromatin binding mediator of the A-tether. Using nuclei of mouse rods as an experimental model lacking peripheral heterochromatin, we expressed a LEMD2 transgene alone or in combination with lamin C in these cells and observed no restoration of peripheral heterochromatin in either case. We conclude that in contrary to the B-tether, the A-tether has a more intricate composition and consists of multiple components that presumably vary, at differing degrees of redundancy, between cell types and differentiation stages.
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