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Communication between the cytoskeleton and the nuclear envelope to position the nucleus. 2007
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In most eukaryotic cells, the nucleus is localized to a specific location. This highlight article focuses on recent advances describing the mechanisms of nuclear migration and anchorage. Central to nuclear positioning mechanisms is the communication between the nuclear envelope and the cytoskeleton. All three components of the cytoskeleton, microtubules, actin filaments and intermediate filaments, are involved in nuclear positioning to varying degrees in different cell types. KASH proteins on the outer nuclear membrane connect to SUN proteins on the inner nuclear membrane. Together they transfer forces between the cytoskeleton and the nuclear lamina. Once at the outer nuclear membrane, KASH proteins can interact with the cytoskeleton. Nuclear migrations are a component of many cellular migration events and defects in nuclear positioning lead to human diseases, most notably lissencephaly. |
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UNC-84 is required to localize UNC-83 to the nuclear envelope where it functions during nuclear migration. A KASH domain in UNC-83 was identified. KASH domains are conserved in the nuclear envelope proteins Syne/nesprins, Klarsicht, MSP-300, and ANC-1. C. elegans UNC-83 was shown to localize to the outer nuclear membrane and UNC-84 to the inner nuclear membrane in transfected mammalian cells, suggesting the KASH and SUN protein targeting mechanisms are conserved. Deletion of the KASH domain of UNC-83 blocked nuclear migration and localization to the C. elegans nuclear envelope. Some point mutations in the UNC-83 KASH domain disrupted nuclear migration, even if they localized normally. At least two separable portions of the C-terminal half of UNC-84 were found to interact with the UNC-83 KASH domain in a membrane-bound, split-ubiquitin yeast two-hybrid system. However, the SUN domain was essential for UNC-84 function and UNC-83 loclization in vivo. These data support the model that KASH and SUN proteins bridge the nuclear envelope, connecting the nuclear lamina to cytoskeletal components. This mechanism appears conserved across eukaryotes and is the first proposed mechanism to target proteins specifically to the outer nuclear membrane. |
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During late stages of Drosophila oogenesis, the cytoplasm of nurse cells in the egg chamber is rapidly transferred (‘‘dumped’’) to oocytes, while the nurse cell nuclei are anchored by a mechanism that involves the actin cytoskeleton. The factors that mediate this interaction between nuclei and actin cytoskeleton are unknown. MSP-300 is the likely Drosophila ortholog of the mammalian Syne-1 and -2 and C. elegans ANC-1 proteins, contained both actin-binding and nuclear envelope localization domains. By using an antibody against C-terminus of MSP-300, we find that MSP-300 is distributed throughout the cytoplasm and accumulates at the nuclear envelope of nurse cells and the oocyte. A GFP fusion protein containing the C-terminal region of MSP-300 is also sufficient to localize protein on the nuclear envelope in oocytes. To eliminate the maternal gene activity during oogenesis, we generated homozygous germ-line clones of a loss-of-function mutation in msp-300 in otherwise heterozygous mothers. In the mutant egg chambers that develop from such clones, cytoplasmic dumping of nurse cells is severely disturbed. The nuclei of nurse cells and the oocyte are mislocalized and the usually well-organized actin structures are severely disrupted. These results indicate that maternal MSP-300 plays an important role in actin-dependent nuclear anchorage during cytoplasmic transport. |
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KASH 'n Karry: the KASH domain family of cargo-specific cytoskeletal adaptor proteins. 2005
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A diverse family of proteins has been discovered with a small C-terminal KASH domain in common. KASH domain proteins are localized uniquely to the outer nuclear envelope, enabling their cytoplasmic extensions to tether the nucleus to actin filaments or microtubules. KASH domains are targeted to the outer nuclear envelope by SUN domains of inner nuclear envelope proteins. Several KASH protein genes were discovered as mutant alleles in model organisms with defects in developmentally regulated nuclear positioning. Recently, KASH-less isoforms have been found that connect the cytoskeleton to organelles other than the nucleus. A widened view of these proteins is now emerging, where KASH proteins and their KASH-less counterparts are cargo-specific adaptors that not only link organelles to the cytoskeleton but also regulate developmentally specific organelle movements. |
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Syne proteins anchor muscle nuclei at the neuromuscular junction. 2005
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Vertebrate skeletal muscle fibers contain hundreds of nuclei, of which three to six are functionally specialized and stably anchored beneath the postsynaptic membrane at the neuromuscular junction (NMJ). The mechanisms that localize synaptic nuclei and the roles they play in neuromuscular development are unknown. Syne-1 is concentrated at the nuclear envelope of synaptic nuclei; its Caenorhabditis elegans orthologue ANC-1 functions to tether nuclei to the cytoskeleton. To test the involvement of Syne proteins in nuclear anchoring, we generated transgenic mice overexpressing the conserved C-terminal Klarsicht ANC-1 Syne homology domain of Syne-1. The transgene acted in a dominant interfering fashion, displacing endogenous Syne-1 from the nuclear envelope. Muscle nuclei failed to aggregate at the NMJ in transgenic mice, demonstrating that localization and positioning of synaptic nuclei require Syne proteins. We then exploited this phenotype to show that synaptic nuclear aggregates are dispensable for maturation of the NMJ. |
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A genetic approach to study the role of nuclear envelope components in nuclear positioning. 2005
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In many cell types, the nucleus is positioned to a specific location. Our work and others have demonstrated that several integral nuclear envelope proteins function to move the nucleus and to anchor it in place. Our forward genetic approach has identified three components of the nuclear envelope involved in nuclear positioning. ANC-1 consists of two actin-binding calponin domains, a huge central coiled domain, and a nuclear envelope targeting domain termed the KASH domain. ANC-1 functions to physically tether the actin cytoskeleton to the outer nuclear membrane. UNC-83 is a novel protein that functions in an unknown manner during nuclear migration. UNC-83 contains a domain with weak homology to the KASH domain of ANC-1. UNC-84 is a SUN protein that is required for both nuclear migration and anchorage. UNC-84 recruits both UNC-83 and ANC-1 to the nuclear envelope. We propose a model where UNC-84 is an integral component of the inner nuclear membrane, with its SUN domain in the perinuclear space. The SUN domain then recruits ANC-1 and UNC-83, through interactions with their KASH domains, to the outer nuclear envelope. Together these proteins function to bridge the two membranes of the nuclear envelope, connecting the nuclear matrix to the cytoskeleton. |
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ANChors away: an actin based mechanism of nuclear positioning. 2003
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Mechanisms for nuclear migration and nuclear anchorage function together to control nuclear positioning. Both tubulin and actin networks play important roles in nuclear positioning. The actin cytoskeleton has been shown to position nuclei in a variety of systems from yeast to plants and animals. It can either act as a stable skeleton to anchor nuclei or supply the active force to move nuclei. Two C. elegans genes and their homologues play important roles in these processes. Syne/ANC-1 anchors nuclei by directly tethering the nuclear envelope to the actin cytoskeleton, and UNC-84/SUN functions at the nuclear envelope to recruit Syne/ANC-1. |
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Role of ANC-1 in tethering nuclei to the actin cytoskeleton. 2002 [Supplement]
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Mutations in anc-1 (nuclear anchorage defective) disrupt the positioning of nuclei and mitochondria in Caenorhabditis elegans. ANC-1 is shown to consist of mostly coiled regions with a nuclear envelope localization domain (called the KASH domain) and an actin-binding domain; this structure was conserved with the Drosophila protein Msp-300 and the mammalian Syne proteins. Antibodies against ANC-1 localized cytoplasmically and were enriched at the nuclear periphery in an UNC-84–dependent manner. Overexpression of the KASH domain or the actin-binding domain caused a dominant negative anchorage defect. Thus, ANC-1 may connect nuclei to the cytoskeleton by interacting with UNC-84 at the nuclear envelope and with actin in the cytoplasm. |
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Lamin-dependent localization of UNC-84, a protein required for nuclear migration in C. elegans. 2002
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Mutations in the Caenorhabditis elegans unc-84 gene cause defects in nuclear migration and anchoring. We show that endogenous UNC-84 protein colocalizes with Ce-lamin at the nuclear envelope and that the envelope localization of UNC-84 requires Ce-lamin. We also show that during mitosis, UNC-84 remains at the nuclear periphery until late anaphase, similar to known inner nuclear membrane proteins. UNC-84 protein is first detected at the 26-cell stage and thereafter is present in most cells during development and in adults. UNC-84 is properly expressed in unc-83 and anc-1 lines, which have phenotypes similar to unc-84, suggesting that neither the expression nor nuclear envelope localization of UNC-84 depends on UNC-83 or ANC-1 proteins. The envelope localization of Ce-lamin, Ce-emerin, Ce-MAN1, and nucleoporins are unaffected by the loss of UNC-84. UNC-84 is not required for centrosome attachment to the nucleus because centrosomes are localized normally in unc-84 hyp7 cells despite a nuclear migration defect. Models for UNC-84 localization are discussed. |
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Worm Breeding for Super Geniuses: A guide to genetic mapping in C. elegans. 2001
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What this guide is and isn't. This guide is neither a basic text in Mendelian genetics nor is it in any way a comprehensive description of C. elegans biology. Detailed information about the latter can be gleaned from C. elegans I and C. elegans II by Cold Spring Harbor Press Inc. This guide does assume a working knowledge of basic genetics and will be of limited use to those who lack some background in this area. It is our hope that this text may serve as a supplement to existing published materials and that it will facilitate the successful breeding of worms by those new to the field. |
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Nuclear migration plays an essential role in the growth and development of a wide variety of eukaryotes. Mutations in unc-84, which encodes a conserved component of the nuclear envelope, have been shown to disrupt nuclear migration in two C. elegans tissues. We show that mutations in unc-83 disrupt nuclear migration in a similar manner in migrating P cells, hyp7 precursors and the intestinal primordium, but have no obvious defects in the association of centrosomes with nuclei or the structure of the nuclear lamina of migrating nuclei. We also show that unc-83 encodes a novel transmembrane protein. We identified three unc-83 transcripts that are expressed in a tissuespecific manner. Antibodies against UNC-83 co-localized to the nuclear envelope with lamin and UNC-84. Unlike UNC- 84, UNC-83 localized to only specific nuclei, many of which were migratory. UNC-83 failed to localize to the nuclear envelope in unc-84 mutants with lesions in the conserved SUN domain of UNC-84, and UNC-83 interacted with the SUN domain of UNC-84 in vitro, suggesting that these two proteins function together during nuclear migration. We favor a model in which UNC-84 directly recruits UNC-83 to the nuclear envelope where they help transfer force between the cytoskeleton and the nucleus. |
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