Schuh, Melina

[["dc.bibliographiccitation.issue","6447"],["dc.bibliographiccitation.journal","Science"],["dc.bibliographiccitation.volume","364"],["dc.contributor.author","So, Chun"],["dc.contributor.author","Seres, K. Bianka"],["dc.contributor.author","Steyer, Anna M."],["dc.contributor.author","Mönnich, Eike"],["dc.contributor.author","Clift, Dean"],["dc.contributor.author","Pejkovska, Anastasija"],["dc.contributor.author","Möbius, Wiebke"],["dc.contributor.author","Schuh, Melina"],["dc.date.accessioned","2022-03-01T11:47:19Z"],["dc.date.available","2022-03-01T11:47:19Z"],["dc.date.issued","2019"],["dc.description.abstract","A new phase in egg biology Chromosome segregation typically requires centrosomes, which generate the microtubule spindle. However, mammalian eggs build a spindle and segregate chromosomes without centrosomes. How acentrosomal spindles are organized has remained elusive. So et al. show that centrosomal and microtubule-associated proteins are repurposed into a large “liquid-like meiotic spindle domain” (LISD) in eggs. The domains localized to spindle poles and also extended to the spindle fibers that connect to kinetochores. LISDs formed by phase separation and were required for spindle assembly, serving as reservoirs that locally sequester and mobilize spindle assembly factors within the large egg cytoplasm. Science , this issue p. eaat9557"],["dc.description.abstract","Phase separation of microtubule regulatory factors promotes acentrosomal spindle assembly in mammalian oocytes."],["dc.description.abstract","INTRODUCTION Mammalian embryos frequently develop abnormally, resulting in miscarriages and genetic disorders such as Down syndrome. The major cause for aberrant embryonic development is chromosome segregation errors during egg meiosis. Unlike somatic cells and male germ cells, eggs segregate chromosomes with a specialized microtubule spindle that lacks centrosomes. Canonical centrosomes consist of a pair of centrioles surrounded by pericentriolar material and are the main microtubule organizing centers in centrosomal spindles. How acentrosomal spindles are organized in mammalian eggs is still poorly understood. RATIONALE Despite the absence of centrosomes, mammalian eggs express many centrosomal proteins. We set out to investigate systematically how these centrosomal proteins localize to acentrosomal spindles and organize microtubules in mammalian eggs. RESULTS We analysed the localization of 70 centrosomal and spindle-related proteins by combining high-resolution microscopy in live and fixed mouse eggs. Unexpectedly, 19 of these proteins localized to a domain that permeated a large region of the spindle and formed prominent spherical protrusions, which were dynamic, fused with each other, and extended well beyond the spindle poles. The domain included centrosomal proteins (AKAP450, CEP170, and KIZ), centriolar satellite proteins (CEP72, PCM1, and LRRC36), minus-end binding proteins (CAMSAP3 and KANSL3), dynein-related proteins (HOOK3, NDE1, NDEL1, and SPDL1), and proteins that control microtubule nucleation and stability (CHC17, chTOG, GTSE1, HAUS6, MCAK, MYO10, and TACC3). Proteins within this domain were dynamic and could redistribute rapidly throughout the entire spindle region. By combining in vitro and in vivo assays, we found that the domain forms by phase separation and behaves similar to a liquid. We hence termed it the liquid-like meiotic spindle domain (LISD). The LISD was also present in spindles in bovine, ovine, and porcine eggs and is thus widely conserved. Many LISD proteins have been studied extensively in mitosis, yet a similar structure has not been reported in somatic cells, suggesting that the LISD is likely exclusive to acentrosomal spindles in oocytes. Assembly of the LISD was controlled by the regulatory kinase aurora A and dependent on the aurora A substrate TACC3 as well as the clathrin heavy chain CHC17, which binds to microtubules together with TACC3. Disruption of the LISD via different means released microtubule regulatory factors within this domain into the cytoplasm and led to severe spindle defects. Spindles were smaller and less stable and took longer to segregate chromosomes. Microtubule growth rates were significantly decreased, and their overall turnover was significantly increased. Both the microtubules that bind to the chromosomes’ kinetochores (kinetochore fibers) as well as microtubules that overlap in an antiparallel manner in the spindle midzone (interpolar microtubules) were strongly depleted. Together, these data establish that the LISD is required for efficient microtubule assembly and to form stable acentrosomal spindles. CONCLUSION Our data uncover a previously unknown principle of acentrosomal spindle assembly in mammalian eggs: Meiotic spindle assembly is facilitated by a prominent liquid-like domain that contains multiple microtubule regulatory factors and sequesters them in a dynamic manner in proximity to spindle microtubules. Enriching microtubule regulatory factors in local proximity to the spindle may be particularly important in large cells such as eggs, where they would otherwise be dispersed throughout the cytoplasm. Liquid-liquid phase separation may be an ideal principle for such an enrichment: It sequesters factors within proximity to microtubules but still allows them to diffuse dynamically throughout the spindle. This could help to promote the even distribution of spindle assembly factors throughout the spindle and to titrate their local concentration to drive efficient spindle assembly within the large egg cytoplasm. Acentrosomal spindle in a mouse egg. A liquid-like meiotic spindle domain (LISD, cyan) forms prominent spherical protrusions at acentrosomal spindle poles and extends into the spindle region (magenta, right) toward chromosomes (magenta, left). The LISD forms by phase separation and is required for spindle assembly, serving as a reservoir that locally sequesters and mobilizes microtubule regulatory factors within the large egg cytoplasm. Scale bar, 5 μm."],["dc.description.abstract","Mammalian oocytes segregate chromosomes with a microtubule spindle that lacks centrosomes, but the mechanisms by which acentrosomal spindles are organized and function are largely unclear. In this study, we identify a conserved subcellular structure in mammalian oocytes that forms by phase separation. This structure, which we term the liquid-like meiotic spindle domain (LISD), permeates the spindle poles and forms dynamic protrusions that extend well beyond the spindle. The LISD selectively concentrates multiple microtubule regulatory factors and allows them to diffuse rapidly within the spindle volume. Disruption of the LISD via different means disperses these factors and leads to severe spindle assembly defects. Our data suggest a model whereby the LISD promotes meiotic spindle assembly by serving as a reservoir that sequesters and mobilizes microtubule regulatory factors in proximity to spindle microtubules."],["dc.description.abstract","A new phase in egg biology Chromosome segregation typically requires centrosomes, which generate the microtubule spindle. However, mammalian eggs build a spindle and segregate chromosomes without centrosomes. How acentrosomal spindles are organized has remained elusive. So et al. show that centrosomal and microtubule-associated proteins are repurposed into a large “liquid-like meiotic spindle domain” (LISD) in eggs. The domains localized to spindle poles and also extended to the spindle fibers that connect to kinetochores. LISDs formed by phase separation and were required for spindle assembly, serving as reservoirs that locally sequester and mobilize spindle assembly factors within the large egg cytoplasm. Science , this issue p. eaat9557"],["dc.description.abstract","Phase separation of microtubule regulatory factors promotes acentrosomal spindle assembly in mammalian oocytes."],["dc.description.abstract","INTRODUCTION Mammalian embryos frequently develop abnormally, resulting in miscarriages and genetic disorders such as Down syndrome. The major cause for aberrant embryonic development is chromosome segregation errors during egg meiosis. Unlike somatic cells and male germ cells, eggs segregate chromosomes with a specialized microtubule spindle that lacks centrosomes. Canonical centrosomes consist of a pair of centrioles surrounded by pericentriolar material and are the main microtubule organizing centers in centrosomal spindles. How acentrosomal spindles are organized in mammalian eggs is still poorly understood. RATIONALE Despite the absence of centrosomes, mammalian eggs express many centrosomal proteins. We set out to investigate systematically how these centrosomal proteins localize to acentrosomal spindles and organize microtubules in mammalian eggs. RESULTS We analysed the localization of 70 centrosomal and spindle-related proteins by combining high-resolution microscopy in live and fixed mouse eggs. Unexpectedly, 19 of these proteins localized to a domain that permeated a large region of the spindle and formed prominent spherical protrusions, which were dynamic, fused with each other, and extended well beyond the spindle poles. The domain included centrosomal proteins (AKAP450, CEP170, and KIZ), centriolar satellite proteins (CEP72, PCM1, and LRRC36), minus-end binding proteins (CAMSAP3 and KANSL3), dynein-related proteins (HOOK3, NDE1, NDEL1, and SPDL1), and proteins that control microtubule nucleation and stability (CHC17, chTOG, GTSE1, HAUS6, MCAK, MYO10, and TACC3). Proteins within this domain were dynamic and could redistribute rapidly throughout the entire spindle region. By combining in vitro and in vivo assays, we found that the domain forms by phase separation and behaves similar to a liquid. We hence termed it the liquid-like meiotic spindle domain (LISD). The LISD was also present in spindles in bovine, ovine, and porcine eggs and is thus widely conserved. Many LISD proteins have been studied extensively in mitosis, yet a similar structure has not been reported in somatic cells, suggesting that the LISD is likely exclusive to acentrosomal spindles in oocytes. Assembly of the LISD was controlled by the regulatory kinase aurora A and dependent on the aurora A substrate TACC3 as well as the clathrin heavy chain CHC17, which binds to microtubules together with TACC3. Disruption of the LISD via different means released microtubule regulatory factors within this domain into the cytoplasm and led to severe spindle defects. Spindles were smaller and less stable and took longer to segregate chromosomes. Microtubule growth rates were significantly decreased, and their overall turnover was significantly increased. Both the microtubules that bind to the chromosomes’ kinetochores (kinetochore fibers) as well as microtubules that overlap in an antiparallel manner in the spindle midzone (interpolar microtubules) were strongly depleted. Together, these data establish that the LISD is required for efficient microtubule assembly and to form stable acentrosomal spindles. CONCLUSION Our data uncover a previously unknown principle of acentrosomal spindle assembly in mammalian eggs: Meiotic spindle assembly is facilitated by a prominent liquid-like domain that contains multiple microtubule regulatory factors and sequesters them in a dynamic manner in proximity to spindle microtubules. Enriching microtubule regulatory factors in local proximity to the spindle may be particularly important in large cells such as eggs, where they would otherwise be dispersed throughout the cytoplasm. Liquid-liquid phase separation may be an ideal principle for such an enrichment: It sequesters factors within proximity to microtubules but still allows them to diffuse dynamically throughout the spindle. This could help to promote the even distribution of spindle assembly factors throughout the spindle and to titrate their local concentration to drive efficient spindle assembly within the large egg cytoplasm. Acentrosomal spindle in a mouse egg. A liquid-like meiotic spindle domain (LISD, cyan) forms prominent spherical protrusions at acentrosomal spindle poles and extends into the spindle region (magenta, right) toward chromosomes (magenta, left). The LISD forms by phase separation and is required for spindle assembly, serving as a reservoir that locally sequesters and mobilizes microtubule regulatory factors within the large egg cytoplasm. Scale bar, 5 μm."],["dc.description.abstract","Mammalian oocytes segregate chromosomes with a microtubule spindle that lacks centrosomes, but the mechanisms by which acentrosomal spindles are organized and function are largely unclear. In this study, we identify a conserved subcellular structure in mammalian oocytes that forms by phase separation. This structure, which we term the liquid-like meiotic spindle domain (LISD), permeates the spindle poles and forms dynamic protrusions that extend well beyond the spindle. The LISD selectively concentrates multiple microtubule regulatory factors and allows them to diffuse rapidly within the spindle volume. Disruption of the LISD via different means disperses these factors and leads to severe spindle assembly defects. Our data suggest a model whereby the LISD promotes meiotic spindle assembly by serving as a reservoir that sequesters and mobilizes microtubule regulatory factors in proximity to spindle microtubules."],["dc.identifier.doi","10.1126/science.aat9557"],["dc.identifier.uri","https://resolver.sub.uni-goettingen.de/purl?gro-2/103988"],["dc.language.iso","en"],["dc.notes.intern","DOI-Import GROB-531"],["dc.relation.eissn","1095-9203"],["dc.relation.issn","0036-8075"],["dc.title","A liquid-like spindle domain promotes acentrosomal spindle assembly in mammalian oocytes"],["dc.type","journal_article"],["dc.type.internalPublication","unknown"],["dspace.entity.type","Publication"]]

[["dc.bibliographiccitation.artnumber","7217"],["dc.bibliographiccitation.journal","Nature Communications"],["dc.bibliographiccitation.volume","6"],["dc.contributor.author","Clift, Dean"],["dc.contributor.author","Schuh, Melina"],["dc.date.accessioned","2017-09-07T11:43:42Z"],["dc.date.available","2017-09-07T11:43:42Z"],["dc.date.issued","2015"],["dc.description.abstract","Assembly of a bipolar microtubule spindle is essential for accurate chromosome segregation. In somatic cells, spindle bipolarity is determined by the presence of exactly two centrosomes. Remarkably, mammalian oocytes do not contain canonical centrosomes. This study reveals that mouse oocytes assemble a bipolar spindle by fragmenting multiple acentriolar microtubule-organizing centres (MTOCs) into a high number of small MTOCs to be able to then regroup and merge them into two equal spindle poles. We show that MTOCs are fragmented in a three-step process. First, PLK1 triggers a decondensation of the MTOC structure. Second, BicD2-anchored dynein stretches the MTOCs into fragmented ribbons along the nuclear envelope. Third, KIF11 further fragments the MTOCs following nuclear envelope breakdown so that they can be evenly distributed towards the two spindle poles. Failure to fragment MTOCs leads to defects in spindle assembly, which delay chromosome individualization and congression, putting the oocyte at risk of aneuploidy."],["dc.format.extent","12"],["dc.identifier.doi","10.1038/ncomms8217"],["dc.identifier.gro","3141870"],["dc.identifier.isi","000358839900001"],["dc.identifier.pmid","26147444"],["dc.identifier.uri","https://resolver.sub.uni-goettingen.de/purl?gro-2/1989"],["dc.language.iso","en"],["dc.notes.intern","WoS Import 2017-03-10"],["dc.notes.status","final"],["dc.notes.submitter","PUB_WoS_Import"],["dc.relation.issn","2041-1723"],["dc.title","A three-step MTOC fragmentation mechanism facilitates bipolar spindle assembly in mouse oocytes"],["dc.type","journal_article"],["dc.type.internalPublication","yes"],["dc.type.peerReviewed","yes"],["dc.type.subtype","original"],["dspace.entity.type","Publication"]]

[["dc.bibliographiccitation.firstpage","1692.e18"],["dc.bibliographiccitation.issue","7"],["dc.bibliographiccitation.journal","Cell"],["dc.bibliographiccitation.lastpage","1706.e18"],["dc.bibliographiccitation.volume","171"],["dc.contributor.author","Clift, Dean"],["dc.contributor.author","McEwan, William A."],["dc.contributor.author","Labzin, Larisa I."],["dc.contributor.author","Konieczny, Vera"],["dc.contributor.author","Mogessie, Binyam"],["dc.contributor.author","James, Leo C."],["dc.contributor.author","Schuh, Melina"],["dc.date.accessioned","2018-01-17T13:17:53Z"],["dc.date.available","2018-01-17T13:17:53Z"],["dc.date.issued","2017"],["dc.description.abstract","Methods for the targeted disruption of protein function have revolutionized science and greatly expedited the systematic characterization of genes. Two main approaches are currently used to disrupt protein function: DNA knockout and RNA interference, which act at the genome and mRNA level, respectively. A method that directly alters endogenous protein levels is currently not available. Here, we present Trim-Away, a technique to degrade endogenous proteins acutely in mammalian cells without prior modification of the genome or mRNA. Trim-Away harnesses the cellular protein degradation machinery to remove unmodified native proteins within minutes of application. This rapidity minimizes the risk that phenotypes are compensated and that secondary, non-specific defects accumulate over time. Because Trim-Away utilizes antibodies, it can be applied to a wide range of target proteins using off-the-shelf reagents. Trim-Away allows the study of protein function in diverse cell types, including non-dividing primary cells where genome- and RNA-targeting methods are limited."],["dc.identifier.doi","10.1016/j.cell.2017.10.033"],["dc.identifier.pmid","29153837"],["dc.identifier.uri","https://resolver.sub.uni-goettingen.de/purl?gro-2/11706"],["dc.language.iso","en"],["dc.notes.status","final"],["dc.relation.eissn","1097-4172"],["dc.title","A Method for the Acute and Rapid Degradation of Endogenous Proteins"],["dc.type","journal_article"],["dc.type.internalPublication","unknown"],["dspace.entity.type","Publication"]]

[["dc.bibliographiccitation.firstpage","2596"],["dc.bibliographiccitation.issue","8"],["dc.bibliographiccitation.journal","Nature Protocols"],["dc.bibliographiccitation.lastpage","2596"],["dc.bibliographiccitation.volume","14"],["dc.contributor.author","Clift, Dean"],["dc.contributor.author","So, Chun"],["dc.contributor.author","McEwan, William A."],["dc.contributor.author","James, Leo C."],["dc.contributor.author","Schuh, Melina"],["dc.date.accessioned","2022-03-01T11:46:03Z"],["dc.date.available","2022-03-01T11:46:03Z"],["dc.date.issued","2018"],["dc.identifier.doi","10.1038/s41596-018-0092-8"],["dc.identifier.pii","92"],["dc.identifier.uri","https://resolver.sub.uni-goettingen.de/purl?gro-2/103542"],["dc.language.iso","en"],["dc.notes.intern","DOI-Import GROB-531"],["dc.relation.eissn","1750-2799"],["dc.relation.issn","1754-2189"],["dc.rights.uri","http://www.springer.com/tdm"],["dc.title","Publisher Correction: Acute and rapid degradation of endogenous proteins by Trim-Away"],["dc.type","journal_article"],["dc.type.internalPublication","unknown"],["dspace.entity.type","Publication"]]

[["dc.bibliographiccitation.firstpage","2149"],["dc.bibliographiccitation.issue","10"],["dc.bibliographiccitation.journal","Nature Protocols"],["dc.bibliographiccitation.lastpage","2175"],["dc.bibliographiccitation.volume","13"],["dc.contributor.author","Clift, Dean"],["dc.contributor.author","So, Chun"],["dc.contributor.author","McEwan, William A."],["dc.contributor.author","James, Leo C."],["dc.contributor.author","Schuh, Melina"],["dc.date.accessioned","2022-03-01T11:46:03Z"],["dc.date.available","2022-03-01T11:46:03Z"],["dc.date.issued","2018"],["dc.identifier.doi","10.1038/s41596-018-0028-3"],["dc.identifier.pii","28"],["dc.identifier.uri","https://resolver.sub.uni-goettingen.de/purl?gro-2/103540"],["dc.language.iso","en"],["dc.notes.intern","DOI-Import GROB-531"],["dc.relation.eissn","1750-2799"],["dc.relation.issn","1754-2189"],["dc.rights.uri","http://www.springer.com/tdm"],["dc.title","Acute and rapid degradation of endogenous proteins by Trim-Away"],["dc.type","journal_article"],["dc.type.internalPublication","unknown"],["dspace.entity.type","Publication"]]

[["dc.bibliographiccitation.firstpage","549"],["dc.bibliographiccitation.journal","Nature Reviews Molecular Cell Biology"],["dc.bibliographiccitation.lastpage","562"],["dc.bibliographiccitation.volume","14"],["dc.contributor.author","Clift, Dean"],["dc.contributor.author","Schuh, Melina"],["dc.date.accessioned","2017-09-07T11:47:37Z"],["dc.date.available","2017-09-07T11:47:37Z"],["dc.date.issued","2013"],["dc.description.abstract","Fertilization triggers a complex cellular programme that transforms two highly specialized meiotic germ cells, the oocyte and the sperm, into a totipotent mitotic embryo. Linkages between sister chromatids are remodelled to support the switch from reductional meiotic to equational mitotic divisions; the centrosome, which is absent from the egg, is reintroduced; cell division shifts from being extremely asymmetric to symmetric; genomic imprinting is selectively erased and re-established; and protein expression shifts from translational control to transcriptional control. Recent work has started to reveal how this remarkable transition from meiosis to mitosis is achieved."],["dc.identifier.doi","10.1038/nrm3643"],["dc.identifier.gro","3142296"],["dc.identifier.isi","000323552400011"],["dc.identifier.pmid","23942453"],["dc.identifier.uri","https://resolver.sub.uni-goettingen.de/purl?gro-2/6709"],["dc.language.iso","en"],["dc.notes.intern","WoS Import 2017-03-10 / Funder: European Community [241548]"],["dc.notes.status","final"],["dc.notes.submitter","PUB_WoS_Import"],["dc.relation.eissn","1471-0080"],["dc.relation.issn","1471-0072"],["dc.title","Restarting life: fertilization and the transition from meiosis to mitosis"],["dc.type","review"],["dc.type.internalPublication","yes"],["dc.type.peerReviewed","yes"],["dspace.entity.type","Publication"]]