@article{SchweigelBatsiosMuellerTaubenbergeretal.2022,
  author    = {Schweigel, Ulrike and Batsios, Petros and M{\"u}ller-Taubenberger, Annette and Gr{\"a}f, Ralph and Grafe, Marianne},
  title     = {Dictyostelium spastin is involved in nuclear envelope dynamics during semi-closed mitosis},
  series = {Nucleus},
  volume    = {13},
  journal   = {Nucleus},
  number    = {1},
  publisher = {Taylor \& Francis Group},
  address   = {Philadelphia},
  issn      = {1949-1034},
  doi       = {10.1080/19491034.2022.2047289},
  pages     = {144 -- 154},
  year      = {2022},
  abstract  = {Dictyostelium amoebae perform a semi-closed mitosis, in which the nuclear envelope is fenestrated at the insertion sites of the mitotic centrosomes and around the central spindle during karyokinesis. During late telophase the centrosome relocates to the cytoplasmic side of the nucleus, the central spindle disassembles and the nuclear fenestrae become closed. Our data indicate that Dictyostelium spastin (DdSpastin) is a microtubule-binding and severing type I membrane protein that plays a role in this process. Its mitotic localization is in agreement with a requirement for the removal of microtubules that would hinder closure of the fenestrae. Furthermore, DdSpastin interacts with the HeH/ LEM-family protein Src1 in BioID analyses as well as the inner nuclear membrane protein Sun1, and shows subcellular co-localizations with Src1, Sun1, the ESCRT component CHMP7 and the IST1-like protein filactin, suggesting that the principal pathway of mitotic nuclear envelope remodeling is conserved between animals and Dictyostelium amoebae.},
  language  = {en}
}
@article{KoonceTikhonenkoGraef2020,
  author    = {Koonce, Michael and Tikhonenko, Irina and Gr{\"a}f, Ralph},
  title     = {Dictyostelium cell fixation},
  series = {Methods and protocols},
  volume    = {3},
  journal   = {Methods and protocols},
  number    = {3},
  publisher = {MDPI},
  address   = {Basel},
  issn      = {2409-9279},
  doi       = {10.3390/mps3030047},
  pages     = {6},
  year      = {2020},
  abstract  = {We share two simple modifications to enhance the fixation and imaging of relatively small, motile, and rounded model cells. These include cell centrifugation and the addition of trace amounts of glutaraldehyde to existing fixation methods. Though they need to be carefully considered in each context, they have been useful to our studies of the spatial relationships of the microtubule cytoskeletal system.},
  language  = {en}
}
@article{GrafeHofmannBatsiosetal.2020,
  author    = {Grafe, Marianne and Hofmann, Phillip and Batsios, Petros and Meyer, Irene and Gr{\"a}f, Ralph},
  title     = {In vivo assembly of a Dictyostelium lamin mutant induced by light, mechanical stress, and pH},
  series = {Cells : open access journal},
  volume    = {9},
  journal   = {Cells : open access journal},
  number    = {8},
  publisher = {MDPI},
  address   = {Basel},
  issn      = {2073-4409},
  doi       = {10.3390/cells9081834},
  pages     = {14},
  year      = {2020},
  abstract  = {We expressedDictyosteliumlamin (NE81) lacking both a functional nuclear localization signal and a CAAX-box for C-terminal lipid modification. This lamin mutant assembled into supramolecular, three-dimensional clusters in the cytosol that disassembled at the onset of mitosis and re-assembled in late telophase, thus mimicking the behavior of the endogenous protein. As disassembly is regulated by CDK1-mediated phosphorylation at serine 122, we generated a phosphomimetic S122E mutant called GFP-NE81-S122E-Delta NLS Delta CLIM. Surprisingly, during imaging, the fusion protein assembled into cytosolic clusters, similar to the protein lacking the phosphomimetic mutation. Clusters disassembled again in the darkness. Assembly could be induced with blue but not green or near ultraviolet light, and it was independent of the fusion tag. Assembly similarly occurred upon cell flattening. Earlier reports and own observations suggested that both blue light and cell flattening could result in a decrease of intracellular pH. Indeed, keeping the cells at low pH also reversibly induced cluster formation. Our results indicate that lamin assembly can be induced by various stress factors and that these are transduced via intracellular acidification. Although these effects have been shown in a phosphomimetic CDK1 mutant of theDictyosteliumlamin, they are likely relevant also for wild-type lamin.},
  language  = {en}
}
@article{GraefGrafeMeyeretal.2021,
  author    = {Gr{\"a}f, Ralph and Grafe, Marianne and Meyer, Irene and Mitic, Kristina and Pitzen, Valentin},
  title     = {The dictyostelium centrosome},
  series = {Cells : open access journal},
  volume    = {10},
  journal   = {Cells : open access journal},
  number    = {10},
  publisher = {MDPI},
  address   = {Basel},
  issn      = {2073-4409},
  doi       = {10.3390/cells10102657},
  pages     = {26},
  year      = {2021},
  abstract  = {The centrosome of Dictyostelium amoebae contains no centrioles and consists of a cylindrical layered core structure surrounded by a corona harboring microtubule-nucleating gamma-tubulin complexes. It is the major centrosomal model beyond animals and yeasts. Proteomics, protein interaction studies by BioID and superresolution microscopy methods led to considerable progress in our understanding of the composition, structure and function of this centrosome type. We discuss all currently known components of the Dictyostelium centrosome in comparison to other centrosomes of animals and yeasts.},
  language  = {en}
}
@article{PitzenSanderBaumannetal.2021,
  author    = {Pitzen, Valentin and Sander, Sophia and Baumann, Otto and Gr{\"a}f, Ralph and Meyer, Irene},
  title     = {Cep192, a novel missing link between the centrosomal core and corona in Dictyostelium amoebae},
  series = {Cells : open access journal},
  volume    = {10},
  journal   = {Cells : open access journal},
  number    = {9},
  publisher = {MDPI},
  address   = {Basel},
  issn      = {2073-4409},
  doi       = {10.3390/cells10092384},
  pages     = {19},
  year      = {2021},
  abstract  = {The Dictyostelium centrosome is a nucleus-associated body with a diameter of approx. 500 nm. It contains no centrioles but consists of a cylindrical layered core structure surrounded by a microtubule-nucleating corona. At the onset of mitosis, the corona disassembles and the core structure duplicates through growth, splitting, and reorganization of the outer core layers. During the last decades our research group has characterized the majority of the 42 known centrosomal proteins. In this work we focus on the conserved, previously uncharacterized Cep192 protein. We use superresolution expansion microscopy (ExM) to show that Cep192 is a component of the outer core layers. Furthermore, ExM with centrosomal marker proteins nicely mirrored all ultrastructurally known centrosomal substructures. Furthermore, we improved the proximity-dependent biotin identification assay (BioID) by adapting the biotinylase BioID2 for expression in Dictyostelium and applying a knock-in strategy for the expression of BioID2-tagged centrosomal fusion proteins. Thus, we were able to identify various centrosomal Cep192 interaction partners, including CDK5RAP2, which was previously allocated to the inner corona structure, and several core components. Studies employing overexpression of GFP-Cep192 as well as depletion of endogenous Cep192 revealed that Cep192 is a key protein for the recruitment of corona components during centrosome biogenesis and is required to maintain a stable corona structure.},
  language  = {en}
}
@article{MeyerPeterBatsiosetal.2017,
  author    = {Meyer, Irene and Peter, Tatjana and Batsios, Petros and Kuhnert, Oliver and Krueger-Genge, Anne and Camurca, Carl and Gr{\"a}f, Ralph},
  title     = {CP39, CP75 and CP91 are major structural components of the Dictyostelium},
  series = {European journal of cell biology},
  volume    = {96},
  journal   = {European journal of cell biology},
  publisher = {Elsevier},
  address   = {Jena},
  issn      = {0171-9335},
  doi       = {10.1016/j.eicb.2017.01.004},
  pages     = {119 -- 130},
  year      = {2017},
  abstract  = {The acentriolar Dictyostelium centrosome is a nucleus-associated body consisting of a core structure with three plaque-like layers, which are surrounded by a microtubule-nucleating corona. The core duplicates once per cell cycle at the G2/M transition, whereby its central layer disappears and the two outer layers form the mitotic spindle poles. Through proteomic analysis of isolated centrosomes, we have identified CP39 and CP75, two essential components of the core structure. Both proteins can be assigned to the central core layer as their centrosomal presence is correlated to the disappearance and reappearance of the central core layer in the course of centrosome duplication. Both proteins contain domains with centrosome-binding activity in their N- and C-terminal halves, whereby the respective N-terminal half is required for cell cycle-dependent regulation. CP39 is capable of self-interaction and GFP-CP39 overexpression elicited supernumerary microtubule-organizing centers and pre-centrosomal cytosolic clusters. Underexpression stopped cell growth and reversed the MTOC amplification phenotype. In contrast, in case of CP75 underexpression of the protein by RNAi treatment elicited supernumerary MTOCs. In addition, CP75RNAi affects correct chromosome segregation and causes co-depletion of CP39 and CP91, another central core layer component. CP39 and CP75 interact with each other directly in a yeast two-hybrid assay. Furthermore, CP39, CP75 and CP91 mutually interact in a proximity-dependent biotin identification (BioID) assay. Our data indicate that these three proteins are all required for proper centrosome biogenesis and make up the major structural components of core structure's central layer.},
  language  = {en}
}
@article{PitzenAskarzadaGraefetal.2018,
  author    = {Pitzen, Valentin and Askarzada, Sophie and Gr{\"a}f, Ralph and Meyer, Irene},
  title     = {CDK5RAP2 Is an Essential Scaffolding Protein of the Corona of the Dictyostelium Centrosome},
  series = {Cells},
  volume    = {7},
  journal   = {Cells},
  number    = {4},
  publisher = {MDPI},
  address   = {Basel},
  issn      = {2073-4409},
  doi       = {10.3390/cells7040032},
  pages     = {17},
  year      = {2018},
  abstract  = {Dictyostelium centrosomes consist of a nucleus-associated cylindrical, three-layered core structure surrounded by a corona consisting of microtubule-nucleation complexes embedded in a scaffold of large coiled-coil proteins. One of them is the conserved CDK5RAP2 protein. Here we focus on the role of Dictyostelium CDK5RAP2 for maintenance of centrosome integrity, its interaction partners and its dynamic behavior during interphase and mitosis. GFP-CDK5RAP2 is present at the centrosome during the entire cell cycle except from a short period during prophase, correlating with the normal dissociation of the corona at this stage. RNAi depletion of CDK5RAP2 results in complete disorganization of centrosomes and microtubules suggesting that CDK5RAP2 is required for organization of the corona and its association to the core structure. This is in line with the observation that overexpressed GFP-CDK5RAP2 elicited supernumerary cytosolic MTOCs. The phenotype of CDK5RAP2 depletion was very reminiscent of that observed upon depletion of CP148, another scaffolding protein of the corona. BioID interaction assays revealed an interaction of CDK5RAP2 not only with the corona markers CP148, gamma-tubulin, and CP248, but also with the core components Cep192, CP75, and CP91. Furthermore, protein localization studies in both depletion strains revealed that CP148 and CDK5RAP2 cooperate in corona organization.},
  language  = {en}
}
@article{GrafeHofmannBatsiosetal.2020,
  author    = {Grafe, Marianne and Hofmann, Phillip and Batsios, Petros and Meyer, Irene and Gr{\"a}f, Ralph},
  title     = {In vivo assembly of a Dictyostelium lamin mutant induced by light, mechanical stress, and pH},
  series = {Cells},
  volume    = {9},
  journal   = {Cells},
  number    = {8},
  publisher = {MDPI},
  address   = {Basel},
  pages     = {14},
  year      = {2020},
  abstract  = {We expressed Dictyostelium lamin (NE81) lacking both a functional nuclear localization signal and a CAAX-box for C-terminal lipid modification. This lamin mutant assembled into supramolecular, three-dimensional clusters in the cytosol that disassembled at the onset of mitosis and re-assembled in late telophase, thus mimicking the behavior of the endogenous protein. As disassembly is regulated by CDK1-mediated phosphorylation at serine 122, we generated a phosphomimetic S122E mutant called GFP-NE81-S122E-∆NLS∆CLIM. Surprisingly, during imaging, the fusion protein assembled into cytosolic clusters, similar to the protein lacking the phosphomimetic mutation. Clusters disassembled again in the darkness. Assembly could be induced with blue but not green or near ultraviolet light, and it was independent of the fusion tag. Assembly similarly occurred upon cell flattening. Earlier reports and own observations suggested that both blue light and cell flattening could result in a decrease of intracellular pH. Indeed, keeping the cells at low pH also reversibly induced cluster formation. Our results indicate that lamin assembly can be induced by various stress factors and that these are transduced via intracellular acidification. Although these effects have been shown in a phosphomimetic CDK1 mutant of the Dictyostelium lamin, they are likely relevant also for wild-type lamin.},
  language  = {en}
}
@article{MaiWolskiPuciulMalinowskaetal.2018,
  author    = {Mai, Tobias and Wolski, Karol and Puciul-Malinowska, Agnieszka and Kopyshev, Alexey and Gr{\"a}f, Ralph and Bruns, Michael and Zapotoczny, Szczepan and Taubert, Andreas},
  title     = {Anionic polymer brushes for biomimetic calcium phosphate mineralization},
  series = {Polymers},
  volume    = {10},
  journal   = {Polymers},
  number    = {10},
  publisher = {MDPI},
  address   = {Basel},
  issn      = {2073-4360},
  doi       = {10.3390/polym10101165},
  pages     = {17},
  year      = {2018},
  abstract  = {This article describes the synthesis of anionic polymer brushes and their mineralization with calcium phosphate. The brushes are based on poly(3-sulfopropyl methacrylate potassium salt) providing a highly charged polymer brush surface. Homogeneous brushes with reproducible thicknesses are obtained via surface-initiated atom transfer radical polymerization. Mineralization with doubly concentrated simulated body fluid yields polymer/inorganic hybrid films containing AB-Type carbonated hydroxyapatite (CHAP), a material resembling the inorganic component of bone. Moreover, growth experiments using Dictyostelium discoideum amoebae demonstrate that the mineral-free and the mineral-containing polymer brushes have a good biocompatibility suggesting their use as biocompatible surfaces in implantology or related fields.},
  language  = {en}
}
@article{BatsiosGraefKoonceetal.2019,
  author    = {Batsios, Petros and Gr{\"a}f, Ralph and Koonce, Michael P. and Larochelle, Denis A. and Meyer, Irene},
  title     = {Nuclear envelope organization in Dictyostelium discoideum},
  series = {The international journal of developmental biology},
  volume    = {63},
  journal   = {The international journal of developmental biology},
  number    = {8-10},
  publisher = {UBC Pr},
  address   = {Bilbao},
  issn      = {0214-6282},
  doi       = {10.1387/ijdb.190184rg},
  pages     = {509 -- 519},
  year      = {2019},
  abstract  = {The nuclear envelope consists of the outer and the inner nuclear membrane, the nuclear lamina and the nuclear pore complexes, which regulate nuclear import and export.The major constituent of the nuclear lamina of Dictyostelium is the lamin NE81. It can form filaments like B-type lamins and it interacts with Sun 1, as well as with the LEM/HeH-family protein Src1. Sun 1 and Src1 are nuclear envelope transmembrane proteins involved in the centrosome-nucleus connection and nuclear envelope stability at the nucleolar regions, respectively. In conjunction with a KASH-domain protein, Sun 1 usually forms a so-called LINC complex.Two proteins with functions reminiscent of KASH-domain proteins at the outer nuclear membrane of Dictyostelium are known; interaptin which serves as an actin connector and the kinesin Kif9 which plays a role in the microtubule-centrosome connector. However, both of these lack the conserved KASH-domain. The link of the centrosome to the nuclear envelope is essential for the insertion of the centrosome into the nuclear envelope and the appropriate spindle formation. Moreover, centrosome insertion is involved in perm eabilization of the mitotic nucleus, which ensures access of tubulin dimers and spindle assembly factors. Our recent progress in identifying key molecular players at the nuclear envelope of Dictyostelium promises further insights into the mechanisms of nuclear envelope dynamics.},
  language  = {en}
}