54533
2022
2021
eng
14
3
11
article
MDPI
Basel
1
2022-01-25
2021-12-31
--
Partial Disassembly of the Nuclear Pore Complex Proteins during Semi-Closed Mitosis in Dictyostelium discoideum
Dictyostelium cells undergo a semi-closed mitosis, during which the nuclear envelope (NE) persists; however, free diffusion between the cytoplasm and the nucleus takes place. To permit the formation of the mitotic spindle, the nuclear envelope must be permeabilized in order to allow diffusion of tubulin dimers and spindle assembly factors into the nucleus. In Aspergillus, free diffusion of proteins between the cytoplasm and the nucleus is achieved by a partial disassembly of the nuclear pore complexes (NPCs) prior to spindle assembly. In order to determine whether this is also the case in Dictyostelium, we analysed components of the NPC by immunofluorescence microscopy and live cell imaging and studied their behaviour during interphase and mitosis. We observed that the NPCs are absent from the contact area of the nucleoli and that some nucleoporins also localize to the centrosome and the spindle poles. In addition, we could show that, during mitosis, the central FG protein NUP62, two inner ring components and Gle1 depart from the NPCs, while all other tested NUPs remained at the NE. This leads to the conclusion that indeed a partial disassembly of the NPCs takes place, which contributes to permeabilisation of the NE during semi-closed mitosis.
Cells
10.3390/cells11030407
2073-4409
35159217
Universität Potsdam
PA 2022_019
2038.77
<a href="https://doi.org/10.25932/publishup-54534">Zweitveröffentlichung in der Schriftenreihe Postprints der Universität Potsdam : Mathematisch-Naturwissenschaftliche Reihe ; 1233</a>
WOS:000754912100001
2661518-6
Meyer, Irene
CC-BY - Namensnennung 4.0 International
Kristina Mitic
Marianne Grafe
Petros Batsios
Irene Meyer
eng
uncontrolled
nuclear pore complex
eng
uncontrolled
nucleoporins
eng
uncontrolled
semi-closed mitosis
eng
uncontrolled
centrosome
eng
uncontrolled
Dictyostelium
Biowissenschaften; Biologie
Institut für Biochemie und Biologie
Referiert
Publikationsfonds der Universität Potsdam
Gold Open-Access
57392
2020
2020
eng
14
8
9
article
MDPI
Basel
1
2020-08-04
2020-08-04
--
In vivo assembly of a Dictyostelium lamin mutant induced by light, mechanical stress, and pH
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.
Cells : open access journal
10.3390/cells9081834
32759812
2073-4409
outputup:dataSource:MDPI:2020
1834
WOS:000564724200001
Graf, R (corresponding author), Univ Potsdam, Dept Cell Biol, Karl Liebknecht Str 24-25, D-14476 Golm, Germany., mgrafe@uni-potsdam.de; phillip.hofmann@outlook.com; <br /> batsios@uni-potsdam.de; irene.meyer@uni-potsdam.de; <br /> rgraef@uni-potsdam.de
Deutsche Forschungsgemeinschaft (DFG) German Research Foundation (DFG); [GR1642/4-3]
Gräf, Ralph
2023-01-09T11:15:31+00:00
sword
importub
filename=package.tar
7b0ae2863e9d32088d36d078ac22b2b1
Gräf, Ralph
CC-BY - Namensnennung 4.0 International
Marianne Grafe
Phillip Hofmann
Petros Batsios
Irene Meyer
Ralph Gräf
eng
uncontrolled
lamin
eng
uncontrolled
NE81
eng
uncontrolled
Dictyostelium
eng
uncontrolled
nuclear envelope
eng
uncontrolled
nuclear lamina
Biowissenschaften; Biologie
Institut für Biochemie und Biologie
Referiert
Import
Gold Open-Access
DOAJ gelistet
55214
2017
2017
eng
119
130
12
96
article
Elsevier
Jena
1
2017-01-12
2017-01-12
--
CP39, CP75 and CP91 are major structural components of the Dictyostelium
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.
European journal of cell biology
10.1016/j.eicb.2017.01.004
28104305
0171-9335
1618-1298
wos:2017
WOS:000412150200004
Meyer, I; Graf, R (reprint author), Univ Potsdam, Inst Biochem & Biol, Dept Cell Biol, Karl Liebknecht Str 24-25,Haus 26, D-14476 Potsdam, Germany., irene.meyer@uni-potsdam.de; rgraef@uni-porsdam.de
2022-06-17T08:19:48+00:00
sword
importub
filename=package.tar
34ca817be97cbd9caebb62bd54a64cd3
false
true
Irene Meyer
Tatjana Peter
Petros Batsios
Oliver Kuhnert
Anne Krueger-Genge
Carl Camurca
Ralph Gräf
eng
uncontrolled
Dictyostelium
eng
uncontrolled
Mitosis
eng
uncontrolled
Microtubules
eng
uncontrolled
Centrosome
eng
uncontrolled
Nucleus
Biowissenschaften; Biologie
Institut für Biochemie und Biologie
Referiert
Import
53310
2018
2018
eng
17
4
7
article
MDPI
Basel
1
2018-04-23
2018-04-23
--
CDK5RAP2 Is an Essential Scaffolding Protein of the Corona of the Dictyostelium Centrosome
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.
Cells
10.3390/cells7040032
29690637
2073-4409
wos:2018
32
WOS:000435179500009
Meyer, I (reprint author), Univ Potsdam, Dept Cell Biol, Karl Liebknecht Str 24-25, D-14476 Potsdam, Germany., valentin.pitzen@uni-potsdam.de; askarzad@uni-potsdam.de; rgraef@uni-potsdam.de; irene.meyer@uni-potsdam.de
University of Potsdam
2022-01-06T09:32:16+00:00
sword
importub
filename=package.tar
232be3c088811ff155ba132c367759d1
Meyer, Irene
false
true
Valentin Pitzen
Sophie Askarzada
Ralph Gräf
Irene Meyer
eng
uncontrolled
centrosome
eng
uncontrolled
centriole
eng
uncontrolled
Dictyostelium
eng
uncontrolled
microtubules
eng
uncontrolled
mitosis
Biowissenschaften; Biologie
Institut für Biochemie und Biologie
Referiert
Import
Gold Open-Access
DOAJ gelistet
52741
2020
2020
eng
14
8
9
article
MDPI
Basel
1
2020-08-04
2020-04-23
--
In vivo assembly of a Dictyostelium lamin mutant induced by light, mechanical stress, and pH
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.
Cells
Universität Potsdam
<a href="https://doi.org/10.25932/publishup-52507">Zweitveröffentlichung in der Schriftenreihe Postprints der Universität Potsdam : Mathematisch-Naturwissenschaftliche Reihe ; 1213</a>
Version of record
false
false
CC-BY - Namensnennung 4.0 International
Marianne Grafe
Phillip Hofmann
Petros Batsios
Irene Meyer
Ralph Gräf
eng
uncontrolled
lamin
eng
uncontrolled
NE81
eng
uncontrolled
Dictyostelium
eng
uncontrolled
nuclear envelope
eng
uncontrolled
nuclear lamina
Biowissenschaften; Biologie
Institut für Biochemie und Biologie
Referiert
Gold Open-Access
42597
2019
2019
eng
17
682
postprint
0
2019-03-13
2019-03-13
--
Supramolecular Structures of the Dictyostelium Lamin NE81
Nuclear lamins are nucleus-specific intermediate filaments (IF) found at the inner nuclear membrane (INM) of the nuclear envelope (NE). Together with nuclear envelope transmembrane proteins, they form the nuclear lamina and are crucial for gene regulation and mechanical robustness of the nucleus and the whole cell. Recently, we characterized Dictyostelium NE81 as an evolutionarily conserved lamin-like protein, both on the sequence and functional level. Here, we show on the structural level that the Dictyostelium NE81 is also capable of assembling into filaments, just as metazoan lamin filament assemblies. Using field-emission scanning electron microscopy, we show that NE81 expressed in Xenopous oocytes forms filamentous structures with an overall appearance highly reminiscent of Xenopus lamin B2. The in vitro assembly properties of recombinant His-tagged NE81 purified from Dictyostelium extracts are very similar to those of metazoan lamins.
Super-resolution stimulated emission depletion (STED) and expansion microscopy (ExM), as well as transmission electron microscopy of negatively stained purified NE81, demonstrated its capability of forming filamentous structures under low-ionic-strength conditions. These results recommend Dictyostelium as a non-mammalian model organism with a well-characterized nuclear envelope involving all relevant protein components known in animal cells.
Potsprint der Universität Potsdam Mathematisch-Naturwissenschaftliche Reihe
10.25932/publishup-42597
urn:nbn:de:kobv:517-opus4-425976
1866-8372
Cells 8 (2019) 2 DOI: 10.3390/cells8020162
162
<a href="http://publishup.uni-potsdam.de/opus4-ubp/frontdoor/index/index/docId/42596">Bibliographieeintrag der Originalveröffentlichung/Quelle</a>
false
false
CC-BY - Namensnennung 4.0 International
Marianne Grafe
Petros Batsios
Irene Meyer
Daria Lisin
Otto Baumann
Martin W. Goldberg
Ralph Gräf
Zweitveröffentlichungen der Universität Potsdam : Mathematisch-Naturwissenschaftliche Reihe
682
eng
uncontrolled
lamin
eng
uncontrolled
NE81
eng
uncontrolled
Dictyostelium
eng
uncontrolled
nuclear envelope
eng
uncontrolled
nuclear lamina
eng
uncontrolled
expansion microscopy
Biowissenschaften; Biologie
open_access
Mathematisch-Naturwissenschaftliche Fakultät
Referiert
Open Access
Universität Potsdam
https://publishup.uni-potsdam.de/files/42597/pmnr682.pdf
42596
2019
2019
eng
17
2
8
article
Molecular Diversity Preservation International
Basel
1
2019-02-16
2019-02-16
--
Supramolecular Structures of the Dictyostelium Lamin NE81
Nuclear lamins are nucleus-specific intermediate filaments (IF) found at the inner nuclear membrane (INM) of the nuclear envelope (NE). Together with nuclear envelope transmembrane proteins, they form the nuclear lamina and are crucial for gene regulation and mechanical robustness of the nucleus and the whole cell. Recently, we characterized Dictyostelium NE81 as an evolutionarily conserved lamin-like protein, both on the sequence and functional level. Here, we show on the structural level that the Dictyostelium NE81 is also capable of assembling into filaments, just as metazoan lamin filament assemblies. Using field-emission scanning electron microscopy, we show that NE81 expressed in Xenopous oocytes forms filamentous structures with an overall appearance highly reminiscent of Xenopus lamin B2. The in vitro assembly properties of recombinant His-tagged NE81 purified from Dictyostelium extracts are very similar to those of metazoan lamins.
Super-resolution stimulated emission depletion (STED) and expansion microscopy (ExM), as well as transmission electron microscopy of negatively stained purified NE81, demonstrated its capability of forming filamentous structures under low-ionic-strength conditions. These results recommend Dictyostelium as a non-mammalian model organism with a well-characterized nuclear envelope involving all relevant protein components known in animal cells.
Cells
10.3390/cells8020162
2073-4409
Universität Potsdam
PA 2019_20
1417.54
162
<a href="http://nbn-resolving.de/urn:nbn:de:kobv:517-opus4-425976">Zweitveröffentlichung in der Schriftenreihe Postprints der Universität Potsdam : Mathematisch-Naturwissenschaftliche Reihe ; 682</a>
CC-BY - Namensnennung 4.0 International
Marianne Grafe
Petros Batsios
Irene Meyer
Daria Lisin
Otto Baumann
Martin W. Goldberg
Ralph Gräf
eng
uncontrolled
lamin
eng
uncontrolled
NE81
eng
uncontrolled
Dictyostelium
eng
uncontrolled
nuclear envelope
eng
uncontrolled
nuclear lamina
eng
uncontrolled
expansion microscopy
Biowissenschaften; Biologie
open_access
Mathematisch-Naturwissenschaftliche Fakultät
Referiert
Publikationsfonds der Universität Potsdam
Open Access
37038
2011
2011
eng
89
96
8
1
22
review
Elsevier
London
1
--
--
--
Functional analyses of lissencephaly-related proteins in Dictyostelium
Lissencephaly is a severe brain developmental disease in human infants, which is usually caused by mutations in either of two genes, LIS1 and DCX. These genes encode proteins interacting with both the microtubule and the actin systems. Here, we review the implications of data on Dictyostelium LIS1 for the elucidation of LIS1 function in higher cells and emphasize the role of LIS1 and nuclear envelope proteins in nuclear positioning, which is also important for coordinated cell migration during neocortical development. Furthermore, for the first time we characterize Dictyostelium DCX, the only bona fide orthologue of human DCX outside the animal kingdom. We show that DCX functionally interacts with LIS1 and that both proteins have a cytoskeleton-independent function in chemotactic signaling during development. Dictyostelium LIS1 is also required for proper attachment of the centrosome to the nucleus and, thus, nuclear positioning, where the association of these two organelles has turned out to be crucial. It involves not only dynein and dynein-associated proteins such as LIS1 but also SUN proteins of the nuclear envelope. Analyses of Dictyostelium SUN1 mutants have underscored the importance of these proteins for the linkage of centrosomes and nuclei and for the maintenance of chromatin integrity. Taken together, we show that Dictyostelium amoebae, which provide a well-established model to study the basic aspects of chemotaxis, cell migration and development, are well suited for the investigation of the molecular and cell biological basis of developmental diseases such as lissencephaly.
Seminars in cell & developmental biology
10.1016/j.semcdb.2010.10.007
1084-9521
wos:2011-2013
WOS:000286590800014
Graf, R (reprint author), Univ Potsdam, Inst Biochem & Biol, Dept Cell Biol, Karl Liebknecht Str 24-25,Haus 26, D-14476 Potsdam, Germany., rgraef@uni-potsdam.de
DFG [GR1642/2-2, GR1642/3-1]
Irene Meyer
Oliver Kuhnert
Ralph Gräf
eng
uncontrolled
Dictyostelium
eng
uncontrolled
Lissencephaly
eng
uncontrolled
LIS1
eng
uncontrolled
DCX
eng
uncontrolled
SUN1
eng
uncontrolled
Centrosome
Institut für Biochemie und Biologie
Referiert
38858
2015
2015
eng
249
256
8
6
94
article
Elsevier
Jena
1
--
--
--
Evolution of centrosomes and the nuclear lamina: Amoebozoan assets
The current eukaryotic tree of life groups most eukaryotes into one of five supergroups, the Opisthokonta, Amoebozoa, Archaeplastida, Excavata and SAR (Stramenopile, Alveolata, Rhizaria). Molecular and comparative morphological analyses revealed that the last eukaryotic common ancestor (LECA) already contained a rather sophisticated equipment of organelles including a mitochondrion, an endomembrane system, a nucleus with a lamina, a microtubule-organizing center (MTOC), and a flagellar apparatus. Recent studies of MTOCs, basal bodies/centrioles, and nuclear envelope organization of organisms in different supergroups have clarified our picture of how the nucleus and MTOCs co-evolved from LECA to extant eukaryotes. In this review we summarize these findings with special emphasis on valuable contributions of research on a lamin-like protein, nuclear envelope proteins, and the MTOC in the amoebozoan model organism Dictyostelium discoideum. (C) 2015 Elsevier GmbH. All rights reserved.
European journal of cell biology
10.1016/j.ejcb.2015.04.004
25952183
0171-9335
1618-1298
wos:2015
WOS:000356906600002
Graf, R (reprint author), Univ Potsdam, Inst Biochem & Biol, Karl Liebknecht Str 24-25, D-14476 Potsdam, Germany., rgraef@uni-potsdam.de
Deutsche Forschungsgemeinschaft [GR1642/4-3]
Ralph Gräf
Petros Batsios
Irene Meyer
eng
uncontrolled
LINC complex
eng
uncontrolled
Sun1
eng
uncontrolled
Nuclear lamina
eng
uncontrolled
Lamin
eng
uncontrolled
Nuclear envelope
eng
uncontrolled
Centrosome
eng
uncontrolled
Basal body
eng
uncontrolled
Centriole
eng
uncontrolled
LEM-domain
Institut für Biochemie und Biologie
Referiert
35953
2012
2012
eng
237
243
7
3
3
article
Landes Bioscience
Austin
1
--
--
--
A lamin in lower eukaryotes?
Lamins are the major components of the nuclear lamina and serve not only as a mechanical support, but are also involved in chromatin organization, epigenetic regulation, transcription and mitotic events. Despite these universal tasks, lamins have so far been found only in metazoans. Yet, recently we have identified Dictyostelium NE81 as the first lamin-like protein in a lower eukaryote. Based on the current knowledge, we draw a model for nuclear envelope organization in Dictyostelium in this Extra View and we review the experimental data that justified this classification. Furthermore we provide unpublished data underscoring the requirement of posttranslational CaaX-box processing for proper protein localization at the nuclear envelope. Sequence comparison of NE81 sequences from four Dictyostelia with bona fide lamins illustrates the evolutional relationship between these proteins. Under certain conditions these usually unicellular social amoebae congregate to form a multicellular body. We propose that the evolution of the lamin-like NE81 went along with the invention of multicellularity.
Nucleus
10.4161/nucl.20149
1949-1034
wos:2011-2013
WOS:000315928100008
Graf, R (reprint author), Univ Potsdam, Dept Cell Biol, Inst Biochem & Biol, Potsdam, Germany., rgraef@uni-potsdam.de
DFG [GR1642/4-1]
Petros Batsios
Tatjana Peter
Otto Baumann
Reimer Stick
Irene Meyer
Ralph Gräf
eng
uncontrolled
dictyostelium
eng
uncontrolled
lamin
eng
uncontrolled
intermediate filament
eng
uncontrolled
centrosome
eng
uncontrolled
nucleus
Institut für Biochemie und Biologie
Referiert
37151
2011
2011
eng
275
287
13
2
68
article
Springer
Basel
1
--
--
--
Analysis of dictyostelium TACC reveals differential interactions with CP224 and unusual dynamics of dictyostelium microtubules
We have localized TACC to the microtubule-nucleating centrosomal corona and to microtubule plus ends. Using RNAi we proved that Dictyostelium TACC promotes microtubule growth during interphase and mitosis. For the first time we show in vivo that both TACC and XMAP215 family proteins can be differentially localized to microtubule plus ends during interphase and mitosis and that TACC is mainly required for recruitment of an XMAP215-family protein to interphase microtubule plus ends but not for recruitment to centrosomes and kinetochores. Moreover, we have now a marker to study dynamics and behavior of microtubule plus ends in living Dictyostelium cells. In a combination of live cell imaging of microtubule plus ends and fluorescence recovery after photobleaching (FRAP) experiments of GFP-alpha-tubulin cells we show that Dictyostelium microtubules are dynamic only in the cell periphery, while they remain stable at the centrosome, which also appears to harbor a dynamic pool of tubulin dimers.
Cellular and molecular life sciences
10.1007/s00018-010-0453-0
1420-682X
wos:2011-2013
WOS:000285970400008
Graf, R (reprint author), Univ Potsdam, Inst Biochem & Biol, Dept Cell Biol, Karl Liebknecht Str 24-25,Haus 26, D-14476 Potsdam, Germany., rgraef@uni-potsdam.de
Deutsche Forschungsgemeinschaft [GR1642/2-2]
Matthias Samereier
Otto Baumann
Irene Meyer
Ralph Gräf
eng
uncontrolled
Dictyostelium
eng
uncontrolled
TACC
eng
uncontrolled
DdCP224
eng
uncontrolled
XMAP215
eng
uncontrolled
Microtubules
eng
uncontrolled
Centrosome
Institut für Biochemie und Biologie
Referiert
37088
2011
2011
eng
2
22
conferenceobject
American Society for Cell Biology
Bethesda
1
--
--
--
Functional characterization of CP148, a novel key component for centrosome integrity in Dictyostelium
Molecular biology of the cell : the official publication of the American Society for Cell Biology
1059-1524
wos:2011-2013
Annual Meeting of the American-Society-for-Cell-Biology (ASCB)
DEC 03-07, 2011
WOS:000305505502008
Denver, CO
Oliver Kuhnert
Otto Baumann
Irene Meyer
Ralph Gräf
Institut für Biochemie und Biologie
Referiert
45538
2016
2016
eng
124
135
12
95
article
Royal Society
Jena
1
--
--
--
CP91 is a component of the Dictyostelium centrosome involved in centrosome biogenesis
The Dictyostelium centrosome is a model for acentriolar centrosomes and it consists of a three-layered core structure surrounded by a corona harboring microtubule nucleation complexes. Its core structure duplicates once per cell cycle at the G2/M transition. Through proteomic analysis of isolated centrosomes we have identified CP91, a 91-kDa coiled coil protein that was localized at the centrosomal core structure. While GFP-CP91 showed almost no mobility in FRAP experiments during interphase, both GFP-CP91 and endogenous CP91 dissociated during mitosis and were absent from spindle poles from late prophase to anaphase. Since this behavior correlates with the disappearance of the central layer upon centrosome duplication, CP91 is a putative component of this layer. When expressed as GFP-fusions, CP91 fragments corresponding to the central coiled coil domain and the preceding N-terminal part (GFP-CP91cc and GFP-CP91N, respectively) also localized to the centrosome but did not show the mitotic redistribution of the full length protein suggesting a regulatory role of the C-terminal domain. Expression of all GFP-fusion proteins suppressed expression of endogenous CP91 and elicited supernumerary centrosomes. This was also very prominent upon depletion of CP91 by RNAi. Additionally, CP91-RNAi cells exhibited heavily increased ploidy due to severe defects in chromosome segregation along with increased cell size and defects in the abscission process during cytokinesis. Our results indicate that CP91 is a central centrosomal core component required for centrosomal integrity, proper centrosome biogenesis and, independently, for abscission during cytokinesis. (c) 2016 Elsevier GmbH. All rights reserved.
European journal of cell biology
10.1016/j.ejcb.2016.03.001
27005924
0171-9335
1618-1298
wos2016:2019
WOS:000374916100002
Graf, R (reprint author), Univ Potsdam, Inst Biochem & Biol, Dept Cell Biol, Karl Liebknecht Str 24-25,Haus 26, D-14476 Potsdam, Germany., rgraef@uni-potsdam.de
Deutsche Forschungsgemeinschaft [GR1642/4-1]
importub
2020-03-22T19:22:02+00:00
filename=package.tar
2a476cfdc97c94b60dbd6a1e87b477c1
Sascha Putzler
Irene Meyer
Ralph Gräf
eng
uncontrolled
Dictyostelium
eng
uncontrolled
Mitosis
eng
uncontrolled
Microtubules
eng
uncontrolled
Centrosome
eng
uncontrolled
Nucleus
Institut für Biochemie und Biologie
Referiert
Import
50530
2019
2019
eng
509
519
11
8-10
63
article
UBC Pr
Bilbao
1
2019-07-09
2019-07-09
--
Nuclear envelope organization in Dictyostelium discoideum
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.
The international journal of developmental biology
10.1387/ijdb.190184rg
31840788
0214-6282
1696-3547
wos:2019
WOS:000503981500018
Batsios, P (reprint author), Univ Potsdam, Dept Cell Biol, Karl Liebknecht Str 24-25, D-14476 Potsdam, Germany.; Graf, R (reprint author), Univ Potsdam, Dept Cell Biol, Inst Biochem & Biol, Karl Liebknecht Str 24-25, D-14476 Potsdam, Germany., batsios@uni-potsdam.de; rgraef@uni-potsdam.de
National Science FoundationNational Science Foundation (NSF) [MCB-1510511]
2021-04-26T13:05:20+00:00
sword
importub
filename=package.tar
3d3a3d0083285304cdc172bb5bea38eb
Batsios, Petros
Graef, Ralph
false
true
CC-BY - Namensnennung 4.0 International
Petros Batsios
Ralph Gräf
Michael P. Koonce
Denis A. Larochelle
Irene Meyer
eng
uncontrolled
nuclear envelop
eng
uncontrolled
Dictyostelium
eng
uncontrolled
lamin
eng
uncontrolled
NET
eng
uncontrolled
centrosome
eng
uncontrolled
centromere
Biowissenschaften; Biologie
Institut für Biochemie und Biologie
Referiert
Import
Gold Open-Access
56748
2021
2021
eng
26
10
10
article
MDPI
Basel
1
2021-10-05
2021-10-05
--
The dictyostelium centrosome
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.
Cells : open access journal
10.3390/cells10102657
34685637
2073-4409
outputup:dataSource:PubMed:2021
2657
WOS:000712350600001
Graef, R (corresponding author), Univ Potsdam, Dept Cell Biol, Karl Liebknecht Str 24-25, D-14476 Potsdam, Germany., rgraef@uni-potsdam.de; mgrafe@uni-potsdam.de; <br /> irene.meyer@uni-potsdam.de; mitic@uni-potsdam.de; <br /> valentin.pitzen@uni-potsdam.de
Deutsche Forschungsgemeinschaft (DFG)German Research Foundation (DFG) [GR1642/9-1, GR1642/11-1, ME3690/2-1]
Gräf, Ralph
2022-11-21T06:16:50+00:00
sword
importub
filename=package.tar
5af7e28ba2818b95ace8f03b63f96f6c
2661518-6
false
true
CC-BY - Namensnennung 4.0 International
Ralph Gräf
Marianne Grafe
Irene Meyer
Kristina Mitic
Valentin Pitzen
eng
uncontrolled
microtubule-organizing center
eng
uncontrolled
microtubule-organization
eng
uncontrolled
centrosome
eng
uncontrolled
Dictyostelium
eng
uncontrolled
mitosis
Biowissenschaften; Biologie
Institut für Biochemie und Biologie
Referiert
Import
Gold Open-Access
DOAJ gelistet
35564
2012
2012
eng
3651
3664
14
21
69
article
Springer
Basel
1
--
--
--
CP55, a novel key component of centrosomal organization in dictyostelium
Dictyostelium centrosomes consist of a layered core structure surrounded by a microtubule-nucleating corona. At the G2/M transition, the corona dissociates and the core structure duplicates, yielding two spindle pole bodies. Finally, in telophase, the spindle poles mature into two new, complete centrosomes. CP55 was identified in a centrosomal proteome analysis. It is a component of the centrosomal core structure, and persists at the centrosome throughout the entire cell cycle. FRAP experiments revealed that during interphase the majority of centrosomal GFP-CP55 is immobile, which indicates a structural task of CP55 at the centrosome. The CP55null mutant is characterized by increased ploidy, a less structured, slightly enlarged corona, and by supernumerary, cytosolic MTOCs, containing only corona proteins and lacking a core structure. Live cell imaging showed that supernumerary MTOCs arise in telophase. Lack of CP55 also caused premature recruitment of the corona organizer CP148 to mitotic spindle poles, already in metaphase instead of telophase. Forces transmitted through astral microtubules may expel prematurely acquired or loosely attached corona fragments into the cytosol, where they act as independent MTOCs. CP55null cells were also impaired in growth, most probably due to difficulties in centrosome splitting during prophase. Furthermore, although they were still capable of phagocytosis, they appeared unable to utilize phagocytosed nutrients. This inability may be attributed to their partially disorganized Golgi apparatus.
Cellular and molecular life sciences
10.1007/s00018-012-1040-3
1420-682X
wos:2011-2013
WOS:000310083100010
Graf, R (reprint author), Univ Potsdam, Inst Biochem & Biol, Dept Cell Biol, Karl Liebknecht Str 24-25,Haus 26, D-14476 Potsdam, Germany., rgraef@uni-potsdam.de
DFG [GR1642/3-1, GR1642/4-1]
Oliver Kuhnert
Otto Baumann
Irene Meyer
Ralph Gräf
eng
uncontrolled
Dictyostelium
eng
uncontrolled
Corona
eng
uncontrolled
Microtubules
eng
uncontrolled
Centrosome
eng
uncontrolled
Nucleus
Institut für Biochemie und Biologie
Referiert
35871
2012
2012
eng
1875
1888
14
11
69
article
Springer
Basel
1
--
--
--
Functional characterization of CP148, a novel key component for centrosome integrity in Dictyostelium
The centrosome consists of a layered core structure surrounded by a microtubule-nucleating corona. A tight linkage through the nuclear envelope connects the cytosolic centrosome with the clustered centromeres within the nuclear matrix. At G2/M the corona dissociates, and the core structure duplicates, yielding two spindle poles. CP148 is a novel coiled coil protein of the centrosomal corona. GFP-CP148 exhibited cell cycle-dependent presence and absence at the centrosome, which correlates with dissociation of the corona in prophase and its reformation in late telophase. During telophase, GFP-CP148 formed cytosolic foci, which coalesced and joined the centrosome. This explains the hypertrophic appearance of the corona upon strong overexpression of GFP-CP148. Depletion of CP148 by RNAi caused virtual loss of the corona and disorganization of interphase microtubules. Surprisingly, formation of the mitotic spindle and astral microtubules was unaffected. Thus, microtubule nucleation complexes associate with centrosomal core components through different means during interphase and mitosis. Furthermore, CP148 RNAi caused dispersal of centromeres and altered Sun1 distribution at the nuclear envelope, suggesting a role of CP148 in the linkage between centrosomes and centromeres. Taken together, CP148 is an essential factor for the formation of the centrosomal corona, which in turn is required for centrosome/centromere linkage.
Cellular and molecular life sciences
10.1007/s00018-011-0904-2
1420-682X
wos:2011-2013
WOS:000304121600009
Graf, R (reprint author), Univ Potsdam, Dept Cell Biol, Inst Biochem & Biol, Karl Liebknecht Str 24-25, D-14476 Potsdam, Germany., rgraef@uni-potsdam.de
DFG [GR1642/3-1, GR1642/4-1]
Oliver Kuhnert
Otto Baumann
Irene Meyer
Ralph Gräf
eng
uncontrolled
Dictyostelium
eng
uncontrolled
Corona
eng
uncontrolled
Microtubules
eng
uncontrolled
Centrosome
eng
uncontrolled
Nucleus
Institut für Biochemie und Biologie
Referiert
36196
2012
2012
eng
360
370
11
2
23
article
American Society for Cell Biology
Bethesda
1
--
--
--
Characterization of NE81, the first lamin-like nucleoskeleton protein in a unicellular organism
Lamins build the nuclear lamina and are required for chromatin organization, gene expression, cell cycle progression, and mechanical stabilization. Despite these universal functions, lamins have so far been found only in metazoans. We have identified protein NE81 in Dictyostelium, which has properties that justify its denomination as a lamin-like protein in a lower eukaryote. This is based on its primary structure, subcellular localization, and regulation during mitosis, and its requirement of the C-terminal CaaX box as a posttranslational processing signal for proper localization. Our knockout and overexpression mutants revealed an important role for NE81 in nuclear integrity, chromatin organization, and mechanical stability of cells. All our results are in agreement with a role for NE81 in formation of a nuclear lamina. This function is corroborated by localization of Dictyostelium NE81 at the nuclear envelope in human cells. The discovery of a lamin-like protein in a unicellular organism is not only intriguing in light of evolution, it may also provide a simple experimental platform for studies of the molecular basis of laminopathies.
Molecular biology of the cell : the official publication of the American Society for Cell Biology
10.1091/mbc.E11-07-0595
1059-1524
wos:2011-2013
WOS:000299108000011
Graf, R (reprint author), Univ Potsdam, Inst Biochem & Biol, Dept Cell Biol, D-14469 Potsdam, Germany., irene.meyer@uni-potsdam.de; rgraef@uni-potsdam.de
Deutsche Forschungsgemeinschaft [GR1642/3-1, GR1642/4-1]
Anne Krüger
Petros Batsios
Otto Baumann
Eva Luckert
Heinz Schwarz
Reimer Stick
Irene Meyer
Ralph Gräf
Institut für Biochemie und Biologie
Referiert
54534
2022
2022
eng
16
3
postprint
1
2022-03-29
2022-03-29
--
Partial Disassembly of the Nuclear Pore Complex Proteins during Semi-Closed Mitosis in Dictyostelium discoideum
Dictyostelium cells undergo a semi-closed mitosis, during which the nuclear envelope (NE) persists; however, free diffusion between the cytoplasm and the nucleus takes place. To permit the formation of the mitotic spindle, the nuclear envelope must be permeabilized in order to allow diffusion of tubulin dimers and spindle assembly factors into the nucleus. In Aspergillus, free diffusion of proteins between the cytoplasm and the nucleus is achieved by a partial disassembly of the nuclear pore complexes (NPCs) prior to spindle assembly. In order to determine whether this is also the case in Dictyostelium, we analysed components of the NPC by immunofluorescence microscopy and live cell imaging and studied their behaviour during interphase and mitosis. We observed that the NPCs are absent from the contact area of the nucleoli and that some nucleoporins also localize to the centrosome and the spindle poles. In addition, we could show that, during mitosis, the central FG protein NUP62, two inner ring components and Gle1 depart from the NPCs, while all other tested NUPs remained at the NE. This leads to the conclusion that indeed a partial disassembly of the NPCs takes place, which contributes to permeabilisation of the NE during semi-closed mitosis.
Postprints der Universität Potsdam : Mathematisch-Naturwissenschaftliche Reihe
10.25932/publishup-54534
urn:nbn:de:kobv:517-opus4-545341
1866-8372
Cells 11 (2022) 3, 407 DOI: 10.3390/cells11030407
<a href="http://publishup.uni-potsdam.de/54533">Bibliographieeintrag der Originalveröffentlichung/Quelle</a>
true
true
CC-BY - Namensnennung 4.0 International
Kristina Mitic
Marianne Grafe
Petros Batsios
Irene Meyer
Zweitveröffentlichungen der Universität Potsdam : Mathematisch-Naturwissenschaftliche Reihe
1233
eng
uncontrolled
nuclear pore complex
eng
uncontrolled
nucleoporins
eng
uncontrolled
semi-closed mitosis
eng
uncontrolled
centrosome
eng
uncontrolled
Dictyostelium
Biowissenschaften; Biologie
open_access
Institut für Biochemie und Biologie
Referiert
Green Open-Access
Universität Potsdam
https://publishup.uni-potsdam.de/files/54534/pmnr1233.pdf
52507
2020
2020
eng
16
8
postprint
1
2020-08-04
2020-04-23
--
In vivo assembly of a Dictyostelium lamin mutant induced by light, mechanical stress, and pH
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.
Postprints der Universität Potsdam : Mathematisch-Naturwissenschaftliche Reihe
10.25932/publishup-52507
urn:nbn:de:kobv:517-opus4-525075
1866-8372
<a href="http://publishup.uni-potsdam.de/52741">Bibliographieeintrag der Originalveröffentlichung/Quelle</a>
1834
Version of record
Cells 2020, 9(8), 1834; https://doi.org/10.3390/cells9081834
true
true
CC-BY - Namensnennung 4.0 International
Marianne Grafe
Phillip Hofmann
Petros Batsios
Irene Meyer
Ralph Gräf
Zweitveröffentlichungen der Universität Potsdam : Mathematisch-Naturwissenschaftliche Reihe
1213
eng
uncontrolled
lamin
eng
uncontrolled
NE81
lat
uncontrolled
Dictyostelium
eng
uncontrolled
nuclear envelope
eng
uncontrolled
nuclear lamina
Biowissenschaften; Biologie
open_access
Institut für Biochemie und Biologie
Referiert
Green Open-Access
Universität Potsdam
https://publishup.uni-potsdam.de/files/52507/pmnr1213.pdf
56288
2021
eng
19
9
10
article
MDPI
Basel
1
--
2021-09-10
--
Cep192, a novel missing link between the centrosomal core and corona in Dictyostelium amoebae
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.
Cells : open access journal
10.3390/cells10092384
34572033
2073-4409
outputup:dataSource:PubMed:2021
2384
WOS:000699256700001
Meyer, I (corresponding author), Univ Potsdam, Dept Cell Biol, Karl Liebknecht Str 24-25, D-14476 Potsdam, Germany., valentin.pitzen@uni-potsdam.de; s.sander88@gmail.com; <br /> obaumann@uni-potsdam.de; rgraef@uni-potsdam.de; <br /> irene.meyer@uni-potsdam.de
Deutsche ForschungsgemeinschaftGerman Research Foundation (DFG) [ME3690/2-1]; Deutsche Forschungsgemeinschaft (DFG)German Research Foundation (DFG) [INST 336/114-1 FUGG]
Meyer, Irene
2022-10-13T11:57:12+00:00
sword
importub
filename=package.tar
93bc8c5a48b63cf4a46a4ba5562fc002
2661518-6
CC-BY - Namensnennung 4.0 International
Valentin Pitzen
Sophia Sander
Otto Baumann
Ralph Gräf
Irene Meyer
eng
uncontrolled
Cep192
eng
uncontrolled
SPD-2
eng
uncontrolled
centrosome
eng
uncontrolled
Dictyostelium
eng
uncontrolled
microtubule-organization
eng
uncontrolled
MTOC
Biowissenschaften; Biologie
Institut für Biochemie und Biologie
Referiert
Import
Gold Open-Access
DOAJ gelistet