2024-03-28T14:19:23Zhttp://repository.helmholtz-hzi.de/oai/requestoai:repository.helmholtz-hzi.de:10033/141222019-08-30T11:33:57Zcom_10033_6807com_10033_6799col_10033_6884
Steffen, Anika
b99fd7c905e8178bcafe15b15aa53ec6
500
Faix, Jan
3a94710daa2c4770050204bff0b1c340
500
Resch, Guenter P
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500
Linkner, Joern
33d47b5dff855e8473faa00bfafe102a
500
Wehland, Juergen
2657a52b0963f15ed66cd386dc4cd124
500
Small, J Victor
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500
Rottner, Klemens
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Stradal, Theresia E B
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2007-10-17T11:25:31Z
2007-10-17T11:25:31Z
2006-06-01
Mol. Biol. Cell 2006, 17(6):2581-91
1059-1524
16597702
10.1091/mbc.E05-11-1088
http://hdl.handle.net/10033/14122
Cell migration is initiated by plasma membrane protrusions, in the form of lamellipodia and filopodia. The latter rod-like projections may exert sensory functions and are found in organisms as distant in evolution as mammals and amoeba such as Dictyostelium discoideum. In mammals, lamellipodia protrusion downstream of the small GTPase Rac1 requires a multimeric protein assembly, the WAVE-complex, which activates Arp2/3-mediated actin filament nucleation and actin network assembly. A current model of filopodia formation postulates that these structures arise from a dendritic network of lamellipodial actin filaments by selective elongation and bundling. Here, we have analyzed filopodia formation in mammalian cells abrogated in expression of essential components of the lamellipodial actin polymerization machinery. Cells depleted of the WAVE-complex component Nck-associated protein 1 (Nap1), and, in consequence, of lamellipodia, exhibited normal filopodia protrusion. Likewise, the Arp2/3-complex, which is essential for lamellipodia protrusion, is dispensable for filopodia formation. Moreover, genetic disruption of nap1 or the WAVE-orthologue suppressor of cAMP receptor (scar) in Dictyostelium was also ineffective in preventing filopodia protrusion. These data suggest that the molecular mechanism of filopodia formation is conserved throughout evolution from Dictyostelium to mammals and show that lamellipodia and filopodia formation are functionally separable.
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Filopodia formation in the absence of functional WAVE- and Arp2/3-complexes.
Article
2018-06-12T23:54:41Z
Cell migration is initiated by plasma membrane protrusions, in the form of lamellipodia and filopodia. The latter rod-like projections may exert sensory functions and are found in organisms as distant in evolution as mammals and amoeba such as Dictyostelium discoideum. In mammals, lamellipodia protrusion downstream of the small GTPase Rac1 requires a multimeric protein assembly, the WAVE-complex, which activates Arp2/3-mediated actin filament nucleation and actin network assembly. A current model of filopodia formation postulates that these structures arise from a dendritic network of lamellipodial actin filaments by selective elongation and bundling. Here, we have analyzed filopodia formation in mammalian cells abrogated in expression of essential components of the lamellipodial actin polymerization machinery. Cells depleted of the WAVE-complex component Nck-associated protein 1 (Nap1), and, in consequence, of lamellipodia, exhibited normal filopodia protrusion. Likewise, the Arp2/3-complex, which is essential for lamellipodia protrusion, is dispensable for filopodia formation. Moreover, genetic disruption of nap1 or the WAVE-orthologue suppressor of cAMP receptor (scar) in Dictyostelium was also ineffective in preventing filopodia protrusion. These data suggest that the molecular mechanism of filopodia formation is conserved throughout evolution from Dictyostelium to mammals and show that lamellipodia and filopodia formation are functionally separable.
ORIGINAL
Steffen et al.pdf
Steffen et al.pdf
original document
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oai:hzi.openrepository.com:10033/14122
2019-08-30 11:33:57.272
Helmholtz Zentrum für Infektionsforschung Repository
hzi@openrepository.com
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oai:repository.helmholtz-hzi.de:10033/181552019-08-30T11:36:04Zcom_10033_6807com_10033_6799col_10033_6884
Chmielowiec, Jolanta
16fcef17ec3fb6e96ed5a15fee3cf8f9
500
Borowiak, Malgorzata
e114a1f6c7baff026345637fc2690d87
500
Morkel, Markus
4aa64050799c8b28eeea93bc1bde29e3
500
Stradal, Theresia
519b5828df98bb932437f00cdf87244d
500
Munz, Barbara
5977f35dfc866650d761a89070c34eea
500
Werner, Sabine
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500
Wehland, Jürgen
3b0b73bd5f49e3aae0a84586e2cc78db
500
Birchmeier, Carmen
587411739a186e9059ca71df04fb5879
500
Birchmeier, Walter
5d7aaaab4b5bbe74789a648df1090af4
500
Department of Cancer Biology, Max-Delbrück-Center for Molecular Medicine, 13125 Berlin, Germany.
2008-02-13T14:22:33Z
2008-02-13T14:22:33Z
2007-04-09
c-Met is essential for wound healing in the skin. 2007, 177 (1):151-62 J. Cell Biol.
0021-9525
17403932
10.1083/jcb.200701086
http://hdl.handle.net/10033/18155
The Journal of cell biology
Wound healing of the skin is a crucial regenerative process in adult mammals. We examined wound healing in conditional mutant mice, in which the c-Met gene that encodes the receptor of hepatocyte growth factor/scatter factor was mutated in the epidermis by cre recombinase. c-Met-deficient keratinocytes were unable to contribute to the reepithelialization of skin wounds. In conditional c-Met mutant mice, wound closure was slightly attenuated, but occurred exclusively by a few (5%) keratinocytes that had escaped recombination. This demonstrates that the wound process selected and amplified residual cells that express a functional c-Met receptor. We also cultured primary keratinocytes from the skin of conditional c-Met mutant mice and examined them in scratch wound assays. Again, closure of scratch wounds occurred by the few remaining c-Met-positive cells. Our data show that c-Met signaling not only controls cell growth and migration during embryogenesis but is also essential for the generation of the hyperproliferative epithelium in skin wounds, and thus for a fundamental regenerative process in the adult.
en
Animals
Autocrine Communication
Cells, Cultured
Hepatocyte Growth Factor
Integrases
Keratinocytes
Mice
Mice, Inbred Strains
Mutation
Proto-Oncogene Proteins c-met
Signal Transduction
Skin Physiology
Wound Healing
c-Met is essential for wound healing in the skin.
Article
2018-06-12T23:05:20Z
Wound healing of the skin is a crucial regenerative process in adult mammals. We examined wound healing in conditional mutant mice, in which the c-Met gene that encodes the receptor of hepatocyte growth factor/scatter factor was mutated in the epidermis by cre recombinase. c-Met-deficient keratinocytes were unable to contribute to the reepithelialization of skin wounds. In conditional c-Met mutant mice, wound closure was slightly attenuated, but occurred exclusively by a few (5%) keratinocytes that had escaped recombination. This demonstrates that the wound process selected and amplified residual cells that express a functional c-Met receptor. We also cultured primary keratinocytes from the skin of conditional c-Met mutant mice and examined them in scratch wound assays. Again, closure of scratch wounds occurred by the few remaining c-Met-positive cells. Our data show that c-Met signaling not only controls cell growth and migration during embryogenesis but is also essential for the generation of the hyperproliferative epithelium in skin wounds, and thus for a fundamental regenerative process in the adult.
ORIGINAL
Chmielowiec et al_final.pdf
Chmielowiec et al_final.pdf
original publication
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10033/18155
oai:hzi.openrepository.com:10033/18155
2019-08-30 11:36:04.899
Helmholtz Zentrum für Infektionsforschung Repository
hzi@openrepository.com
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oai:repository.helmholtz-hzi.de:10033/478092019-08-30T11:31:23Zcom_10033_6807com_10033_6799col_10033_6884
Derivery, Emmanuel
5d12857ab4be85e0dd809c4034df0134
500
Fink, Jenny
7249c33fb68fb7ebc675bd76e5ad964a
500
Martin, Davy
fcd0d5c3e689dea705d347de61bb44a6
500
Houdusse, Anne
ee5aed2372945b3df3469060eff3acfc
500
Piel, Matthieu
59bf8302816b0eb7e4bc327dd297bf55
500
Stradal, Theresia E
58958e12d8699368d0442aad9238fb13
500
Louvard, Daniel
b18ae80515672747d9f38ef53e745011
500
Gautreau, Alexis
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500
Institut Curie, Centre de Recherche, Morphogenesis and Cell Signaling laboratory, Paris, France.
2009-01-21T15:28:19Z
2009-01-21T15:28:19Z
2008
Free Brick1 is a trimeric precursor in the assembly of a functional wave complex. 2008, 3 (6):e2462 PLoS ONE
1932-6203
18560548
10.1371/journal.pone.0002462
http://hdl.handle.net/10033/47809
PLoS ONE
BACKGROUND: The Wave complex activates the Arp2/3 complex, inducing actin polymerization in lamellipodia and membrane ruffles. The Wave complex is composed of five subunits, the smallest of which, Brick1/Hspc300 (Brk1), is the least characterized. We previously reported that, unlike the other subunits, Brk1 also exists as a free form. PRINCIPAL FINDINGS: Here we report that this free form of Brk1 is composed of homotrimers. Using a novel assay in which purified free Brk1 is electroporated into HeLa cells, we were able to follow its biochemical fate in cells and to show that free Brk1 becomes incorporated into the Wave complex. Importantly, incorporation of free Brk1 into the Wave complex was blocked upon inhibition of protein synthesis and incorporated Brk1 was found to associate preferentially with neosynthesized subunits. Brk1 depleted HeLa cells were found to bleb, as were Nap1, Wave2 or ARPC2 depleted cells, suggesting that this blebbing phenotype of Brk1 depleted cells is due to an impairment of the Wave complex function rather than a specific function of free Brk1. Blebs of Brk1 depleted cells were emitted at sites where lamellipodia and membrane ruffles were normally emitted. In Brk1 depleted cells, the electroporation of free Brk1 was sufficient to restore Wave complex assembly and to rescue the blebbing phenotype. CONCLUSION: Together these results establish that the free form of Brk1 is an essential precursor in the assembly of a functional Wave complex.
en
http://www.plosone.org/article/info:doi/10.1371/journal.pone.0002462
Amino Acid Sequence
Biopolymers
Cell Line
Cytoskeletal Proteins
Electrophoresis, Polyacrylamide Gel
Electroporation
Humans
Immunoprecipitation
Molecular Sequence Data
Transfection
Free Brick1 is a trimeric precursor in the assembly of a functional wave complex.
Article
2018-06-12T22:02:10Z
BACKGROUND: The Wave complex activates the Arp2/3 complex, inducing actin polymerization in lamellipodia and membrane ruffles. The Wave complex is composed of five subunits, the smallest of which, Brick1/Hspc300 (Brk1), is the least characterized. We previously reported that, unlike the other subunits, Brk1 also exists as a free form. PRINCIPAL FINDINGS: Here we report that this free form of Brk1 is composed of homotrimers. Using a novel assay in which purified free Brk1 is electroporated into HeLa cells, we were able to follow its biochemical fate in cells and to show that free Brk1 becomes incorporated into the Wave complex. Importantly, incorporation of free Brk1 into the Wave complex was blocked upon inhibition of protein synthesis and incorporated Brk1 was found to associate preferentially with neosynthesized subunits. Brk1 depleted HeLa cells were found to bleb, as were Nap1, Wave2 or ARPC2 depleted cells, suggesting that this blebbing phenotype of Brk1 depleted cells is due to an impairment of the Wave complex function rather than a specific function of free Brk1. Blebs of Brk1 depleted cells were emitted at sites where lamellipodia and membrane ruffles were normally emitted. In Brk1 depleted cells, the electroporation of free Brk1 was sufficient to restore Wave complex assembly and to rescue the blebbing phenotype. CONCLUSION: Together these results establish that the free form of Brk1 is an essential precursor in the assembly of a functional Wave complex.
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Helmholtz Zentrum für Infektionsforschung Repository
hzi@openrepository.com
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oai:repository.helmholtz-hzi.de:10033/1068892019-08-30T11:35:39Zcom_10033_6807com_10033_6799col_10033_6884
Tahirovic, Sabina
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500
Hellal, Farida
1ede5b9eb35bdbab2bcb40ebd4a67dbe
500
Neukirchen, Dorothee
922ad9d1ce622b05fb4bf805d01996ec
500
Hindges, Robert
1e49a87e73b30cf449735f189f0143aa
500
Garvalov, Boyan K
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500
Flynn, Kevin C
fcc823d6821e9947d6206af44c897a5b
500
Stradal, Theresia E
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500
Chrostek-Grashoff, Anna
ea9229bbf9c685205c2a3e90a773f250
500
Brakebusch, Cord
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Bradke, Frank
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500
Axonal Growth and Regeneration Group, Max Planck Institute of Neurobiology, 82152 Martinsried, Germany.
2010-06-28T08:50:24Z
2010-06-28T08:50:24Z
2010-05-19
Rac1 regulates neuronal polarization through the WAVE complex. 2010, 30 (20):6930-43 J. Neurosci.
1529-2401
20484635
10.1523/JNEUROSCI.5395-09.2010
http://hdl.handle.net/10033/106889
The Journal of neuroscience : the official journal of the Society for Neuroscience
Neuronal migration and axon growth, key events during neuronal development, require distinct changes in the cytoskeleton. Although many molecular regulators of polarity have been identified and characterized, relatively little is known about their physiological role in this process. To study the physiological function of Rac1 in neuronal development, we have generated a conditional knock-out mouse, in which Rac1 is ablated in the whole brain. Rac1-deficient cerebellar granule neurons, which do not express other Rac isoforms, showed impaired neuronal migration and axon formation both in vivo and in vitro. In addition, Rac1 ablation disrupts lamellipodia formation in growth cones. The analysis of Rac1 effectors revealed the absence of the Wiskott-Aldrich syndrome protein (WASP) family verprolin-homologous protein (WAVE) complex from the plasma membrane of knock-out growth cones. Loss of WAVE function inhibited axon growth, whereas overexpression of a membrane-tethered WAVE mutant partially rescued axon growth in Rac1-knock-out neurons. In addition, pharmacological inhibition of the WAVE complex effector Arp2/3 also reduced axon growth. We propose that Rac1 recruits the WAVE complex to the plasma membrane to enable actin remodeling necessary for axon growth.
en
Angiopoietins
Animals
Animals, Newborn
Apoptosis
Axons
Bromodeoxyuridine
Cell Movement
Cell Proliferation
Cells, Cultured
Cerebellum
Cofilin 1
Enzyme Inhibitors
Enzyme-Linked Immunosorbent Assay
Growth Cones
Ki-67 Antigen
Luminescent Proteins
Mice
Mice, Knockout
Mutation
Nerve Tissue Proteins
Neurons
Organ Culture Techniques
RNA Interference
RNA, Small Interfering
Transfection
Wiskott-Aldrich Syndrome Protein Family
cdc42 GTP-Binding Protein
rac1 GTP-Binding Protein
rhoA GTP-Binding Protein
Rac1 regulates neuronal polarization through the WAVE complex.
Article
2010-11-20T00:00:00Z
Neuronal migration and axon growth, key events during neuronal development, require distinct changes in the cytoskeleton. Although many molecular regulators of polarity have been identified and characterized, relatively little is known about their physiological role in this process. To study the physiological function of Rac1 in neuronal development, we have generated a conditional knock-out mouse, in which Rac1 is ablated in the whole brain. Rac1-deficient cerebellar granule neurons, which do not express other Rac isoforms, showed impaired neuronal migration and axon formation both in vivo and in vitro. In addition, Rac1 ablation disrupts lamellipodia formation in growth cones. The analysis of Rac1 effectors revealed the absence of the Wiskott-Aldrich syndrome protein (WASP) family verprolin-homologous protein (WAVE) complex from the plasma membrane of knock-out growth cones. Loss of WAVE function inhibited axon growth, whereas overexpression of a membrane-tethered WAVE mutant partially rescued axon growth in Rac1-knock-out neurons. In addition, pharmacological inhibition of the WAVE complex effector Arp2/3 also reduced axon growth. We propose that Rac1 recruits the WAVE complex to the plasma membrane to enable actin remodeling necessary for axon growth.
ORIGINAL
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2019-08-30 11:35:39.742
Helmholtz Zentrum für Infektionsforschung Repository
hzi@openrepository.com
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oai:repository.helmholtz-hzi.de:10033/1347622019-08-30T11:37:24Zcom_10033_6807com_10033_6799col_10033_6884
Emadi Baygi, Modjtaba
88583d466fb6e7bd6812da8c616b2fd4
500
Soheili, Zahra Soheila
ed875ae3286f0bd42049794ce8990886
500
Schmitz, Ingo
33252c7e3ac40c72ff226b0ae1187c4b
500
Sameie, Shahram
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Schulz, Wolfgang A
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Department of Genetics, Faculty of Biological Sciences, Tarbiat Modares University, Tehran, Iran.
2011-06-28T13:47:38Z
2011-06-28T13:47:38Z
2010-12
Snail regulates cell survival and inhibits cellular senescence in human metastatic prostate cancer cell lines. 2010, 26 (6):553-67 Cell Biol. Toxicol.
1573-6822
20397042
10.1007/s10565-010-9163-5
http://hdl.handle.net/10033/134762
Cell biology and toxicology
The epithelial-mesenchymal transition (EMT) is regarded as an important step in cancer metastasis. Snail, a master regulator of EMT, has been recently proposed to act additionally as a cell survival factor and inducer of motility. We have investigated the function of Snail (SNAI1) in prostate cancer cells by downregulating its expression via short (21-mer) interfering RNA (siRNA) and measuring the consequences on EMT markers, cell viability, death, cell cycle, senescence, attachment, and invasivity. Of eight carcinoma cell lines, the prostate carcinoma cell lines LNCaP and PC-3 showed the highest and moderate expression of SNAI1 mRNA, respectively, as measured by quantitative RT-PCR. Long-term knockdown of Snail induced a severe decline in cell numbers in LNCaP and PC-3 and caspase activity was accordingly enhanced in both cell lines. In addition, suppression of Snail expression induced senescence in LNCaP cells. SNAI1-siRNA-treated cells did not tolerate detachment from the extracellular matrix, probably due to downregulation of integrin α6. Expression of E-cadherin, vimentin, and fibronectin was also affected. Invasiveness of PC-3 cells was not significantly diminished by Snail knockdown. Our data suggest that Snail acts primarily as a survival factor and inhibitor of cellular senescence in prostate cancer cell lines. We therefore propose that Snail can act as early driver of prostate cancer progression.
en
Cell Aging
Cell Line, Tumor
Cell Survival
Down-Regulation
Epithelial-Mesenchymal Transition
Gene Expression Regulation, Neoplastic
Humans
Integrin alpha6
Male
Neoplasm Metastasis
Prostatic Neoplasms
RNA, Messenger
Transcription Factors
Snail regulates cell survival and inhibits cellular senescence in human metastatic prostate cancer cell lines.
Article
2018-06-13T21:41:35Z
The epithelial-mesenchymal transition (EMT) is regarded as an important step in cancer metastasis. Snail, a master regulator of EMT, has been recently proposed to act additionally as a cell survival factor and inducer of motility. We have investigated the function of Snail (SNAI1) in prostate cancer cells by downregulating its expression via short (21-mer) interfering RNA (siRNA) and measuring the consequences on EMT markers, cell viability, death, cell cycle, senescence, attachment, and invasivity. Of eight carcinoma cell lines, the prostate carcinoma cell lines LNCaP and PC-3 showed the highest and moderate expression of SNAI1 mRNA, respectively, as measured by quantitative RT-PCR. Long-term knockdown of Snail induced a severe decline in cell numbers in LNCaP and PC-3 and caspase activity was accordingly enhanced in both cell lines. In addition, suppression of Snail expression induced senescence in LNCaP cells. SNAI1-siRNA-treated cells did not tolerate detachment from the extracellular matrix, probably due to downregulation of integrin α6. Expression of E-cadherin, vimentin, and fibronectin was also affected. Invasiveness of PC-3 cells was not significantly diminished by Snail knockdown. Our data suggest that Snail acts primarily as a survival factor and inhibitor of cellular senescence in prostate cancer cell lines. We therefore propose that Snail can act as early driver of prostate cancer progression.
ORIGINAL
Emadi et al_final.pdf
Emadi et al_final.pdf
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2019-08-30 11:37:24.056
Helmholtz Zentrum für Infektionsforschung Repository
hzi@openrepository.com
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oai:repository.helmholtz-hzi.de:10033/1460102019-08-30T11:25:11Zcom_10033_6807com_10033_6799col_10033_6884
Oelkers, J Margit
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Vinzenz, Marlene
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Nemethova, Maria
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Jacob, Sonja
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Lai, Frank P L
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Block, Jennifer
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Szczodrak, Malgorzata
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Kerkhoff, Eugen
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Backert, Steffen
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Schlüter, Kai
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Stradal, Theresia E B
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Small, J Victor
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Koestler, Stefan A
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Rottner, Klemens
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Helmholtz Centre for Infection Research, Braunschweig, Germany.
2011-10-19T13:44:11Z
2011-10-19T13:44:11Z
2011
Microtubules as platforms for assaying actin polymerization in vivo. 2011, 6 (5):e19931 PLoS ONE
1932-6203
21603613
10.1371/journal.pone.0019931
http://hdl.handle.net/10033/146010
PloS one
The actin cytoskeleton is continuously remodeled through cycles of actin filament assembly and disassembly. Filaments are born through nucleation and shaped into supramolecular structures with various essential functions. These range from contractile and protrusive assemblies in muscle and non-muscle cells to actin filament comets propelling vesicles or pathogens through the cytosol. Although nucleation has been extensively studied using purified proteins in vitro, dissection of the process in cells is complicated by the abundance and molecular complexity of actin filament arrays. We here describe the ectopic nucleation of actin filaments on the surface of microtubules, free of endogenous actin and interfering membrane or lipid. All major mechanisms of actin filament nucleation were recapitulated, including filament assembly induced by Arp2/3 complex, formin and Spir. This novel approach allows systematic dissection of actin nucleation in the cytosol of live cells, its genetic re-engineering as well as screening for new modifiers of the process.
en
Microtubules as platforms for assaying actin polymerization in vivo.
Article
2018-06-12T23:42:40Z
The actin cytoskeleton is continuously remodeled through cycles of actin filament assembly and disassembly. Filaments are born through nucleation and shaped into supramolecular structures with various essential functions. These range from contractile and protrusive assemblies in muscle and non-muscle cells to actin filament comets propelling vesicles or pathogens through the cytosol. Although nucleation has been extensively studied using purified proteins in vitro, dissection of the process in cells is complicated by the abundance and molecular complexity of actin filament arrays. We here describe the ectopic nucleation of actin filaments on the surface of microtubules, free of endogenous actin and interfering membrane or lipid. All major mechanisms of actin filament nucleation were recapitulated, including filament assembly induced by Arp2/3 complex, formin and Spir. This novel approach allows systematic dissection of actin nucleation in the cytosol of live cells, its genetic re-engineering as well as screening for new modifiers of the process.
ORIGINAL
Oelkers et al_final.pdf
Oelkers et al_final.pdf
Open Access publication
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TEXT
Oelkers et al_final.pdf.txt
Oelkers et al_final.pdf.txt
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10033/146010
oai:hzi.openrepository.com:10033/146010
2019-08-30 11:25:11.21
Helmholtz Zentrum für Infektionsforschung Repository
hzi@openrepository.com
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oai:repository.helmholtz-hzi.de:10033/1963292019-08-30T11:25:43Zcom_10033_6807com_10033_6799col_10033_6884
Linkner, Joern
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Witte, Gregor
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Stradal, Theresia
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Curth, Ute
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Faix, Jan
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Institute for Biophysical Chemistry, Hannover Medical School, Hannover, Germany.
2011-12-07T15:36:57Z
2011-12-07T15:36:57Z
2011
High-resolution X-ray structure of the trimeric Scar/WAVE-complex precursor Brk1. 2011, 6 (6):e21327 PLoS ONE
1932-6203
21701600
10.1371/journal.pone.0021327
http://hdl.handle.net/10033/196329
PloS one
The Scar/WAVE-complex links upstream Rho-GTPase signaling to the activation of the conserved Arp2/3-complex. Scar/WAVE-induced and Arp2/3-complex-mediated actin nucleation is crucial for actin assembly in protruding lamellipodia to drive cell migration. The heteropentameric Scar/WAVE-complex is composed of Scar/WAVE, Abi, Nap, Pir and a small polypeptide Brk1/HSPC300, and recent work suggested that free Brk1 serves as a homooligomeric precursor in the assembly of this complex. Here we characterized the Brk1 trimer from Dictyostelium by analytical ultracentrifugation and gelfiltration. We show for the first time its dissociation at concentrations in the nanomolar range as well as an exchange of subunits within different DdBrk1 containing complexes. Moreover, we determined the three-dimensional structure of DdBrk1 at 1.5 Å resolution by X-ray crystallography. Three chains of DdBrk1 are associated with each other forming a parallel triple coiled-coil bundle. Notably, this structure is highly similar to the heterotrimeric α-helical bundle of HSPC300/WAVE1/Abi2 within the human Scar/WAVE-complex. This finding, together with the fact that Brk1 is collectively sandwiched by the remaining subunits and also constitutes the main subunit connecting the triple-coil domain of the HSPC300/WAVE1/Abi2/ heterotrimer to Sra1(Pir1), implies a critical function of this subunit in the assembly process of the entire Scar/WAVE-complex.
en
Blotting, Western
Chromatography, Gel
Crystallography, X-Ray
Dictyostelium
Protozoan Proteins
Ultracentrifugation
Wiskott-Aldrich Syndrome Protein Family
High-resolution X-ray structure of the trimeric Scar/WAVE-complex precursor Brk1.
Article
2018-06-12T22:09:08Z
The Scar/WAVE-complex links upstream Rho-GTPase signaling to the activation of the conserved Arp2/3-complex. Scar/WAVE-induced and Arp2/3-complex-mediated actin nucleation is crucial for actin assembly in protruding lamellipodia to drive cell migration. The heteropentameric Scar/WAVE-complex is composed of Scar/WAVE, Abi, Nap, Pir and a small polypeptide Brk1/HSPC300, and recent work suggested that free Brk1 serves as a homooligomeric precursor in the assembly of this complex. Here we characterized the Brk1 trimer from Dictyostelium by analytical ultracentrifugation and gelfiltration. We show for the first time its dissociation at concentrations in the nanomolar range as well as an exchange of subunits within different DdBrk1 containing complexes. Moreover, we determined the three-dimensional structure of DdBrk1 at 1.5 Å resolution by X-ray crystallography. Three chains of DdBrk1 are associated with each other forming a parallel triple coiled-coil bundle. Notably, this structure is highly similar to the heterotrimeric α-helical bundle of HSPC300/WAVE1/Abi2 within the human Scar/WAVE-complex. This finding, together with the fact that Brk1 is collectively sandwiched by the remaining subunits and also constitutes the main subunit connecting the triple-coil domain of the HSPC300/WAVE1/Abi2/ heterotrimer to Sra1(Pir1), implies a critical function of this subunit in the assembly process of the entire Scar/WAVE-complex.
ORIGINAL
Linkner et al_final.pdf
Linkner et al_final.pdf
Open Access publication
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THUMBNAIL
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TEXT
Linkner et al_final.pdf.txt
Linkner et al_final.pdf.txt
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10033/196329
oai:hzi.openrepository.com:10033/196329
2019-08-30 11:25:43.566
Helmholtz Zentrum für Infektionsforschung Repository
hzi@openrepository.com
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oai:repository.helmholtz-hzi.de:10033/2008892019-08-30T11:31:49Zcom_10033_6807com_10033_6799col_10033_6884
Kabaso, Doron
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Shlomovitz, Roie
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Schloen, Kathrin
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Stradal, Theresia
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Gov, Nir S
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Department of Chemical Physics, The Weizmann Institute of Science, Rehovot, Israel.
2012-01-09T10:43:20Z
2012-01-09T10:43:20Z
2011-05
Theoretical model for cellular shapes driven by protrusive and adhesive forces. 2011, 7 (5):e1001127 PLoS Comput. Biol.
1553-7358
21573201
10.1371/journal.pcbi.1001127
http://hdl.handle.net/10033/200889
PLoS computational biology
The forces that arise from the actin cytoskeleton play a crucial role in determining the cell shape. These include protrusive forces due to actin polymerization and adhesion to the external matrix. We present here a theoretical model for the cellular shapes resulting from the feedback between the membrane shape and the forces acting on the membrane, mediated by curvature-sensitive membrane complexes of a convex shape. In previous theoretical studies we have investigated the regimes of linear instability where spontaneous formation of cellular protrusions is initiated. Here we calculate the evolution of a two dimensional cell contour beyond the linear regime and determine the final steady-state shapes arising within the model. We find that shapes driven by adhesion or by actin polymerization (lamellipodia) have very different morphologies, as observed in cells. Furthermore, we find that as the strength of the protrusive forces diminish, the system approaches a stabilization of a periodic pattern of protrusions. This result can provide an explanation for a number of puzzling experimental observations regarding cellular shape dependence on the properties of the extra-cellular matrix.
en
Actins
Animals
Biomechanics
Cell Adhesion
Cell Shape
Cells, Cultured
Cytoskeleton
Extracellular Matrix
Fibroblasts
Mice
Models, Biological
Pseudopodia
Theoretical model for cellular shapes driven by protrusive and adhesive forces.
Article
2018-06-12T22:44:32Z
The forces that arise from the actin cytoskeleton play a crucial role in determining the cell shape. These include protrusive forces due to actin polymerization and adhesion to the external matrix. We present here a theoretical model for the cellular shapes resulting from the feedback between the membrane shape and the forces acting on the membrane, mediated by curvature-sensitive membrane complexes of a convex shape. In previous theoretical studies we have investigated the regimes of linear instability where spontaneous formation of cellular protrusions is initiated. Here we calculate the evolution of a two dimensional cell contour beyond the linear regime and determine the final steady-state shapes arising within the model. We find that shapes driven by adhesion or by actin polymerization (lamellipodia) have very different morphologies, as observed in cells. Furthermore, we find that as the strength of the protrusive forces diminish, the system approaches a stabilization of a periodic pattern of protrusions. This result can provide an explanation for a number of puzzling experimental observations regarding cellular shape dependence on the properties of the extra-cellular matrix.
ORIGINAL
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2019-08-30 11:31:49.281
Helmholtz Zentrum für Infektionsforschung Repository
hzi@openrepository.com
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oai:repository.helmholtz-hzi.de:10033/2183772019-08-30T11:34:48Zcom_10033_6807com_10033_6799col_10033_6884
Dubielecka, Patrycja M
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500
Ladwein, Kathrin I
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500
Xiong, Xiaoling
fe06b84da28bdfb8ffce5855308a639c
500
Migeotte, Isabelle
143e5696de313171593b199f5ca8c0ca
500
Chorzalska, Anna
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500
Anderson, Kathryn V
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500
Sawicki, Janet A
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500
Rottner, Klemens
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Stradal, Theresia E
58958e12d8699368d0442aad9238fb13
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Kotula, Leszek
327d1f03367d4ad338b63f9add3050fa
500
Laboratory of Cell Signaling, New York Blood Center, New York, NY 10065, USA.
2012-04-13T13:42:12Z
2012-04-13T13:42:12Z
2011-04-26
Essential role for Abi1 in embryonic survival and WAVE2 complex integrity. 2011, 108 (17):7022-7 Proc. Natl. Acad. Sci. U.S.A.
1091-6490
21482783
10.1073/pnas.1016811108
http://hdl.handle.net/10033/218377
Proceedings of the National Academy of Sciences of the United States of America
Abl interactor 1 (Abi1) plays a critical function in actin cytoskeleton dynamics through participation in the WAVE2 complex. To gain a better understanding of the specific role of Abi1, we generated a conditional Abi1-KO mouse model and MEFs lacking Abi1 expression. Abi1-KO cells displayed defective regulation of the actin cytoskeleton, and this dysregulation was ascribed to altered activity of the WAVE2 complex. Changes in motility of Abi1-KO cells were manifested by a decreased migration rate and distance but increased directional persistence. Although these phenotypes did not correlate with peripheral ruffling, which was unaffected, Abi1-KO cells exhibited decreased dorsal ruffling. Western blotting analysis of Abi1-KO cell lysates indicated reduced levels of the WAVE complex components WAVE1 and WAVE2, Nap1, and Sra-1/PIR121. Although relative Abi2 levels were more than doubled in Abi1-KO cells, the absolute Abi2 expression in these cells amounted only to a fifth of Abi1 levels in the control cell line. This finding suggests that the presence of Abi1 is critical for the integrity and stability of WAVE complex and that Abi2 levels are not sufficiently increased to compensate fully for the loss of Abi1 in KO cells and to restore the integrity and function of the WAVE complex. The essential function of Abi1 in WAVE complexes and their regulation might explain the observed embryonic lethality of Abi1-deficient embryos, which survived until approximately embryonic day 11.5 and displayed malformations in the developing heart and brain. Cells lacking Abi1 and the conditional Abi1-KO mouse will serve as critical models for defining Abi1 function.
en
Archived with thanks to Proceedings of the National Academy of Sciences of the United States of America
Adaptor Proteins, Signal Transducing
Animals
Brain
Cell Line
Cell Movement
Cytoskeletal Proteins
Embryo, Mammalian
Heart
Homeodomain Proteins
Membrane Proteins
Mice
Mice, Knockout
Multiprotein Complexes
Wiskott-Aldrich Syndrome Protein Family
Essential role for Abi1 in embryonic survival and WAVE2 complex integrity.
Article
2018-06-12T21:30:30Z
Abl interactor 1 (Abi1) plays a critical function in actin cytoskeleton dynamics through participation in the WAVE2 complex. To gain a better understanding of the specific role of Abi1, we generated a conditional Abi1-KO mouse model and MEFs lacking Abi1 expression. Abi1-KO cells displayed defective regulation of the actin cytoskeleton, and this dysregulation was ascribed to altered activity of the WAVE2 complex. Changes in motility of Abi1-KO cells were manifested by a decreased migration rate and distance but increased directional persistence. Although these phenotypes did not correlate with peripheral ruffling, which was unaffected, Abi1-KO cells exhibited decreased dorsal ruffling. Western blotting analysis of Abi1-KO cell lysates indicated reduced levels of the WAVE complex components WAVE1 and WAVE2, Nap1, and Sra-1/PIR121. Although relative Abi2 levels were more than doubled in Abi1-KO cells, the absolute Abi2 expression in these cells amounted only to a fifth of Abi1 levels in the control cell line. This finding suggests that the presence of Abi1 is critical for the integrity and stability of WAVE complex and that Abi2 levels are not sufficiently increased to compensate fully for the loss of Abi1 in KO cells and to restore the integrity and function of the WAVE complex. The essential function of Abi1 in WAVE complexes and their regulation might explain the observed embryonic lethality of Abi1-deficient embryos, which survived until approximately embryonic day 11.5 and displayed malformations in the developing heart and brain. Cells lacking Abi1 and the conditional Abi1-KO mouse will serve as critical models for defining Abi1 function.
ORIGINAL
Dubielecka etal_final.pdf
Dubielecka etal_final.pdf
original manuscript
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oai:hzi.openrepository.com:10033/218377
2019-08-30 11:34:48.174
Helmholtz Zentrum für Infektionsforschung Repository
hzi@openrepository.com
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