Chemical manipulations to facilitate membrane blebbing and vesicle shedding on the cellular cortex

2020 ◽  
Vol 42 (7) ◽  
pp. 1137-1145
Author(s):  
Saya Okada ◽  
Yuta Fukai ◽  
Fumiya Yoshimoto ◽  
Hisato Saitoh
Keyword(s):  
Membrane Blebbing ◽  
Vesicle Shedding

Absence of temporal ordering of apoptotic features in heat-shock treated leukemia and lymphoma cell lines

1996 ◽  
Vol 54 ◽  
pp. 758-759
Author(s):  
D. W. Fairbain ◽  
M.D. Standing ◽  
K.L. O'Neill
Keyword(s):  
Heat Shock ◽  
Cell Lines ◽  
Chromatin Condensation ◽  
Membrane Integrity ◽  
Resistant Cell ◽  
Membrane Blebbing ◽  
Late Loss ◽  
Induced Apoptosis ◽  
Dying Cells ◽  
In Cells

Apoptosis is a genetically defined response to physiological stimuli that results in cellular suicide. Features common to apoptotic cells include chromatin condensation, oligonucleosomal DNA fragmentation, membrane blebbing, nuclear destruction, and late loss of ability to exclude vital dyes. These characteristics contrast markedly from pathological necrosis, in which membrane integrity loss is demonstrated early, and other features of apoptosis, which allow a non-inflammatory removal of dead and dying cells, are absent. Using heat shock-induced apoptosis as a model for examining stress response in cells, we undertook to categorize a variety of human leukemias and lymphomas with regard to their response to heat shock. We were also interested in determining whether a common temporal order was followed in cells dying by apoptosis. In addition, based on our previous results, we investigated whether increasing heat load resulted in increased apoptosis, with particular interest in relatively resistant cell lines, or whether the mode of death changed from apoptosis to necrosis.

scholarly journals Caspase-mediated Cleavage of p130cas in Etoposide-induced Apoptotic Rat-1 Cells

2000 ◽  
Vol 11 (3) ◽  
pp. 929-939 ◽  
Author(s):  
Seunghyi Kook ◽  
Sang Ryeol Shim ◽  
Soo Jeon Choi ◽  
Joohong Ahnn ◽  
Jae Il Kim ◽  
...  
Keyword(s):  
Extracellular Matrix ◽  
Focal Adhesion ◽  
Caspase 3 ◽  
Morphological Changes ◽  
Point Mutations ◽  
Membrane Blebbing ◽  
Adhesion Sites ◽  
Focal Adhesion Proteins ◽  
Induced Apoptosis

Apoptosis causes characteristic morphological changes in cells, including membrane blebbing, cell detachment from the extracellular matrix, and loss of cell–cell contacts. We investigated the changes in focal adhesion proteins during etoposide-induced apoptosis in Rat-1 cells and found that during apoptosis, p130cas (Crk-associated substrate [Cas]) is cleaved by caspase-3. Sequence analysis showed that Cas contains 10 DXXD consensus sites preferred by caspase-3. We identified two of these sites (DVPD416G and DSPD748G) in vitro, and point mutations substituting the Asp of DVPD416G and DSPD748G with Glu blocked caspase-3-mediated cleavage. Cleavage at DVPD416G generated a 74-kDa fragment, which was in turn cleaved at DSPD748G, yielding 47- and 31-kDa fragments. Immunofluorescence microscopy revealed well-developed focal adhesion sites in control cells that dramatically declined in number in etoposide-treated cells. Cas cleavage correlated temporally with the onset of apoptosis and coincided with the loss of p125FAK (focal adhesion kinase [FAK]) from focal adhesion sites and the attenuation of Cas–paxillin interactions. Considering that Cas associates with FAK, paxillin, and other molecules involved in the integrin signaling pathway, these results suggest that caspase-mediated cleavage of Cas contributes to the disassembly of focal adhesion complexes and interrupts survival signals from the extracellular matrix.

scholarly journals Genetic disruption of calpain correlates with loss of membrane blebbing and differential expression of RhoGDI-1, cofilin and tropomyosin

2008 ◽  
Vol 411 (3) ◽  
pp. 657-666 ◽  
Author(s):  
Anna K. Larsen ◽  
René Lametsch ◽  
John S. Elce ◽  
Jørgen K. Larsen ◽  
Bo Thomsen ◽  
...  
Keyword(s):  
Membrane Surface ◽  
Proteome Analysis ◽  
Time Lapse ◽  
Wild Type ◽  
Genetic Rescue ◽  
Dynamic Regulation ◽  
Membrane Blebbing ◽  
Signalling Molecules ◽  
Membrane Blebs

Dynamic regulation of the actin cytoskeleton is important for cell motility, spreading and the formation of membrane surface extensions such as lamellipodia, ruffles and blebs. The ubiquitous calpains contribute to integrin-mediated cytoskeletal remodelling during cell migration and spreading, by cleavage of focal adhesion components and signalling molecules. In the present study, the live-cell morphology of calpain-knockout and wild-type cells was examined by time-lapse fluorescence microscopy, and a role of calpain in mediating the formation of sporadic membrane blebs was established. Membrane blebbing was significantly reduced in calpain-knockout cells, and genetic rescue fully restored the wild-type phenotype in knockout cells. Proteomic comparison of wild-type and knockout cells identified decreased levels of RhoGDI-1 (Rho GDP-dissociation inhibitor) and cofilin 1, and increased levels of tropomyosin in calpain-knockout cells, suggesting a role of calpain in regulating membrane extensions involving these proteins. RhoGDI, cofilin and tropomyosin are known regulators of actin filament dynamics and membrane extensions. The reduced levels of RhoGDI-1 in calpain-knockout cells observed by proteome analysis were confirmed by immunoblotting. Genetic rescue of the calpain-knockout cells enhanced RhoGDI-1-expression 2-fold above that normally present in wild-type cells. These results suggest a regulatory connection between calpain and RhoGDI-1 in promoting formation of membrane blebs.

scholarly journals P2X7 Receptor-mediated Membrane Blebbing in Salivary Epithelial Cells

2009 ◽  
Vol 13 (3) ◽  
pp. 175 ◽  
Author(s):  
Sung-Min Hwang ◽  
Na-Youn Koo ◽  
Se-Young Choi ◽  
Gae-Sig Chun ◽  
Joong-Soo Kim ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
P2x7 Receptor ◽  
Membrane Blebbing ◽  
Salivary Epithelial Cells

scholarly journals Membrane blebbing as an assessment of functional rescue of dysferlin-deficient human myotubes via nonsense suppression

2010 ◽  
Vol 109 (3) ◽  
pp. 901-905 ◽  
Author(s):  
Bingjing Wang ◽  
Zhaohui Yang ◽  
Becky K. Brisson ◽  
Huisheng Feng ◽  
Zhiqian Zhang ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Stop Codon ◽  
Premature Stop Codon ◽  
Nonsense Suppression ◽  
Muscle Membrane ◽  
Membrane Repair ◽  
Vitro Assay ◽  
Membrane Blebbing ◽  
Human Myotubes

Mutations that result in the loss of the protein dysferlin result in defective muscle membrane repair and cause either a form of limb girdle muscular dystrophy (type 2B) or Miyoshi myopathy. Most patients are compound heterozygotes, often carrying one allele with a nonsense mutation. Using dysferlin-deficient mouse and human myocytes, we demonstrated that membrane blebbing in skeletal muscle myotubes in response to hypotonic shock requires dysferlin. Based on this, we developed an in vitro assay to assess rescue of dysferlin function in skeletal muscle myotubes. This blebbing assay may be useful for drug discovery/validation for dysferlin deficiency. With this assay, we demonstrate that the nonsense suppression drug, ataluren (PTC124), is able to induce read-through of the premature stop codon in a patient with a R1905X mutation in dysferlin and produce sufficient functional dysferlin (∼15% of normal levels) to rescue myotube membrane blebbing. Thus ataluren is a potential therapeutic for dysferlin-deficient patients harboring nonsense mutations.

scholarly journals Osteoblast-conditioned media stimulate membrane vesicle shedding in prostate cancer cells

2006 ◽  
Author(s):  
Danilo Millimaggi ◽  
Claudio Festuccia ◽  
Adriano Angelucci ◽  
Sandra D'Ascenzo ◽  
Nadia Rucci ◽  
...  
Keyword(s):  
Prostate Cancer ◽  
Cancer Cells ◽  
Membrane Vesicle ◽  
Conditioned Media ◽  
Prostate Cancer Cells ◽  
Vesicle Shedding

scholarly journals General principles for the formation and proliferation of a wall-free (L-form) state in bacteria

eLife ◽  
2014 ◽  
Vol 3 ◽  
Author(s):  
Romain Mercier ◽  
Yoshikazu Kawai ◽  
Jeff Errington
Keyword(s):  
Cell Wall ◽  
Molecular Biology ◽  
Gram Negative Bacteria ◽  
Systematic Analysis ◽  
Gram Negative ◽  
Membrane Blebbing ◽  
Precursor Synthesis ◽  
Peptidoglycan Cell Wall ◽  
Synthetic Cells ◽  
Opening Up

The peptidoglycan cell wall is a defining structural feature of the bacterial kingdom. Curiously, some bacteria have the ability to switch to a wall-free or ‘L-form’ state. Although known for decades, the general properties of L-forms are poorly understood, largely due to the lack of systematic analysis of L-forms in the molecular biology era. Here we show that inhibition of peptidoglycan precursor synthesis promotes the generation of L-forms from both Gram-positive and Gram-negative bacteria. We show that the L-forms generated have in common a mechanism of proliferation involving membrane blebbing and tubulation, which is dependent on an altered rate of membrane synthesis. Crucially, this mode of proliferation is independent of the essential FtsZ based division machinery. Our results suggest that the L-form mode of proliferation is conserved across the bacterial kingdom, reinforcing the idea that it could have been used in primitive cells, and opening up its use in the generation of synthetic cells.

scholarly journals MARCKS domain phosphorylation regulates the differential interaction of Diacylglycerol Kinase ζ with Rac1, RhoA and Syntrophin

2020 ◽  
Author(s):  
Ryan Ard ◽  
Jean-Christian Maillet ◽  
Elias Daher ◽  
Michael Phan ◽  
Radoslav Zinoviev ◽  
...  
Keyword(s):  
Catalytic Activity ◽  
Molecular Mechanisms ◽  
Rho Gtpase ◽  
Pdz Domain ◽  
Diacylglycerol Kinase ◽  
Binding Motif ◽  
Membrane Blebbing ◽  
Rhoa Activity ◽  
C Terminus ◽  
Mechanistic Basis

AbstractCells can switch between Rac1, lamellipodia-based and RhoA, blebbing-based migration modes but the molecular mechanisms regulating this choice are not fully understood. Diacylglycerol kinase ζ (DGKζ), which phosphorylates diacylglycerol to yield phosphatidic acid, forms independent complexes with Rac1 and RhoA, selectively dissociating each from RhoGDI. DGKζ catalytic activity is required for Rac1 dissociation but is dispensable for RhoA dissociation. Instead, DGKζ functions as a scaffold that stimulates RhoA release by enhancing RhoGDI phosphorylation by protein kinase Cα (PKCα). Here, PKCα-mediated phosphorylation of the DGKζ MARCKS domain increased DGKζ association with RhoA and decreased its interaction with Rac1. The same modification increased binding of the DGKζ C-terminus to the α1-syntrophin PDZ domain. Expression of a phosphomimetic DGKζ mutant stimulated membrane blebbing in mouse embryonic fibroblasts and C2C12 myoblasts, which was augmented by inhibition of endogenous Rac1. DGKζ expression in differentiated C2 myotubes, which have low endogenous Rac1 levels, also induced substantial membrane blebbing via the Rho-ROCK pathway. These events were independent of DGKζ catalytic activity, but dependent upon a functional C-terminal PDZ-binding motif. Rescue of RhoA activity in DGKζ-null cells required the PDZ-binding motif, suggesting syntrophin interaction is necessary for optimal RhoA activation. Collectively, our results define a switch-like mechanism involving DGKζ phosphorylation by PKCα that favours RhoA-driven blebbing over Rac1-driven lamellipodia formation and macropinocytosis. These findings provide a mechanistic basis for the effect of PKCα signaling on Rho GTPase activity and suggest PKCα activity plays a role in the interconversion between Rac1 and RhoA signaling that underlies different migration modes.

scholarly journals Mechanisms of programmed cell death

2019 ◽  
Vol 6 (4) ◽  
pp. 156-158
Author(s):  
Abdu-Alhameed A Ali Azzwali ◽  
 Azab Elsayed Azab
Keyword(s):  
Dna Damage ◽  
Cell Death ◽  
Programmed Cell Death ◽  
Nuclear Dna ◽  
Chromatin Condensation ◽  
Cell Shrinkage ◽  
Membrane Blebbing ◽  
Development And Aging ◽  
Nuclear Dna Fragmentation ◽  
Dependent Pathway

The present review aims to spotlight on the mechanisms and stages of programmed cell death. Apoptosis, known as programmed cell death, is a homeostatic mechanism that generally occurs during development and aging in order to keep cells in tissue. It can also act as a protective mechanism, for example, in immune response or if cells are damaged by toxin agents or diseases. In cancer treatment, drugs and irradiation used in chemotherapy leads to DNA damage, which results in triggering apoptosis through the p53 dependent pathway in cancer treatment, drugs and irradiation used in chemotherapy leads to DNA damage, which results in triggering apoptosis through the p53 dependent pathway. Corticosteroids can cause apoptotic death in a number of cells. A number of changes in cell morphology are related to the different stages of apoptosis, which includes nuclear DNA fragmentation, cell shrinkage, chromatin condensation, membrane blebbing, and the formation of apoptotic bodies. There are three pathways for apoptosis, the intrinsic (mitochondrial) and extrinsic (death receptor) are the two major paths that are interlinked and that can effect one another. Conclusion: It can be concluded that apoptosis is a homeostatic mechanism that generally occurs during development and aging in order to keep cells in tissue. Drugs and irradiation used in chemotherapy leads to DNA damage, which results in triggering apoptosis through the p53 dependent pathway. The apoptosis, stages are includes nuclear DNA fragmentation, cell shrinkage, chromatin condensation, membrane blebbing, and the formation of apoptotic bodies. There are three pathways for apoptosis.

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