scholarly journals Regulated changes in material properties underlie centrosome disassembly during mitotic exit

2020 ◽  
Vol 219 (4) ◽  
Author(s):  
Matthäus Mittasch ◽  
Vanna M. Tran ◽  
Manolo U. Rios ◽  
Anatol W. Fritsch ◽  
Stephen J. Enos ◽  
...  
Keyword(s):  
Material Properties ◽  
Mitotic Exit ◽  
C Elegans ◽  
Anaphase Onset ◽  
Pulling Forces ◽  
Mechanical Transition ◽  
Induced Flow ◽  
Brittle State ◽  
Strength And Ductility

Centrosomes must resist microtubule-mediated forces for mitotic chromosome segregation. During mitotic exit, however, centrosomes are deformed and fractured by those same forces, which is a key step in centrosome disassembly. How the functional material properties of centrosomes change throughout the cell cycle, and how they are molecularly tuned, remain unknown. Here, we used optically induced flow perturbations to determine the molecular basis of centrosome strength and ductility in C. elegans embryos. We found that both properties declined sharply at anaphase onset, long before natural disassembly. This mechanical transition required PP2A phosphatase and correlated with inactivation of PLK-1 (Polo kinase) and SPD-2 (Cep192). In vitro, PLK-1 and SPD-2 directly protected centrosome scaffolds from force-induced disassembly. Our results suggest that, before anaphase, PLK-1 and SPD-2 respectively confer strength and ductility to the centrosome scaffold so that it can resist microtubule-pulling forces. In anaphase, centrosomes lose PLK-1 and SPD-2 and transition to a weak, brittle state that enables force-mediated centrosome disassembly.

scholarly journals Material aging causes centrosome weakening and disassembly during mitotic exit

2019 ◽  
Author(s):  
Matthäus Mittasch ◽  
Vanna M. Tran ◽  
Manolo U. Rios ◽  
Anatol W. Fritsch ◽  
Stephen J. Enos ◽  
...  
Keyword(s):  
Mitotic Chromosome ◽  
Mitotic Exit ◽  
C Elegans ◽  
Anaphase Onset ◽  
Pulling Forces ◽  
Mechanical Transition ◽  
Induced Flow ◽  
Brittle State ◽  
Strength And Ductility

ABSTRACTCentrosomes must resist microtubule-mediated forces for mitotic chromosome segregation. During mitotic exit, however, centrosomes are deformed and fractured by those same forces, which is a key step in centrosome disassembly. How the functional material properties of centrosomes change throughout the cell cycle, and how they are molecularly tuned remain unknown. Here, we used optically-induced flow perturbations to determine the molecular basis of centrosome strength and ductility in C. elegans embryos. We found that both properties declined sharply at anaphase onset, long before natural disassembly. This mechanical transition required PP2A phosphatase and correlated with inactivation of PLK-1 (Polo Kinase) and SPD-2 (Cep192). In vitro, PLK-1 and SPD-2 directly protected centrosome scaffolds from force-induced disassembly. Our results suggest that, prior to anaphase, PLK-1 and SPD-2 confer strength and ductility to the centrosome scaffold so that it can resist microtubule-pulling forces. In anaphase, centrosomes lose PLK-1 and SPD-2 and transition to a weak, brittle state that enables force-mediated centrosome disassembly.

scholarly journals Protein Phosphatase 1 inactivates Mps1 to ensure efficient Spindle Assembly Checkpoint silencing

eLife ◽  
2017 ◽  
Vol 6 ◽  
Author(s):  
Margarida Moura ◽  
Mariana Osswald ◽  
Nelson Leça ◽  
João Barbosa ◽  
António J Pereira ◽  
...  
Keyword(s):  
Protein Phosphatase ◽  
Spindle Assembly Checkpoint ◽  
Spindle Assembly ◽  
Mitotic Exit ◽  
Protein Phosphatase 1 ◽  
Anaphase Onset ◽  
Phosphorylation Cascade ◽  
Spindle Microtubules ◽  
Early Mitosis

Faithfull genome partitioning during cell division relies on the Spindle Assembly Checkpoint (SAC), a conserved signaling pathway that delays anaphase onset until all chromosomes are attached to spindle microtubules. Mps1 kinase is an upstream SAC regulator that promotes the assembly of an anaphase inhibitor through a sequential multi-target phosphorylation cascade. Thus, the SAC is highly responsive to Mps1, whose activity peaks in early mitosis as a result of its T-loop autophosphorylation. However, the mechanism controlling Mps1 inactivation once kinetochores attach to microtubules and the SAC is satisfied remains unknown. Here we show in vitro and in Drosophila that Protein Phosphatase 1 (PP1) inactivates Mps1 by dephosphorylating its T-loop. PP1-mediated dephosphorylation of Mps1 occurs at kinetochores and in the cytosol, and inactivation of both pools of Mps1 during metaphase is essential to ensure prompt and efficient SAC silencing. Overall, our findings uncover a mechanism of SAC inactivation required for timely mitotic exit.

scholarly journals Phosphatase PP2A and microtubule pulling forces disassemble centrosomes during mitotic exit

2017 ◽  
Author(s):  
Stephen J. Enos ◽  
Martin Dressler ◽  
Beatriz Ferreira Gomes ◽  
Anthony A. Hyman ◽  
Jeffrey B. Woodruff
Keyword(s):  
Cell Cycle ◽  
Chromosome Segregation ◽  
Mitotic Exit ◽  
Scaffold Proteins ◽  
Regulatory Subunit ◽  
Pulling Forces ◽  
Dividing Cells ◽  
Pericentriolar Material ◽  
Embryonic Death

ABSTRACTCentrosomes are major microtubule-nucleating organelles that facilitate chromosome segregation and cell division in metazoans. Centrosomes comprise centrioles that organize a micron-scale mass of protein called pericentriolar material (PCM) from which microtubules nucleate. During each cell cycle, PCM accumulates around centrioles through phosphorylation-mediated assembly of PCM scaffold proteins. During mitotic exit, PCM swiftly disassembles by an unknown mechanism. Here, we used Caenorhabditis elegans embryos to determine the mechanism and importance of PCM disassembly in dividing cells. We found that the phosphatase PP2A and its regulatory subunit SUR-6 (PP2ASUR-6), together with cortically directed microtubule pulling forces, actively disassemble PCM. In embryos depleted of these activities, ~25% of PCM persisted from one cell cycle into the next, resulting in cytokinetic furrow ingression errors, excessive centrosome accumulation, and embryonic death. Purified pp2ASUR-6 could dephosphorylate the major PCM scaffold protein SPD-5 in vitro. Our data suggest that PCM disassembly occurs through a combination of dephosphorylation of PCM components and catastrophic rupture of the PCM scaffold.

scholarly journals LET-99-dependent spatial restriction of active force generators makes spindle’s position robust

2017 ◽  
Author(s):  
H. Bouvrais ◽  
L. Chesneau ◽  
S. Pastezeur ◽  
M. Delattre ◽  
J. Pécréaux
Keyword(s):  
Biochemical Networks ◽  
Active Force ◽  
Mitotic Progression ◽  
Contact Density ◽  
Posterior Displacement ◽  
C Elegans ◽  
Anaphase Onset ◽  
Pulling Forces ◽  
Pulling Force ◽  
Force Generator

AbstractDuring the asymmetric division of the Caenorhabditis elegans nematode zygote, the polarity cues distribution and daughter cell fates depend on the correct positioning of the mitotic spindle, which results from both centering and cortical pulling forces. Revealed by anaphase spindle rocking, these pulling forces are regulated by the force generator dynamics, which are in turn consequent of mitotic progression. We found a novel, additional, regulation of these forces by the spindle position. It controls astral microtubule availability at the cortex, on which the active force generators can pull. Importantly, this positional control relies on the polarity dependent LET-99 cortical band, which restricts or concentrates generators to a posterior crescent. After delaying anaphase onset, we detected this positional pulling force regulation in C. elegans as a precocious spindle rocking with respect to anaphase onset. We ascribed this control to the microtubule dynamics at the cortex. Indeed, in mapping the cortical contacts, we found a correlation between the centrosome–cortex distance and the microtubule contact density. In turn, it modulates pulling force generator activity. We modelled this control, predicting and experimentally validating that the posterior crescent extent controlled where the anaphase oscillations started, in addition to mitotic progression. We found in particular that the oscillation onset position resists changes in cellular geometry and moderate variations of active force generator count. Finally, we propose that spatially restricting force generator to a posterior crescent sets the spindle’s final position, reflecting polarity through the LET-99 dependent restriction of force generators to a posterior crescent. This regulation superimposes that of force generator processivity. This novel control confers a low dependence on microtubule and active force generator exact numbers or dynamics, provided that they exceed the threshold needed for posterior displacement. Interestingly, this robustness originates in cell mechanics rather than biochemical networks.

scholarly journals Dynactin binding to tyrosinated microtubules promotes centrosome centration in C. elegans by enhancing dynein-mediated organelle transport

2017 ◽  
Author(s):  
Daniel José Barbosa ◽  
Joana Duro ◽  
Dhanya K. Cheerambathur ◽  
Bram Prevo ◽  
Ana Xavier Carvalho ◽  
...  
Keyword(s):  
Binding Activity ◽  
Organelle Transport ◽  
Cell Cortex ◽  
Cell Periphery ◽  
Animal Cells ◽  
Spindle Positioning ◽  
C Elegans ◽  
Pulling Forces

ABSTRACTThe microtubule-based motor dynein generates pulling forces for centrosome centration and mitotic spindle positioning in animal cells. How the essential dynein activator dynactin regulates these functions of the motor is incompletely understood. Here, we dissect the role of dynactin’s microtubule binding activity, located in p150’s CAP-Gly domain and an adjacent basic patch, in the C. elegans zygote. Using precise mutants engineered by genome editing, we show that microtubule tip tracking of dynein-dynactin is dispensable for targeting the motor to the cell cortex and for generating cortical pulling forces. Instead, p150 CAP-Gly mutants inhibit cytoplasmic pulling forces responsible for centration of centrosomes and attached pronuclei. The centration defects are mimicked by mutations of the C-terminal tyrosine of α-tubulin, and both p150 CAP-Gly and tubulin tyrosination mutants decrease the frequency of organelle transport from the cell periphery towards centrosomes during centration. In light of recent work on dynein-dynactin motility in vitro, our results suggest that p150 GAP-Gly domain binding to tyrosinated microtubules promotes initiation of dynein-mediated organelle transport in the dividing embryo, and that this function of dynactin is important for generating robust cytoplasmic pulling forces for centrosome centration.

Cytocompatibility and Material Properties of Poly-carbonate Urethane/Carbon Nanofiber Composites for Neural Applications

2003 ◽  
Vol 774 ◽  
Author(s):  
Janice L. McKenzie ◽  
Michael C. Waid ◽  
Riyi Shi ◽  
Thomas J. Webster
Keyword(s):  
Mechanical Properties ◽  
Carbon Nanofibers ◽  
Material Properties ◽  
Carbon Nanofiber ◽  
Scar Tissue ◽  
Tissue Response ◽  
Positive Interactions ◽  
Weight Percentage ◽  
Poly Carbonate

AbstractCarbon nanofibers possess excellent conductivity properties, which may be beneficial in the design of more effective neural prostheses, however, limited evidence on their cytocompatibility properties exists. The objective of the present in vitro study was to determine cytocompatibility and material properties of formulations containing carbon nanofibers to predict the gliotic scar tissue response. Poly-carbonate urethane was combined with carbon nanofibers in varying weight percentages to provide a supportive matrix with beneficial bulk electrical and mechanical properties. The substrates were tested for mechanical properties and conductivity. Astrocytes (glial scar tissue-forming cells) were seeded onto the substrates for adhesion. Results provided the first evidence that astrocytes preferentially adhered to the composite material that contained the lowest weight percentage of carbon nanofibers. Positive interactions with neurons, and, at the same time, limited astrocyte functions leading to decreased gliotic scar tissue formation are essential for increased neuronal implant efficacy.

Streblus asper Lour. exerts MAPK and SKN-1 mediated anti-aging, anti-photoaging activities and imparts neuroprotection by ameliorating Aβ in Caenorhabditis elegans

2021 ◽  
pp. 1-17
Author(s):  
Mani Iyer Prasanth ◽  
James Michael Brimson ◽  
Dicson Sheeja Malar ◽  
Anchalee Prasansuklab ◽  
Tewin Tencomnao
Keyword(s):  
Oxidative Stress ◽  
Caenorhabditis Elegans ◽  
Stress Resistance ◽  
Vital Role ◽  
Transgenic Strain ◽  
Wild Type ◽  
Neuroprotective Effects ◽  
C Elegans ◽  
Streblus Asper

BACKGROUND: Streblus asper Lour., has been reported to have anti-aging and neuroprotective efficacies in vitro. OBJECTIVE: To analyze the anti-aging, anti-photoaging and neuroprotective efficacies of S. asper in Caenorhabditis elegans. METHODS: C. elegans (wild type and gene specific mutants) were treated with S. asper extract and analyzed for lifespan and other health benefits through physiological assays, fluorescence microscopy, qPCR and Western blot. RESULTS: The plant extract was found to increase the lifespan, reduce the accumulation of lipofuscin and modulate the expression of candidate genes. It could extend the lifespan of both daf-16 and daf-2 mutants whereas the pmk-1 mutant showed no effect. The activation of skn-1 was observed in skn-1::GFP transgenic strain and in qPCR expression. Further, the extract can extend the lifespan of UV-A exposed nematodes along with reducing ROS levels. Additionally, the extract also extends lifespan and reduces paralysis in Aβ transgenic strain, apart from reducing Aβ expression. CONCLUSIONS: S. asper was able to extend the lifespan and healthspan of C. elegans which was independent of DAF-16 pathway but dependent on SKN-1 and MAPK which could play a vital role in eliciting the anti-aging, anti-photoaging and neuroprotective effects, as the extract could impart oxidative stress resistance and neuroprotection.

scholarly journals LRIG1 is a conserved EGFR regulator involved in melanoma development, survival and treatment resistance

Oncogene ◽  
2021 ◽  
Author(s):  
Ola Billing ◽  
Ylva Holmgren ◽  
Daniel Nosek ◽  
Håkan Hedman ◽  
Oskar Hemmingsson
Keyword(s):  
Growth Factor Receptor ◽  
Braf Inhibitor ◽  
Treatment Resistance ◽  
Negative Regulator ◽  
C Elegans ◽  
Rtk Signaling ◽  
Egfr Expression ◽  
Inhibitor Resistance ◽  
Leucine Rich Repeats

AbstractLeucine-rich repeats and immunoglobulin-like domains 1 (LRIG1) is a pan-negative regulator of receptor tyrosine kinase (RTK) signaling and a tumor suppressor in several cancers, but its involvement in melanoma is largely unexplored. Here, we aim to determine the role of LRIG1 in melanoma tumorigenesis, RTK signaling, and BRAF inhibitor resistance. We find that LRIG1 is downregulated during early tumorigenesis and that LRIG1 affects activation of the epidermal growth factor receptor (EGFR) in melanoma cells. LRIG1-dependent regulation of EGFR signaling is evolutionary conserved to the roundworm C. elegans, where negative regulation of the EGFR-Ras-Raf pathway by sma-10/LRIG completely depends on presence of the receptor let-23/EGFR. In a cohort of metastatic melanoma patients, we observe an association between LRIG1 and survival in the triple wild-type subtype and in tumors with high EGFR expression. During in vitro development of BRAF inhibitor resistance, LRIG1 expression decreases; and mimics LRIG1 knockout cells for increased EGFR expression. Treating resistant cells with recombinant LRIG1 suppresses AKT activation and proliferation. Together, our results show that sma-10/LRIG is a conserved regulator of RTK signaling, add to our understanding of LRIG1 in melanoma and identifies recombinant LRIG1 as a potential therapeutic against BRAF inhibitor-resistant melanoma.

scholarly journals The double-stranded DNA-binding proteins TEBP-1 and TEBP-2 form a telomeric complex with POT-1

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Sabrina Dietz ◽  
Miguel Vasconcelos Almeida ◽  
Emily Nischwitz ◽  
Jan Schreier ◽  
Nikenza Viceconte ◽  
...  
Keyword(s):  
Quantitative Proteomics ◽  
Wild Type ◽  
C Elegans ◽  
Double Stranded Dna ◽  
Telomere Sequence ◽  
Proteomics Approach ◽  
Telomeric Protein ◽  
Regulate Telomere Length

AbstractTelomeres are bound by dedicated proteins, which protect them from DNA damage and regulate telomere length homeostasis. In the nematode Caenorhabditis elegans, a comprehensive understanding of the proteins interacting with the telomere sequence is lacking. Here, we harnessed a quantitative proteomics approach to identify TEBP-1 and TEBP-2, two paralogs expressed in the germline and embryogenesis that associate to telomeres in vitro and in vivo. tebp-1 and tebp-2 mutants display strikingly distinct phenotypes: tebp-1 mutants have longer telomeres than wild-type animals, while tebp-2 mutants display shorter telomeres and a Mortal Germline. Notably, tebp-1;tebp-2 double mutant animals have synthetic sterility, with germlines showing signs of severe mitotic and meiotic arrest. Furthermore, we show that POT-1 forms a telomeric complex with TEBP-1 and TEBP-2, which bridges TEBP-1/-2 with POT-2/MRT-1. These results provide insights into the composition and organization of a telomeric protein complex in C. elegans.

Focal TLR4 activation mediates disturbed flow-induced endothelial inflammation

2019 ◽  
Vol 116 (1) ◽  
pp. 226-236 ◽  
Author(s):  
Dan Qu ◽  
Li Wang ◽  
Mingyu Huo ◽  
Wencong Song ◽  
Chi-Wai Lau ◽  
...  
Keyword(s):  
Vascular Inflammation ◽  
Subsequent Development ◽  
Tlr4 Expression ◽  
Disturbed Flow ◽  
Atherosclerotic Lesions ◽  
Endothelial Inflammation ◽  
En Face ◽  
Induced Flow ◽  
Tlr4 Activation

Abstract Aims Disturbed blood flow at arterial branches and curvatures modulates endothelial function and predisposes the region to endothelial inflammation and subsequent development of atherosclerotic lesions. Activation of the endothelial Toll-like receptors (TLRs), in particular TLR4, contributes to vascular inflammation. Therefore, we investigate whether TLR4 can sense disturbed flow (DF) to mediate the subsequent endothelial inflammation. Methods and results En face staining of endothelium revealed that TLR4 expression, activation, and its downstream inflammatory markers were elevated in mouse aortic arch compared with thoracic aorta, which were absent in Tlr4mut mice. Similar results were observed in the partial carotid ligation model where TLR4 signalling was activated in response to ligation-induced flow disturbance in mouse carotid arteries, and such effect was attenuated in Tlr4mut mice. DF in vitro increased TLR4 expression and activation in human endothelial cells (ECs) and promoted monocyte-EC adhesion, which were inhibited in TLR4-knockdown ECs. Among endogenous TLR4 ligands examined as candidate mediators of DF-induced TLR4 activation, fibronectin containing the extra domain A (FN-EDA) expressed by ECs was increased by DF and was revealed to directly interact with and activate TLR4. Conclusion Our findings demonstrate the indispensable role of TLR4 in DF-induced endothelial inflammation and pinpoint FN-EDA as the endogenous TLR4 activator in this scenario. This novel mechanism of vascular inflammation under DF condition may serve as a critical initiating step in atherogenesis.

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