scholarly journals The Beta-Tubulin Isotype TUBB6 Controls Microtubule and Actin Dynamics in Osteoclasts

2021 ◽  
Vol 9 ◽  
Author(s):  
Justine Maurin ◽  
Anne Morel ◽  
David Guérit ◽  
Julien Cau ◽  
Serge Urbach ◽  
...  
Keyword(s):  
Bone Resorption ◽  
Actin Cytoskeleton ◽  
Molecular Mechanisms ◽  
Life Time ◽  
Negative Regulator ◽  
Actin Dynamics ◽  
Growth Speed ◽  
Actin Organization ◽  
Cytoskeleton Organization ◽  
Tubulin Isotype

Osteoclasts are bone resorbing cells that participate in the maintenance of bone health. Pathological increase in osteoclast activity causes bone loss, eventually resulting in osteoporosis. Actin cytoskeleton of osteoclasts organizes into a belt of podosomes, which sustains the bone resorption apparatus and is maintained by microtubules. Better understanding of the molecular mechanisms regulating osteoclast cytoskeleton is key to understand the mechanisms of bone resorption, in particular to propose new strategies against osteoporosis. We reported recently that β-tubulin isotype TUBB6 is key for cytoskeleton organization in osteoclasts and for bone resorption. Here, using an osteoclast model CRISPR/Cas9 KO for Tubb6, we show that TUBB6 controls both microtubule and actin dynamics in osteoclasts. Osteoclasts KO for Tubb6 have reduced microtubule growth speed with longer growth life time, higher levels of acetylation, and smaller EB1-caps. On the other hand, lack of TUBB6 increases podosome life time while the belt of podosomes is destabilized. Finally, we performed proteomic analyses of osteoclast microtubule-associated protein enriched fractions. This highlighted ARHGAP10 as a new microtubule-associated protein, which binding to microtubules appears to be negatively regulated by TUBB6. ARHGAP10 is a negative regulator of CDC42 activity, which participates in actin organization in osteoclasts. Our results suggest that TUBB6 plays a key role in the control of microtubule and actin cytoskeleton dynamics in osteoclasts. Moreover, by controlling ARHGAP10 association with microtubules, TUBB6 may participate in the local control of CDC42 activity to ensure efficient bone resorption.

scholarly journals The beta-tubulin isotype TUBB6 controls microtubule and actin dynamics in osteoclasts.

2021 ◽  
Author(s):  
Justine Maurin ◽  
Anne Morel ◽  
David Guérit ◽  
Julien Cau ◽  
Serge Urbach ◽  
...  
Keyword(s):  
Bone Resorption ◽  
Actin Cytoskeleton ◽  
Molecular Mechanisms ◽  
Life Time ◽  
Negative Regulator ◽  
Actin Dynamics ◽  
Growth Speed ◽  
Actin Organization ◽  
Cytoskeleton Organization ◽  
Tubulin Isotype

Osteoclasts are bone resorbing cells that participate in the maintenance of bone health. Pathological increase in osteoclast activity causes bone loss eventually resulting in osteoporosis. Actin cytoskeleton of osteoclasts organizes into a belt of podosomes, which sustains the bone resorption apparatus and is maintained by microtubules. Better understanding of the molecular mechanisms regulating osteoclast cytoskeleton is key to understand the mechanisms of bone resorption, in particular to propose new strategies against osteoporosis. We reported recently that β-tubulin isotype TUBB6 is key for cytoskeleton organization in osteoclasts and for bone resorption. Here, using an osteoclast model CRISPR/Cas9 KO for Tubb6, we show that TUBB6 controls both microtubule and actin dynamics in osteoclasts. Osteoclasts KO for Tubb6 have reduced microtubule growth speed with longer growth life time, higher levels of acetylation and smaller EB1-caps. On the other hand, lack of TUBB6 increases podosome life time while the belt of podosomes is destabilized. Finally, we performed proteomic analyses of osteoclast microtubule-associated protein enriched fractions. This highlighted ARHGAP10 as a new microtubule-associated protein, which binding to microtubules appears to be negatively regulated by TUBB6. ARHGAP10 is a negative regulator of CDC42 activity, which participates in actin organization in osteoclasts. Our results suggest that TUBB6 plays a key role in the control of microtubule and actin cytoskeleton dynamics in osteoclasts. Moreover, by controlling ARHGAP10 association with microtubules, TUBB6 may participate in the local control CDC42 activity to ensure efficient bone resorption.

The actin cytoskeleton is required to elaborate and maintain spatial patterning during trichome cell morphogenesis in Arabidopsis thaliana

Development ◽  
1999 ◽  
Vol 126 (24) ◽  
pp. 5559-5568 ◽  
Author(s):  
J. Mathur ◽  
P. Spielhofer ◽  
B. Kost ◽  
N. Chua
Keyword(s):  
Arabidopsis Thaliana ◽  
Actin Cytoskeleton ◽  
Spatial Patterning ◽  
Actin Microfilaments ◽  
Cell Morphogenesis ◽  
Actin Organization ◽  
Cytoskeleton Organization ◽  
Branch Formation ◽  
Actin Cytoskeleton Organization ◽  
Trichome Cell

Arabidopsis thaliana trichomes provide an attractive model system to dissect molecular processes involved in the generation of shape and form in single cell morphogenesis in plants. We have used transgenic Arabidopsis plants carrying a GFP-talin chimeric gene to analyze the role of the actin cytoskeleton in trichome cell morphogenesis. We found that during trichome cell development the actin microfilaments assumed an increasing degree of complexity from fine filaments to thick, longitudinally stretched cables. Disruption of the F-actin cytoskeleton by actin antagonists produced distorted but branched trichomes which phenocopied trichomes of mutants belonging to the ‘distorted’ class. Subsequent analysis of the actin cytoskeleton in trichomes of the distorted mutants, alien, crooked, distorted1, gnarled, klunker and wurm uncovered actin organization defects in each case. Treatments of wild-type seedlings with microtubule-interacting drugs elicited a radically different trichome phenotype characterized by isotropic growth and a severe inhibition of branch formation; these trichomes did not show defects in actin cytoskeleton organization. A normal actin cytoskeleton was also observed in trichomes of the zwichel mutant which have reduced branching. ZWICHEL, which was previously shown to encode a kinesin-like protein is thought to be involved in microtubule-linked processes. Based on our results we propose that microtubules establish the spatial patterning of trichome branches whilst actin microfilaments elaborate and maintain the overall trichome pattern during development.

scholarly journals The Pleckstrin Homology Domain Proteins Slm1 and Slm2 Are Required for Actin Cytoskeleton Organization in Yeast and Bind Phosphatidylinositol-4,5-Bisphosphate and TORC2

2005 ◽  
Vol 16 (4) ◽  
pp. 1883-1900 ◽  
Author(s):  
Maria Fadri ◽  
Alexes Daquinag ◽  
Shimei Wang ◽  
Tao Xue ◽  
Jeannette Kunz
Keyword(s):  
Cell Growth ◽  
Actin Cytoskeleton ◽  
Membrane Dynamics ◽  
Effector Proteins ◽  
Pleckstrin Homology Domain ◽  
Ph Domain ◽  
Pleckstrin Homology ◽  
Yeast Saccharomyces Cerevisiae ◽  
Actin Organization ◽  
Cytoskeleton Organization

Phosphatidylinositol-4,5-bisphosphate [PtdIns(4,5)P2] is a key second messenger that regulates actin and membrane dynamics, as well as other cellular processes. Many of the effects of PtdIns(4,5)P2are mediated by binding to effector proteins that contain a pleckstrin homology (PH) domain. Here, we identify two novel effectors of PtdIns(4,5)P2in the budding yeast Saccharomyces cerevisiae: the PH domain containing protein Slm1 and its homolog Slm2. Slm1 and Slm2 serve redundant roles essential for cell growth and actin cytoskeleton polarization. Slm1 and Slm2 bind PtdIns(4,5)P2through their PH domains. In addition, Slm1 and Slm2 physically interact with Avo2 and Bit61, two components of the TORC2 signaling complex, which mediates Tor2 signaling to the actin cytoskeleton. Together, these interactions coordinately regulate Slm1 targeting to the plasma membrane. Our results thus identify two novel effectors of PtdIns(4,5)P2regulating cell growth and actin organization and suggest that Slm1 and Slm2 integrate inputs from the PtdIns(4,5)P2and TORC2 to modulate polarized actin assembly and growth.

scholarly journals Combined strategy of siRNA and osteoclast actin cytoskeleton automated imaging to identify novel regulators of bone resorption shows a non-mitotic function for anillin

2018 ◽  
Author(s):  
Justine Maurin ◽  
Anne Morel ◽  
Cedric Hassen-Khodja ◽  
Virginie Vives ◽  
Pierre Jurdic ◽  
...  
Keyword(s):  
Bone Resorption ◽  
Actin Cytoskeleton ◽  
Public Health Problem ◽  
Major Public Health Problem ◽  
Cytoskeleton Organization ◽  
Multinucleated Cells ◽  
Automated Imaging ◽  
Actin Cytoskeleton Organization ◽  
Set Up ◽  
Combined Strategy

AbstractOsteoclasts are the main cells responsible for the resorption of mineralized extracellular matrices. They are the major targets for anti-resorptive therapies to manage osteoporosis, a major public health problem. Osteoclasts are giant multinucleated cells that can organize their a unique adhesion structure based on a belt of podosomes, which is the keystone of the bone resorption apparatus. We combined differential transcriptomics and siRNA screening approaches to get a broader view of cytoskeletal regulators that participate in the control of osteoclast cytoskeleton and identify novel regulators of bone resorption by osteoclasts. We identified 20 new candidate regulators of osteoclasts cytoskeleton including Fkbp15, Spire1, Tacc2 and RalA, for which we confirmed they are necessary for proper organization of the podosome belt. We also showed that Anillin, well known for its role during cytokinesis, is essential in osteoclasts for correct podosome patterning and efficient bone resorption. In particular, Anillin controls the levels of the GTPase RhoA, a known regulator of osteoclast cytoskeleton and resorption activity. Finally, we set up and validated an automated imaging strategy based on open-source software for automatic and objective measurement of actin cytoskeleton organization in osteoclasts. We provide these pipelines that are useful to automatically assess the effect of collections of siRNAs or chemical compounds on osteoclast cytoskeleton or differentiation.

scholarly journals Gα12/13 regulate epiboly by inhibiting E-cadherin activity and modulating the actin cytoskeleton

2009 ◽  
Vol 184 (6) ◽  
pp. 909-921 ◽  
Author(s):  
Fang Lin ◽  
Songhai Chen ◽  
Diane S. Sepich ◽  
Jennifer Ray Panizzi ◽  
Sherry G. Clendenon ◽  
...  
Keyword(s):  
Actin Cytoskeleton ◽  
Molecular Mechanisms ◽  
Genetic Studies ◽  
Yolk Cell ◽  
Cytoskeleton Organization ◽  
Actin Cytoskeleton Organization ◽  
Cell Layers ◽  
Dependent Pathway ◽  
E Cadherin

Epiboly spreads and thins the blastoderm over the yolk cell during zebrafish gastrulation, and involves coordinated movements of several cell layers. Although recent studies have begun to elucidate the processes that underlie these epibolic movements, the cellular and molecular mechanisms involved remain to be fully defined. Here, we show that gastrulae with altered Gα12/13 signaling display delayed epibolic movement of the deep cells, abnormal movement of dorsal forerunner cells, and dissociation of cells from the blastoderm, phenocopying e-cadherin mutants. Biochemical and genetic studies indicate that Gα12/13 regulate epiboly, in part by associating with the cytoplasmic terminus of E-cadherin, and thereby inhibiting E-cadherin activity and cell adhesion. Furthermore, we demonstrate that Gα12/13 modulate epibolic movements of the enveloping layer by regulating actin cytoskeleton organization through a RhoGEF/Rho-dependent pathway. These results provide the first in vivo evidence that Gα12/13 regulate epiboly through two distinct mechanisms: limiting E-cadherin activity and modulating the organization of the actin cytoskeleton.

scholarly journals Cdc42-dependent actin dynamics controls maturation and secretory activity of dendritic cells

2015 ◽  
Vol 211 (3) ◽  
pp. 553-567 ◽  
Author(s):  
Anna M. Schulz ◽  
Susanne Stutte ◽  
Sebastian Hogl ◽  
Nancy Luckashenak ◽  
Diana Dudziak ◽  
...  
Keyword(s):  
Cell Surface ◽  
Mhc Class Ii ◽  
Molecular Mechanisms ◽  
Secretory Activity ◽  
Class Ii ◽  
Actin Dynamics ◽  
Actin Organization ◽  
Peptide Antigens ◽  
Enhanced Secretion ◽  
Guanosine Triphosphatase

Cell division cycle 42 (Cdc42) is a member of the Rho guanosine triphosphatase family and has pivotal functions in actin organization, cell migration, and proliferation. To further study the molecular mechanisms of dendritic cell (DC) regulation by Cdc42, we used Cdc42-deficient DCs. Cdc42 deficiency renders DCs phenotypically mature as they up-regulate the co-stimulatory molecule CD86 from intracellular storages to the cell surface. Cdc42 knockout DCs also accumulate high amounts of invariant chain–major histocompatibility complex (MHC) class II complexes at the cell surface, which cannot efficiently present peptide antigens (Ag’s) for priming of Ag-specific CD4 T cells. Proteome analyses showed a significant reduction in lysosomal MHC class II–processing proteins, such as cathepsins, which are lost from DCs by enhanced secretion. As these effects on DCs can be mimicked by chemical actin disruption, our results propose that Cdc42 control of actin dynamics keeps DCs in an immature state, and cessation of Cdc42 activity during DC maturation facilitates secretion as well as rapid up-regulation of intracellular molecules to the cell surface.

scholarly journals In Vivo Importance of Actin Nucleotide Exchange Catalyzed by Profilin

2000 ◽  
Vol 150 (4) ◽  
pp. 895-904 ◽  
Author(s):  
Amy K. Wolven ◽  
Lisa D. Belmont ◽  
Nicole M. Mahoney ◽  
Steven C. Almo ◽  
David G. Drubin
Keyword(s):  
Point Mutations ◽  
Fluid Phase ◽  
Intrinsic Rate ◽  
Actin Dynamics ◽  
Actin Binding ◽  
Nucleotide Exchange ◽  
Actin Organization ◽  
Cytoskeleton Organization

The actin monomer-binding protein, profilin, influences the dynamics of actin filaments in vitro by suppressing nucleation, enhancing nucleotide exchange on actin, and promoting barbed-end assembly. Profilin may also link signaling pathways to actin cytoskeleton organization by binding to the phosphoinositide PIP2 and to polyproline stretches on several proteins. Although activities of profilin have been studied extensively in vitro, the significance of each of these activities in vivo needs to be tested. To study profilin function, we extensively mutagenized the Saccharomyces cerevisiae profilin gene (PFY1) and examined the consequences of specific point mutations on growth and actin organization. The actin-binding region of profilin was shown to be critical in vivo. act1-157, an actin mutant with an increased intrinsic rate of nucleotide exchange, suppressed defects in actin organization, cell growth, and fluid-phase endocytosis of pfy1-4, a profilin mutant defective in actin binding. In reactions containing actin, profilin, and cofilin, profilin was required for fast rates of actin filament turnover. However, Act1-157p circumvented the requirement for profilin. Based on the results of these studies, we conclude that in living cells profilin promotes rapid actin dynamics by regenerating ATP actin from ADP actin–cofilin generated during filament disassembly.

Probing cytoplasmic organization and the actin cytoskeleton of plant cells with optical tweezers

2010 ◽  
Vol 38 (3) ◽  
pp. 823-828 ◽  
Author(s):  
Tijs Ketelaar ◽  
Hannie S. van der Honing ◽  
Anne Mie C. Emons
Keyword(s):  
Actin Cytoskeleton ◽  
Physical Properties ◽  
Optical Tweezers ◽  
Intracellular Transport ◽  
Plant Cells ◽  
Actin Dynamics ◽  
Actin Binding ◽  
Structural Basis ◽  
Actin Organization ◽  
Cytoplasmic Organization

In interphase plant cells, the actin cytoskeleton is essential for intracellular transport and organization. To fully understand how the actin cytoskeleton functions as the structural basis for cytoplasmic organization, both molecular and physical aspects of the actin organization have to be considered. In the present review, we discuss literature that gives an insight into how cytoplasmic organization is achieved and in which actin-binding proteins have been identified that play a role in this process. We discuss how physical properties of the actin cytoskeleton in the cytoplasm of live plant cells, such as deformability and elasticity, can be probed by using optical tweezers. This technique allows non-invasive manipulation of cytoplasmic organization. Optical tweezers, integrated in a confocal microscope, can be used to manipulate cytoplasmic organization while studying actin dynamics. By combining this with mutant studies and drug applications, insight can be obtained about how the physical properties of the actin cytoskeleton, and thus the cytoplasmic organization, are influenced by different cellular processes.

scholarly journals MLK3 Is Associated With Poor Prognosis in Patients With Glioblastomas and Actin Cytoskeleton Remodeling in Glioblastoma Cells

2021 ◽  
Vol 10 ◽  
Author(s):  
Yan Zhu ◽  
Jin-Min Sun ◽  
Zi-Chen Sun ◽  
Feng-Jiao Chen ◽  
Yong-Ping Wu ◽  
...  
Keyword(s):  
Actin Cytoskeleton ◽  
Poor Prognosis ◽  
Molecular Mechanisms ◽  
Growth Factor Receptor ◽  
Recurrent Glioblastoma ◽  
Migration And Invasion ◽  
Breast Cancers ◽  
Recurrent Glioblastoma Multiforme ◽  
Genetic Ablation ◽  
Cytoskeleton Organization

Mixed lineage kinase 3 (MLK3) has been implicated in human melanoma and breast cancers. However, the clinical significance of MLK3 in human gliomas and the underlying cellular and molecular mechanisms remain unclear. We found that MLK3 proteins were highly expressed in high-grade human glioma specimens and especially prevalent in primary and recurrent glioblastoma multiforme (GBM). High levels of MLK3 mRNA were correlated with poor prognosis in patients with isocitrate dehydrogenase (IDH)-wild-type (wt) gliomas. Furthermore, genetic ablation of MLK3 significantly suppressed the migration and invasion abilities of GBM cells and disrupted actin cytoskeleton organization. Importantly, MLK3 directly bound to epidermal growth factor receptor kinase substrate 8 (EPS8) and regulated the cellular location of EPS8, which is essential for actin cytoskeleton rearrangement. Overall, these findings provide evidence that MLK3 upregulation predicts progression and poor prognosis in human IDH-wt gliomas and suggest that MLK3 promotes the migration and invasion of GBM cells by remodeling the actin cytoskeleton via MLK3-EPS8 signaling.

scholarly journals RhoE Regulates Actin Cytoskeleton Organization and Cell Migration

1998 ◽  
Vol 18 (8) ◽  
pp. 4761-4771 ◽  
Author(s):  
Rosa M. Guasch ◽  
Peter Scambler ◽  
Gareth E. Jones ◽  
Anne J. Ridley
Keyword(s):  
Actin Cytoskeleton ◽  
Mdck Cells ◽  
Fiber Formation ◽  
Stress Fibers ◽  
Complete Disappearance ◽  
Actin Organization ◽  
Cytoskeleton Organization ◽  
Actin Cytoskeleton Organization ◽  
Rho Family ◽  
As Stress

ABSTRACT The actin cytoskeleton is regulated by Rho family proteins: in fibroblasts, Rho mediates the formation of actin stress fibers, whereas Rac regulates lamellipodium formation and Cdc42 controls filopodium formation. We have cloned the mouse RhoE gene, whose product is a member of the Rho family that shares (except in one amino acid) the conserved effector domain of RhoA, RhoB, and RhoC. RhoE is able to bind GTP but does not detectably bind GDP and has low intrinsic GTPase activity compared with Rac. The role of RhoE in regulating actin organization was investigated by microinjection in Bac1.2F5 macrophages and MDCK cells. In macrophages, RhoE induced actin reorganization, leading to the formation of extensions resembling filopodia and pseudopodia. In MDCK cells, RhoE induced the complete disappearance of stress fibers, together with cell spreading. However, RhoE did not detectably affect the actin bundles that run parallel to the outer membranes of cells at the periphery of colonies, which are known to be dependent on RhoA. In addition, RhoE induced an increase in the speed of migration of hepatocyte growth factor/scatter factor-stimulated MDCK cells, in contrast to the previously reported inhibition produced by activated RhoA. The subcellular localization of RhoE at the lateral membranes of MDCK cells suggests a role in cell-cell adhesion, as has been shown for RhoA. These results suggest that RhoE may act to inhibit signalling downstream of RhoA, altering some RhoA-regulated responses, such as stress fiber formation, but not affecting others, such as peripheral actin bundle formation.

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