scholarly journals WASP and SCAR are evolutionarily conserved in actin-filled pseudopod-based motility

2017 ◽  
Vol 216 (6) ◽  
pp. 1673-1688 ◽  
Author(s):  
Lillian K. Fritz-Laylin ◽  
Samuel J. Lord ◽  
R. Dyche Mullins
Keyword(s):  
Actin Polymerization ◽  
Eukaryotic Cells ◽  
Human Neutrophils ◽  
Evolutionary Relationships ◽  
Actin Assembly ◽  
Complex Environments ◽  
Clear Trend ◽  
Cell Crawling ◽  
Evolutionarily Conserved ◽  
Underlying Mechanisms

Diverse eukaryotic cells crawl through complex environments using distinct modes of migration. To understand the underlying mechanisms and their evolutionary relationships, we must define each mode and identify its phenotypic and molecular markers. In this study, we focus on a widely dispersed migration mode characterized by dynamic actin-filled pseudopods that we call “α-motility.” Mining genomic data reveals a clear trend: only organisms with both WASP and SCAR/WAVE—activators of branched actin assembly—make actin-filled pseudopods. Although SCAR has been shown to drive pseudopod formation, WASP’s role in this process is controversial. We hypothesize that these genes collectively represent a genetic signature of α-motility because both are used for pseudopod formation. WASP depletion from human neutrophils confirms that both proteins are involved in explosive actin polymerization, pseudopod formation, and cell migration. WASP and WAVE also colocalize to dynamic signaling structures. Moreover, retention of WASP together with SCAR correctly predicts α-motility in disease-causing chytrid fungi, which we show crawl at >30 µm/min with actin-filled pseudopods. By focusing on one migration mode in many eukaryotes, we identify a genetic marker of pseudopod formation, the morphological feature of α-motility, providing evidence for a widely distributed mode of cell crawling with a single evolutionary origin.

scholarly journals WASP and SCAR are evolutionarily conserved in actin-filled pseudopod-based motility

2016 ◽  
Author(s):  
Lillian K. Fritz-Laylin ◽  
Samuel J. Lord ◽  
R. Dyche Mullins
Keyword(s):  
Actin Polymerization ◽  
Eukaryotic Cells ◽  
Human Neutrophils ◽  
Evolutionary Relationships ◽  
Actin Assembly ◽  
Complex Environments ◽  
Clear Trend ◽  
Cell Crawling ◽  
Evolutionarily Conserved ◽  
Underlying Mechanisms

AbstractDiverse eukaryotic cells crawl through complex environments using distinct modes of migration. To understand the underlying mechanisms and their evolutionary relationships, we must define each mode, and identify its phenotypic and molecular markers. Here, we focus on a widely dispersed migration mode characterized by dynamic, actin-filled pseudopods that we call “α-motility.” Mining genomic data reveals a clear trend: only organisms with both WASP and SCAR/WAVE—activators of branched actin assembly—make actin-filled pseudopods. While SCAR has been shown to drive pseudopod formation, WASP’s role in this process is controversial. We hypothesize that these genes together represent a genetic signature of α-motility, because both are used for pseudopod formation. WASP depletion from human neutrophils confirms that both proteins are involved in explosive actin polymerization, pseudopod formation, and cell migration, and colocalize to dynamic signaling structures. Moreover, retention of WASP together with SCAR correctly predicts α-motility in disease-causing chytrid fungi, which we show crawl at >30 μm/min with actin-filled pseudopods. By focusing on one migration mode in many eukaryotes, we identify a genetic marker of pseudopod formation, the morphological feature of α-motility, providing evidence for a widely distributed mode of cell crawling with a single evolutionary origin.

scholarly journals Beta 2 integrin engagement triggers actin polymerization and phosphatidylinositol trisphosphate formation in non-adherent human neutrophils.

1993 ◽  
Vol 123 (6) ◽  
pp. 1597-1605 ◽  
Author(s):  
R Löfgren ◽  
J Ng-Sikorski ◽  
A Sjölander ◽  
T Andersson
Keyword(s):  
Actin Polymerization ◽  
Cellular Level ◽  
Human Neutrophils ◽  
Induced Response ◽  
Actin Assembly ◽  
Filamentous Actin ◽  
Integrin Receptors ◽  
Absolute Requirement ◽  
Beta 2

Beta 2 integrins are involved in the adhesion of leukocytes to other cells and surfaces. Although adhesion is required for cell locomotion, little is known regarding the way beta 2 integrin-receptors affect the actin network in leukocytes. In the present study filamentous actin (F-actin) levels in non-adherent human neutrophils have been measured by phalloidin staining after antibody cross-linking of beta 2 integrins. Antibody engagement of beta 2 integrins resulted in a rapid and sustained (146 and 131% after 30 and 300 s, respectively) increase in the neutrophil F-actin content. This is in contrast to stimulation with N-formyl-l-methionyl-l-leucyl-l-phenylalanine (fMLP), which causes a prompt and pronounced but rapidly declining rise in F-actin (214 and 127% after 15 and 300 s, respectively). Priming neutrophils with 1 nM PMA, a low concentration that did not influence the F-actin content per se, increased the magnitude of the beta 2 integrin-induced response but had no effect on the kinetics (199% after 30 s and 169% after 300 s). Removal of extracellular Ca2+ only marginally affected the beta 2 integrin-induced F-actin response for cells that were pretreated with PMA whereas the response for nonprimed cells was reduced by half. This suggests that even though extracellular Ca2+ has a modulatory effect it is not an absolute requirement for beta 2 integrin-induced actin polymerization. beta 2 integrin engagement did not affect the resting cellular level of cAMP arguing against a role of cAMP in beta 2 integrin-induced actin assembly.(ABSTRACT TRUNCATED AT 250 WORDS)

scholarly journals The Human Arp2/3 Complex Is Composed of Evolutionarily Conserved Subunits and Is Localized to Cellular Regions of Dynamic Actin Filament Assembly

1997 ◽  
Vol 138 (2) ◽  
pp. 375-384 ◽  
Author(s):  
Matthew D. Welch ◽  
Angela H. DePace ◽  
Suzie Verma ◽  
Akihiro Iwamatsu ◽  
Timothy J. Mitchison
Keyword(s):  
Actin Polymerization ◽  
Predicted Amino Acid Sequence ◽  
Actin Assembly ◽  
Filament Assembly ◽  
Evolutionarily Conserved ◽  
And Function ◽  
Related Proteins ◽  
Human Complex ◽  
Novel Protein ◽  
In Cells

The Arp2/3 protein complex has been implicated in the control of actin polymerization in cells. The human complex consists of seven subunits which include the actin related proteins Arp2 and Arp3, and five others referred to as p41-Arc, p34-Arc, p21-Arc, p20-Arc, and p16-Arc (Arp complex). We have determined the predicted amino acid sequence of all seven subunits. Each has homologues in diverse eukaryotes, implying that the structure and function of the complex has been conserved through evolution. Human Arp2 and Arp3 are very similar to family members from other species. p41-Arc is a new member of the Sop2 family of WD (tryptophan and aspartate) repeat–containing proteins and may be posttranslationally modified, suggesting that it may be involved in regulating the activity and/or localization of the complex. p34-Arc, p21-Arc, p20-Arc, and p16-Arc define novel protein families. We sought to evaluate the function of the Arp2/3 complex in cells by determining its intracellular distribution. Arp3, p34-Arc, and p21-Arc were localized to the lamellipodia of stationary and locomoting fibroblasts, as well to Listeria monocytogenes assembled actin tails. They were not detected in cellular bundles of actin filaments. Taken together with the ability of the Arp2/3 complex to induce actin polymerization, these observations suggest that the complex promotes actin assembly in lamellipodia and may participate in lamellipodial protrusion.

scholarly journals Actin assembly in electropermeabilized neutrophils: role of intracellular calcium.

1990 ◽  
Vol 110 (6) ◽  
pp. 1975-1982 ◽  
Author(s):  
G P Downey ◽  
C K Chan ◽  
S Trudel ◽  
S Grinstein
Keyword(s):  
Actin Polymerization ◽  
Cytosolic Calcium ◽  
Leukotriene B4 ◽  
Human Neutrophils ◽  
Actin Assembly ◽  
Intact Cells ◽  
Permeabilized Cells ◽  
Sequence Of Events ◽  
Formyl Peptide

Assembly of microfilaments involves the conversion of actin from the monomeric (G) to the filamentous (F) form. The exact sequence of events responsible for this conversion is yet to be defined and, in particular, the role of calcium remains unclear. Intact and electropermeabilized human neutrophils were used to assess more directly the role of cytosolic calcium [( Ca2+]i) in actin assembly. Staining with 7-nitrobenz-2-oxa-1,3-diazole-phallacidin and right angle light scattering were used to monitor the formation of F-actin. Though addition of Ca2+ ionophores can be known to induce actin assembly, the following observations suggest that an increased [Ca2+]i is not directly responsible for receptor-induced actin polymerization: (a) intact cells in Ca2(+)-free medium, depleted of internal Ca2+ by addition of ionophore, responded to the formyl peptide fMLP with actin assembly despite the absence of changes in [Ca2+]i, assessed with Indo-1; (b) fMLP induced a significant increase in F-actin content in permeabilized cells equilibrated with medium containing 0.1 microM free Ca2+, buffered with up to 10 mM EGTA; (c) increasing [Ca2+]i beyond the resting level by direct addition of CaCl2 to permeabilized cells resulted in actin disassembly. Conversely, lowering [Ca2+]i resulted in spontaneous actin assembly. To reconcile these findings with the actin-polymerizing effects of Ca2+ ionophores, we investigated whether A23187 and ionomycin induced actin assembly by a mechanism independent of, or secondary to the increase in [Ca2+]i. We found that the ionophore-induced actin assembly was completely inhibited by the leukotriene B4 (LTB4) antagonist LY-223982, implying that the ionophore effect was secondary to LTB4 formation, possibly by stimulation of phospholipase A2. We conclude that actin assembly is not mediated by an increase in [Ca2+]i, but rather that elevated [Ca2+]i facilitates actin disassembly, an effect possibly mediated by Ca2(+)-sensitive actin filament-severing proteins such as gelsolin. Sequential actin assembly and disassembly may be necessary for functions such as chemotaxis.

Metabolic amplifying pathway increases both phases of insulin secretion independently of β-cell actin microfilaments

2010 ◽  
Vol 299 (2) ◽  
pp. C389-C398 ◽  
Author(s):  
Nizar I. Mourad ◽  
Myriam Nenquin ◽  
Jean-Claude Henquin
Keyword(s):  
Insulin Secretion ◽  
Actin Polymerization ◽  
Second Phase ◽  
Actin Microfilaments ◽  
Actin Depolymerization ◽  
Inhibitory Function ◽  
Insulin Granules ◽  
Priming Process ◽  
Underlying Mechanisms ◽  
Amplifying Pathway

Two pathways control glucose-induced insulin secretion (IS) by β-cells. The triggering pathway involves ATP-sensitive potassium (KATP) channel-dependent depolarization, Ca2+ influx, and a rise in the cytosolic Ca2+ concentration ([Ca2+]c), which triggers exocytosis of insulin granules. The metabolic amplifying pathway augments IS without further increasing [Ca2+]c. The underlying mechanisms are unknown. Here, we tested the hypothesis that amplification implicates actin microfilaments. Mouse islets were treated with latrunculin B and cytochalasin B to depolymerize actin or jasplakinolide to polymerize actin. They were then perifused to measure [Ca2+]c and IS. Metabolic amplification was studied during imposed steady elevation of [Ca2+]c by tolbutamide or KCl or by comparing the magnitude of [Ca2+]c and IS changes produced by glucose and tolbutamide. Both actin polymerization and depolymerization augmented IS triggered by all stimuli without increasing (sometimes decreasing) [Ca2+]c, which indicates a predominantly inhibitory function of microfilaments in exocytosis at a step distal to [Ca2+]c increase. When [Ca2+]c was elevated and controlled by KCl or tolbutamide, the amplifying action of glucose was facilitated by actin depolymerization and unaffected by polymerization. Both phases of IS were larger in response to high-glucose than to tolbutamide in low-glucose, although triggering [Ca2+]c was lower. This difference in IS, due to amplification, persisted when the IS rate was doubled by actin depolymerization or polymerization. In conclusion, metabolic amplification is rapid and influences the first as well as the second phase of IS. It is a late step of stimulus-secretion coupling, which does not require functional actin microfilaments and could correspond to acceleration of the priming process conferring release competence to insulin granules.

Actin Polymerization, Calcium-Transients, and Phospholipid Metabolism in Human Neutrophils After Stimulation with Interleukin-8 and N-formyl Peptide

1994 ◽  
Vol 102 (3) ◽  
pp. 310-314 ◽  
Author(s):  
Johannes Norgauer ◽  
Jean Krutmann ◽  
Gustav J. Dobos ◽  
Alexis E. Traynor-Kaplan ◽  
Zenaida G. Oades ◽  
...  
Keyword(s):  
Actin Polymerization ◽  
Phospholipid Metabolism ◽  
Interleukin 8 ◽  
Calcium Transients ◽  
Human Neutrophils ◽  
Formyl Peptide

Difference of MreBCD Complex Interactome in Salmonella Typhimurium ST19 and ST213 Genotypes on Pathogenesis and Stress Response Pathways

2021 ◽  
Vol 22 ◽  
Author(s):  
Reyna Cristina Zepeda-Gurrola ◽  
Gerardo Vázquez-Marrufo ◽  
Xianwu Guo ◽  
Isabel Cristina Rodríguez-Luna ◽  
Alejandro Sánchez-Varela ◽  
...  
Keyword(s):  
Stress Response ◽  
Bacterial Virulence ◽  
Eukaryotic Cells ◽  
Interaction Patterns ◽  
High Infection Rate ◽  
Differential Interaction ◽  
Sequence Variations ◽  
High Infection ◽  
Underlying Mechanisms

: Salmonella enterica is the etiological agent of salmonellosis, with a high infection rate worldwide. In Mexico, ST213 genotype of S. enterica ser. Typhimurium is displacing the ancestral ST19 genotype. Bacterial cytoskeleton protein complex MreBCD play an important role in S. enterica pathogenesis, but underlying mechanisms are unknown. In this study, 106 interactions among MreBCD and 15 proteins from S. Typhimurium Pathogenicity Islands 1 (SP-I) and 2 (SP-2) involved in both bacterial virulence and stress response were predicted in ST213 and ST19 genotypes, of which 12 interactions were confirmed in vitro. In addition, gene cluster analysis in 100 S. Typhimurium genomes was performed for these genes. The in silico and in vitro results showed a novel MreBCD interactome involved in the regulation of pathogenesis and stress response through interactions with virulence factors located at SPI-1 and SPI-2. Furthermore, both pseudogene presence and sequence variations in four tested proteins between genotypes resulted in differential interaction patterns that are involved in Salmonella motility and survival in eukaryotic cells, which could explain replacement of ST19 by ST213 in Mexico.

The protein kinase C inhibitor H-7 activates human neutrophils: effect on shape, actin polymerization, fluid pinocytosis and locomotion

1990 ◽  
Vol 96 (1) ◽  
pp. 99-106
Author(s):  
H.U. Keller ◽  
V. Niggli ◽  
A. Zimmermann ◽  
R. Portmann
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Actin Polymerization ◽  
Kinase Inhibitor ◽  
Human Neutrophils ◽  
Kinase C ◽  
Chemotactic Peptides ◽  
Protein Kinase C Inhibitor ◽  
Shape Changes ◽  
Stimulation Of

The present study demonstrates new properties of H-7. The protein kinase inhibitor H-7 is a potent activator of several neutrophil functions. Stimulation of initially spherical nonmotile neutrophils elicits vigorous shape changes within a few seconds, increases in cytoskeletal actin, altered F-actin distribution, increased adhesiveness and a relatively small increase in pinocytic activity. H-7 has also chemokinetic activities. Depending on the experimental condition, H-7 may elicit or inhibit neutrophil locomotion. It failed to induce chemotaxis. Thus, the response pattern elicited by H-7 is different from that of other leukocyte activators such as chemotactic peptides, PMA or diacylglycerols. The finding that H-7 can elicit shape changes, actin polymerization and pinocytosis suggests that these events can occur without activation of protein kinase C (PKC). PMA-induced shape changes and stimulation of pinocytosis were not inhibited by H-7.

scholarly journals Activation of human neutrophils by mastoparan. Reorganization of the cytoskeleton, formation of phosphatidylinositol 3,4,5-trisphosphate, secretion up-regulation of complement receptor type 3 and superoxide anion production are stimulated by mastoparan

1992 ◽  
Vol 282 (2) ◽  
pp. 393-397 ◽  
Author(s):  
J Norgauer ◽  
M Eberle ◽  
H D Lemke ◽  
K Aktories
Keyword(s):  
Pertussis Toxin ◽  
Actin Polymerization ◽  
Cytochalasin B ◽  
Superoxide Radical ◽  
Human Neutrophils ◽  
Superoxide Anion Production ◽  
Radical Production ◽  
Rapid Formation ◽  
Primary Granules ◽  
Type 3

In human neutrophils, mastoparan induced rapid F-actin polymerization which was followed by a slow and sustained depolymerization to below the initial F-actin content. Incubation of neutrophils with pertussis toxin inhibited mastoparan-stimulated actin polymerization; however it did not prevent sustained depolymerization of F-actin. Analyses of phospholipids performed in parallel revealed that mastoparan stimulated rapid formation of phosphatidylinositol 3,4,5-trisphosphate (PIP3) and consumption of phosphatidylinositol 4,5-bisphosphate (PIP2). Pertussis toxin treatment blocked mastoparan-induced formation of PIP3. Furthermore, mastoparan stimulated the release of N-acetylglucosaminidase from primary granules. Cytochalasin B enhanced mastoparan-stimulated secretion. Mastoparan triggered superoxide radical production in a cytochalasin B-sensitive manner and induced complement type 3 receptor (CR3) up-regulation.

scholarly journals Coordinated regulation of platelet actin filament barbed ends by gelsolin and capping protein.

1996 ◽  
Vol 134 (2) ◽  
pp. 389-399 ◽  
Author(s):  
K Barkalow ◽  
W Witke ◽  
D J Kwiatkowski ◽  
J H Hartwig
Keyword(s):  
Actin Cytoskeleton ◽  
Actin Filament ◽  
Actin Polymerization ◽  
Residual Activity ◽  
Human Platelets ◽  
Actin Assembly ◽  
Capping Protein ◽  
Capping Proteins ◽  
End Capping ◽  
Human And Mouse

Exposure of cryptic actin filament fast growing ends (barbed ends) initiates actin polymerization in stimulated human and mouse platelets. Gelsolin amplifies platelet actin assembly by severing F-actin and increasing the number of barbed ends. Actin filaments in stimulated platelets from transgenic gelsolin-null mice elongate their actin without severing. F-actin barbed end capping activity persists in human platelet extracts, depleted of gelsolin, and the heterodimeric capping protein (CP) accounts for this residual activity. 35% of the approximately 5 microM CP is associated with the insoluble actin cytoskeleton of the resting platelet. Since resting platelets have an F-actin barbed end concentration of approximately 0.5 microM, sufficient CP is bound to cap these ends. CP is released from OG-permeabilized platelets by treatment with phosphatidylinositol 4,5-bisphosphate or through activation of the thrombin receptor. However, the fraction of CP bound to the actin cytoskeleton of thrombin-stimulated mouse and human platelets increases rapidly to approximately 60% within 30 s. In resting platelets from transgenic mice lacking gelsolin, which have 33% more F-actin than gelsolin-positive cells, there is a corresponding increase in the amount of CP associated with the resting cytoskeleton but no change with stimulation. These findings demonstrate an interaction between the two major F-actin barbed end capping proteins of the platelet: gelsolin-dependent severing produces barbed ends that are capped by CP. Phosphatidylinositol 4,5-bisphosphate release of gelsolin and CP from platelet cytoskeleton provides a mechanism for mediating barbed end exposure. After actin assembly, CP reassociates with the new actin cytoskeleton.

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