Lymphocyte-Specific Protein 1 Expression in Eukaryotic Cells Reproduces the Morphologic and Motile Abnormality of NAD 47/89 Neutrophils

Blood ◽  
1998 ◽  
Vol 91 (12) ◽  
pp. 4786-4795 ◽  
Author(s):  
Thomas H. Howard ◽  
John Hartwig ◽  
Casey Cunningham
Keyword(s):  
Actin Polymerization ◽  
Cell Function ◽  
Specific Protein ◽  
Polymorphonuclear Neutrophils ◽  
Actin Binding ◽  
Actin Network ◽  
Cell Surfaces ◽  
Actin Bundles ◽  
Mixed Polarity ◽  
Low Levels

Abstract Despite its name, the actin-binding protein lymphocyte-specific protein1 (LSP1) is found in all hematopoetic cells, and yet its role in cell function remains unclear. Recently, LSP1 was identified as the 47-kD protein overexpressed in the polymorphonuclear neutrophils of patients with a rare neutrophil disorder, neutrophil actin dysfunction with abnormalities of 47-kD and 89-kD proteins (NAD 47/89). These neutrophils are immotile, defective in actin polymerization in response to agonists, and display distinctive, fine, “hairlike” F-actin-rich projections on their cell surfaces. We now show that overexpression of LSP1 produces F-actin bundles that are likely responsible for the morphologic and motile abnormalities characteristic of the NAD 47/89 phenotype. Coincident with LSP1 overexpression, cells from each of several different eukaryotic lines, including a highly motile human melanoma line, develop hairlike surface projections that branch distinctively and contain F-actin and LSP1. The hairlike projections are supported at their core by thick actin bundles, composed of actin filaments of mixed polarity, which periodically anastomose to generate a branching structure. The motility of the melanoma cells is inhibited even at low levels of LSP1 expression. Therefore, these studies show that overexpression of LSP1 alone can recreate the morphologic and motile defects seen in NAD 47/89 and suggest that LSP1 is distinct from other known actin binding proteins in its effect on F-actin network structure.

Lymphocyte-Specific Protein 1 Expression in Eukaryotic Cells Reproduces the Morphologic and Motile Abnormality of NAD 47/89 Neutrophils

Blood ◽  
1998 ◽  
Vol 91 (12) ◽  
pp. 4786-4795 ◽  
Author(s):  
Thomas H. Howard ◽  
John Hartwig ◽  
Casey Cunningham
Keyword(s):  
Actin Polymerization ◽  
Cell Function ◽  
Specific Protein ◽  
Polymorphonuclear Neutrophils ◽  
Actin Binding ◽  
Actin Network ◽  
Cell Surfaces ◽  
Actin Bundles ◽  
Mixed Polarity ◽  
Low Levels

Despite its name, the actin-binding protein lymphocyte-specific protein1 (LSP1) is found in all hematopoetic cells, and yet its role in cell function remains unclear. Recently, LSP1 was identified as the 47-kD protein overexpressed in the polymorphonuclear neutrophils of patients with a rare neutrophil disorder, neutrophil actin dysfunction with abnormalities of 47-kD and 89-kD proteins (NAD 47/89). These neutrophils are immotile, defective in actin polymerization in response to agonists, and display distinctive, fine, “hairlike” F-actin-rich projections on their cell surfaces. We now show that overexpression of LSP1 produces F-actin bundles that are likely responsible for the morphologic and motile abnormalities characteristic of the NAD 47/89 phenotype. Coincident with LSP1 overexpression, cells from each of several different eukaryotic lines, including a highly motile human melanoma line, develop hairlike surface projections that branch distinctively and contain F-actin and LSP1. The hairlike projections are supported at their core by thick actin bundles, composed of actin filaments of mixed polarity, which periodically anastomose to generate a branching structure. The motility of the melanoma cells is inhibited even at low levels of LSP1 expression. Therefore, these studies show that overexpression of LSP1 alone can recreate the morphologic and motile defects seen in NAD 47/89 and suggest that LSP1 is distinct from other known actin binding proteins in its effect on F-actin network structure.

scholarly journals The 47-kD protein increased in neutrophil actin dysfunction with 47- and 89-kD protein abnormalities is lymphocyte-specific protein

Blood ◽  
1994 ◽  
Vol 83 (1) ◽  
pp. 231-241 ◽  
Author(s):  
T Howard ◽  
Y Li ◽  
M Torres ◽  
A Guerrero ◽  
T Coates
Keyword(s):  
Actin Polymerization ◽  
Binding Protein ◽  
Family Members ◽  
Autosomal Recessive Disorder ◽  
Specific Protein ◽  
Polymorphonuclear Neutrophils ◽  
Actin Binding ◽  
Expression Library ◽  
Actin Binding Protein ◽  
Formyl Peptide

A male child born of related parents suffered recurrent infections because of neutrophil actin dysfunction with increased amounts of a 47- kD protein and decreased amounts of an 89-kD protein (NAD 47/89). The patient and family members were studied to define the nature of the abnormal proteins and to examine their role in the functional defects of neutrophil actin dysfunction (NAD) 47/89 polymorphonuclear neutrophils (PMNs). NAD 47/89 PMNs are defective in motility, microfilamentous cytoskeletal structure, and formyl peptide-induced actin polymerization and express increased amounts of a 47-kD protein and decreased amounts of an 89-kD proteins intermediate abnormality in amount of 47-kD and 89-kD proteins in PMNs from parents and a female sibling suggest the disease is an autosomal recessive disorder. Immunoblots with monoclonal antibody (MoAb1) and polyclonal antibody raised to 47-kD protein showed the 89-kD protein is antigenically distinct from the 47-kD protein and the 89-kD protein is not gelsolin. 125I-actin binding to one-dimensional (1 D) and 2 D gels of PMN proteins from NAD 47/89 proband, family members, and controls showed the 47-kD protein binds actin, is acidic (pl = 4.5 to 4.7), is recognized by the MoAb1, exists on 2-D gels as three distinct actin binding species (MWapp 52 kD, 47-kD, and 44-kD), and is present in control PMNs in lesser amount than in PMNs of NAD 47/89 proband or parents. Immunoaffinity purification of the 47 kD actin binding protein on MoAb1 matrix yielded a multimolecular complex with proteins of MWapp 180 kD, 71 kD, 47 kD and actin. Cloning, sequencing, and expression of a 1.58-kb cDNA selected for MoAb1 reactivity from a HL60 expression library and microsequence of native PMNs, 47-kD actin binding protein showed the overexpressed 47-kD protein is lymphocyte-specific protein 1 (LSP1), which is a known actin binding protein. The results show LSP1 is expressed in PMNs and suggest overexpression of LSP1 is related to the motility and cytoskeletal abnormalities in NAD 47/89 PMNs.

scholarly journals The 47-kD protein increased in neutrophil actin dysfunction with 47- and 89-kD protein abnormalities is lymphocyte-specific protein

Blood ◽  
1994 ◽  
Vol 83 (1) ◽  
pp. 231-241 ◽  
Author(s):  
T Howard ◽  
Y Li ◽  
M Torres ◽  
A Guerrero ◽  
T Coates
Keyword(s):  
Actin Polymerization ◽  
Binding Protein ◽  
Family Members ◽  
Autosomal Recessive Disorder ◽  
Specific Protein ◽  
Polymorphonuclear Neutrophils ◽  
Actin Binding ◽  
Expression Library ◽  
Actin Binding Protein ◽  
Formyl Peptide

Abstract A male child born of related parents suffered recurrent infections because of neutrophil actin dysfunction with increased amounts of a 47- kD protein and decreased amounts of an 89-kD protein (NAD 47/89). The patient and family members were studied to define the nature of the abnormal proteins and to examine their role in the functional defects of neutrophil actin dysfunction (NAD) 47/89 polymorphonuclear neutrophils (PMNs). NAD 47/89 PMNs are defective in motility, microfilamentous cytoskeletal structure, and formyl peptide-induced actin polymerization and express increased amounts of a 47-kD protein and decreased amounts of an 89-kD proteins intermediate abnormality in amount of 47-kD and 89-kD proteins in PMNs from parents and a female sibling suggest the disease is an autosomal recessive disorder. Immunoblots with monoclonal antibody (MoAb1) and polyclonal antibody raised to 47-kD protein showed the 89-kD protein is antigenically distinct from the 47-kD protein and the 89-kD protein is not gelsolin. 125I-actin binding to one-dimensional (1 D) and 2 D gels of PMN proteins from NAD 47/89 proband, family members, and controls showed the 47-kD protein binds actin, is acidic (pl = 4.5 to 4.7), is recognized by the MoAb1, exists on 2-D gels as three distinct actin binding species (MWapp 52 kD, 47-kD, and 44-kD), and is present in control PMNs in lesser amount than in PMNs of NAD 47/89 proband or parents. Immunoaffinity purification of the 47 kD actin binding protein on MoAb1 matrix yielded a multimolecular complex with proteins of MWapp 180 kD, 71 kD, 47 kD and actin. Cloning, sequencing, and expression of a 1.58-kb cDNA selected for MoAb1 reactivity from a HL60 expression library and microsequence of native PMNs, 47-kD actin binding protein showed the overexpressed 47-kD protein is lymphocyte-specific protein 1 (LSP1), which is a known actin binding protein. The results show LSP1 is expressed in PMNs and suggest overexpression of LSP1 is related to the motility and cytoskeletal abnormalities in NAD 47/89 PMNs.

scholarly journals A direct proof that sole actin dynamics drive membrane deformations

2018 ◽  
Author(s):  
Camille Simon ◽  
Rémy Kusters ◽  
Valentina Caorsi ◽  
Antoine Allard ◽  
Majdouline Abou-Ghali ◽  
...  
Keyword(s):  
Mammalian Cells ◽  
Actin Polymerization ◽  
Cell Function ◽  
Turgor Pressure ◽  
Direct Proof ◽  
Actin Dynamics ◽  
Actin Binding ◽  
Associated Proteins ◽  
Low Tension ◽  
Membrane Deformations

AbstractCell membrane deformations are crucial for proper cell function. Specialized protein assemblies initiate inward or outward membrane deformations that turn into, for example, filopodia or endocytic intermediates. Actin dynamics and actin-binding proteins are involved in this process, although their detailed role remains controversial. We show here that a dynamic, branched actin network is sufficient, in absence of any membrane-associated proteins, to initiate both inward and outward membrane deformation. With actin polymerization triggered at the membrane of liposomes, we produce inward filopodia-like structures at low tension, while outward endocytosis-like structures are robustly generated regardless of tension. Our results are reminiscent of endocytosis in mammalian cells, where actin polymerization forces are required when membrane tension is increased, and in yeast, where they are always required to overcome the opposing turgor pressure. By combining experimental observations with physical modeling, we propose a mechanism for actin-driven endocytosis under high tension or high pressure conditions.

scholarly journals Cofilin-mediated actin dynamics promotes actin bundle formation during Drosophila bristle development

2016 ◽  
Vol 27 (16) ◽  
pp. 2554-2564 ◽  
Author(s):  
Jing Wu ◽  
Heng Wang ◽  
Xuan Guo ◽  
Jiong Chen
Keyword(s):  
Actin Filament ◽  
Actin Filaments ◽  
Actin Polymerization ◽  
Model System ◽  
Actin Dynamics ◽  
Actin Network ◽  
Loss Of Function ◽  
Actin Bundle ◽  
Actin Bundles

The actin bundle is an array of linear actin filaments cross-linked by actin-bundling proteins, but its assembly and dynamics are not as well understood as those of the branched actin network. Here we used the Drosophila bristle as a model system to study actin bundle formation. We found that cofilin, a major actin disassembly factor of the branched actin network, promotes the formation and positioning of actin bundles in the developing bristles. Loss of function of cofilin or AIP1, a cofactor of cofilin, each resulted in increased F-actin levels and severe defects in actin bundle organization, with the defects from cofilin deficiency being more severe. Further analyses revealed that cofilin likely regulates actin bundle formation and positioning by the following means. First, cofilin promotes a large G-actin pool both locally and globally, likely ensuring rapid actin polymerization for bundle initiation and growth. Second, cofilin limits the size of a nonbundled actin-myosin network to regulate the positioning of actin bundles. Third, cofilin prevents incorrect assembly of branched and myosin-associated actin filament into bundles. Together these results demonstrate that the interaction between the dynamic dendritic actin network and the assembling actin bundles is critical for actin bundle formation and needs to be closely regulated.

scholarly journals VASP mediated actin dynamics activate and recruit a filopodia myosin

2021 ◽  
Author(s):  
Ashley L Arthur ◽  
Amy Crawford ◽  
Anne Houdusse ◽  
Margaret A Titus
Keyword(s):  
Actin Cytoskeleton ◽  
Actin Polymerization ◽  
The State ◽  
Actin Dynamics ◽  
Actin Binding ◽  
Actin Network ◽  
Cortical Actin ◽  
Close Collaboration ◽  
Actin Binding Protein ◽  
Dynamic Regions

Filopodia are thin, actin-based structures that cells use to interact with their environments. Filopodia initiation requires a suite of conserved proteins but the mechanism remains poorly understood. The actin polymerase VASP and a MyTH-FERM (MF) myosin, DdMyo7 in amoeba, are essential for filopodia initiation. DdMyo7 is localized to dynamic regions of the actin-rich cortex. Analysis of VASP mutants and treatment of cells with anti-actin drugs shows that myosin recruitment and activation in Dictyostelium requires localized VASP-dependent actin polymerization. Targeting of DdMyo7 to the cortex alone is not sufficient for filopodia initiation; VASP activity is also required. The actin regulator locally produces a cortical actin network, that activates the MF myosin and together they shape the actin network to promote extension of parallel bundles during filopodia formation. This work reveals how filopodia initiation requires close collaboration between an actin binding protein, the state of the actin cytoskeleton and MF myosin activity.

scholarly journals VASP mediated actin dynamics activate and recruit a filopodia myosin

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Ashley L Arthur ◽  
Amy Crawford ◽  
Anne Houdusse ◽  
Margaret A Titus
Keyword(s):  
Actin Cytoskeleton ◽  
Actin Polymerization ◽  
The State ◽  
Actin Dynamics ◽  
Actin Binding ◽  
Actin Network ◽  
Cortical Actin ◽  
Close Collaboration ◽  
Actin Binding Protein ◽  
Dynamic Regions

Filopodia are thin, actin-based structures that cells use to interact with their environments. Filopodia initiation requires a suite of conserved proteins but the mechanism remains poorly understood. The actin polymerase VASP and a MyTH-FERM (MF) myosin, DdMyo7 in amoeba, are essential for filopodia initiation. DdMyo7 is localized to dynamic regions of the actin-rich cortex. Analysis of VASP mutants and treatment of cells with anti-actin drugs shows that myosin recruitment and activation in Dictyostelium requires localized VASP-dependent actin polymerization. Targeting of DdMyo7 to the cortex alone is not sufficient for filopodia initiation; VASP activity is also required. The actin regulator locally produces a cortical actin network that activates myosin and together they shape the actin network to promote extension of parallel bundles of actin during filopodia formation. This work reveals how filopodia initiation requires close collaboration between an actin binding protein, the state of the actin cytoskeleton and MF myosin activity.

scholarly journals Active cargo positioning in antiparallel transport networks

2019 ◽  
Author(s):  
Mathieu Richard ◽  
Carles Blanch-Mercader ◽  
Hajer Ennomani ◽  
Wenxiang Cao ◽  
Enrique M. De La Cruz ◽  
...  
Keyword(s):  
Transport Properties ◽  
Molecular Motors ◽  
Actin Filaments ◽  
Actin Polymerization ◽  
Myosin Ii ◽  
Actin Network ◽  
Cargo Transport ◽  
Stochastic Transport ◽  
Mixed Polarity ◽  
Micro Patterns

ABSTRACTCytoskeletal filaments assemble into dense parallel, antiparallel or disordered networks, providing a complex environment for active cargo transport and positioning by molecular motors. The interplay between the network architecture and intrinsic motor properties clearly affects transport properties but remains poorly understood. Here, by using surface micro-patterns of actin polymerization, we investigate stochastic transport properties of colloidal beads in antiparallel networks of overlapping actin filaments. We found that 200-nm beads coated with myosin-Va motors displayed directed movements towards positions where the net polarity of the actin network vanished, accumulating there. The bead distribution was dictated by the spatial profiles of local bead velocity and diffusion coefficient, indicating that a diffusion-drift process was at work. Remarkably, beads coated with heavy mero-myosin-II motors showed a similar behavior. However, although velocity gradients were steeper with myosin II, the much larger bead diffusion observed with this motor resulted in less precise positioning. Our observations are well described by a three-state model, in which active beads locally sense the net polarity of the network by frequently detaching from and reattaching to the filaments. A stochastic sequence of processive runs and diffusive searches results in a biased random walk. The precision of bead positioning is set by the gradient of net actin polarity in the network and by the run length of the cargo in an attached state. Our results unveiled physical rules for cargo transport and positioning in networks of mixed polarity.Significance statementCellular functions rely on small groups of molecular motors to transport their cargoes throughout the cell along polar filaments of the cytoskeleton. Cytoskeletal filaments self-assemble into dense networks comprising intersections and filaments of mixed polarity, challenging directed motor-based transport. Using micro-patterns of actin polymerization in-vitro, we investigated stochastic transport of colloidal beads in antiparallel networks of overlapping actin filaments. We found that beads coated with myosin motors sensed the net polarity of the actin network, resulting in active bead positioning to regions of neutral polarity with a precision depending on the motor type. A theoretical description of our experimental results provides the key physical rules for cargo transport and positioning in filament networks of mixed polarity.

High resolution spot scan imaging of frozen, hydrated actin bundles with 400kV electrons

1992 ◽  
Vol 50 (1) ◽  
pp. 512-513
Author(s):  
J. Jakana ◽  
M.F. Schmid ◽  
P. Matsudaira ◽  
W. Chiu
Keyword(s):  
High Resolution ◽  
Binding Proteins ◽  
Actin Binding ◽  
Eukaryotic Cells ◽  
Dimensional Structure ◽  
Structural Basis ◽  
Actin Bundle ◽  
Actin Bundles ◽  
3 Dimensional ◽  
Macromolecular Assembly

Actin is a protein found in all eukaryotic cells. In its polymerized form, the cells use it for motility, cytokinesis and for cytoskeletal support. An example of this latter class is the actin bundle in the acrosomal process from the Limulus sperm. The different functions actin performs seem to arise from its interaction with the actin binding proteins. A 3-dimensional structure of this macromolecular assembly is essential to provide a structural basis for understanding this interaction in relationship to its development and functions.

scholarly journals Filamin A Regulates Cardiovascular Remodeling

2021 ◽  
Vol 22 (12) ◽  
pp. 6555
Author(s):  
Sashidar Bandaru ◽  
Chandu Ala ◽  
Alex-Xianghua Zhou ◽  
Levent M. Akyürek
Keyword(s):  
Transcription Factors ◽  
Respiratory Diseases ◽  
Cell Function ◽  
Actin Binding ◽  
Cardiovascular Development ◽  
Filamin A ◽  
Cardiovascular Malformations ◽  
Experimental Animal Models ◽  
Actin Networks ◽  
Cardiovascular Remodeling

Filamin A (FLNA) is a large actin-binding cytoskeletal protein that is important for cell motility by stabilizing actin networks and integrating them with cell membranes. Interestingly, a C-terminal fragment of FLNA can be cleaved off by calpain to stimulate adaptive angiogenesis by transporting multiple transcription factors into the nucleus. Recently, increasing evidence suggests that FLNA participates in the pathogenesis of cardiovascular and respiratory diseases, in which the interaction of FLNA with transcription factors and/or cell signaling molecules dictate the function of vascular cells. Localized FLNA mutations associate with cardiovascular malformations in humans. A lack of FLNA in experimental animal models disrupts cell migration during embryogenesis and causes anomalies, including heart and vessels, similar to human malformations. More recently, it was shown that FLNA mediates the progression of myocardial infarction and atherosclerosis. Thus, these latest findings identify FLNA as an important novel mediator of cardiovascular development and remodeling, and thus a potential target for therapy. In this update, we summarized the literature on filamin biology with regard to cardiovascular cell function.

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