scholarly journals Cerebral Ischemia-Reperfusion Is Associated With Upregulation of Cofilin-1 in the Motor Cortex

2021 ◽  
Vol 9 ◽  
Author(s):  
Ming-Shu Xu ◽  
Lei-Miao Yin ◽  
Ai-Fang Cheng ◽  
Ying-Jie Zhang ◽  
Di Zhang ◽  
...  
Keyword(s):  
Cerebral Ischemia ◽  
Motor Cortex ◽  
Binding Proteins ◽  
Binding Protein ◽  
Ischemia Reperfusion ◽  
High Expression ◽  
Actin Binding ◽  
Actin Binding Proteins ◽  
Actin Binding Protein ◽  
Cerebral Ischemia Reperfusion

Cerebral ischemia is one of the leading causes of death. Reperfusion is a critical stage after thrombolysis or thrombectomy, accompanied by oxidative stress, excitotoxicity, neuroinflammation, and defects in synapse structure. The process is closely related to the dephosphorylation of actin-binding proteins (e.g., cofilin-1) by specific phosphatases. Although studies of the molecular mechanisms of the actin cytoskeleton have been ongoing for decades, limited studies have directly investigated reperfusion-induced reorganization of actin-binding protein, and little is known about the gene expression of actin-binding proteins. The exact mechanism is still uncertain. The motor cortex is very important to save nerve function; therefore, we chose the penumbra to study the relationship between cerebral ischemia-reperfusion and actin-binding protein. After transient middle cerebral artery occlusion (MCAO) and reperfusion, we confirmed reperfusion and motor function deficit by cerebral blood flow and gait analysis. PCR was used to screen the high expression mRNAs in penumbra of the motor cortex. The high expression of cofilin in this region was confirmed by immunohistochemistry (IHC) and Western blot (WB). The change in cofilin-1 expression appears at the same time as gait imbalance, especially maximum variation and left front swing. It is suggested that cofilin-1 may partially affect motor cortex function. This result provides a potential mechanism for understanding cerebral ischemia-reperfusion.

scholarly journals Actin-binding Proteins Coronin-1a and IBA-1 are Effective Microglial Markers for Immunohistochemistry

2007 ◽  
Vol 55 (7) ◽  
pp. 687-700 ◽  
Author(s):  
Zeshan Ahmed ◽  
Gerry Shaw ◽  
Ved P. Sharma ◽  
Cui Yang ◽  
Eileen McGowan ◽  
...  
Keyword(s):  
Binding Proteins ◽  
Dynamic Properties ◽  
Binding Protein ◽  
Actin Binding ◽  
Actin Binding Proteins ◽  
Actin Binding Protein ◽  
Membrane Cytoskeleton ◽  
Mouse Tissues ◽  
Cell Type Specific ◽  
Double Immunolabeling

This study identifies the actin-binding protein, coronin-1a, as a novel and effective immunohistochemical marker for microglia in both cell cultures and in formaldehyde-fixed, paraffin-embedded tissue. Antibodies to coronin-1a effectively immunostained microglia in human, monkey, horse, rat, and mouse tissues, even in tissues stored for long periods of time. The identity of coronin-1a-immunoreactive cells as microglia was confirmed using double immunolabeling with cell type-specific markers as well as by morphological features and the distribution of immunoreactive cells. These properties are shared by another actin-binding protein, IBA-1. Unlike IBA-1, coronin-1a immunoreactivity was also detected in lymphocytes and certain other hematopoietic cells. The results indicate that both coronin-1a and IBA-1 are robust markers for microglia that can be used in routinely processed tissue of humans and animals. Because both coronin-1a and IBA-1 are actin-binding proteins that play a role in rearrangement of the membrane cytoskeleton, it suggests that these proteins are critical to dynamic properties of microglia.

scholarly journals Secretory cell actin-binding proteins: identification of a gelsolin-like protein in chromaffin cells.

1986 ◽  
Vol 102 (2) ◽  
pp. 636-646 ◽  
Author(s):  
M F Bader ◽  
J M Trifaró ◽  
O K Langley ◽  
D Thiersé ◽  
D Aunis
Keyword(s):  
Adrenal Medulla ◽  
Chromaffin Cells ◽  
Binding Proteins ◽  
Binding Protein ◽  
Actin Binding ◽  
Secretory Process ◽  
Secretory Cells ◽  
Breakdown Product ◽  
Actin Binding Proteins ◽  
Actin Binding Protein

Chromaffin cells, secretory cells of the adrenal medulla, have been shown to contain actin and other contractile proteins, which might be involved in the secretory process. Actin and Ca++-sensitive actin-binding proteins were purified from bovine adrenal medulla on affinity columns using DNase-I as a ligand. Buffers that contained decreasing Ca++ concentrations were used to elute three major proteins of 93, 91, and 85 kD. The bulk of the actin was eluted with guanidine-HCl buffer plus some 93- and 91-kD proteins. These Ca++-sensitive regulatory proteins were shown to inhibit the gelation of actin using the low-shear falling ball viscometer and by electron microscopy. Actin filaments were found to be shortened by fragmentation. Using antibody raised against rabbit lung macrophage gelsolin, proteolytic digestion with Staphylococcus V8 protease and two-dimensional gel electrophoresis, the 91-kD actin-binding protein was shown to be a gelsolin-like protein. The 93-kD actin-binding protein also showed cross-reactivity with anti-gelsolin antibody, similar peptide maps, and a basic-shift in pHi indicating that this 93-kD protein is a brevin-like protein, derived from blood present abundantly in adrenal medulla. Purification from isolated chromaffin cells demonstrated the presence of 91- and 85-kD proteins, whereas the 93-kD protein was hardly detectable. The 85-kD protein is not a breakdown product of brevin-like or gelsolin-like proteins. It did not cross-react with anti-gelsolin antibody and showed a very different peptide map after mild digestion with V8 protease. Antibodies were raised against the 93- and 91-kD actin-binding proteins and the 85-kD actin-binding protein. Antibody against the 85-kD protein did not cross-react with 93- and 91-kD proteins and vice versa. In vivo, the cytoskeleton organization of chromaffin secretory cells is not known, but appears to be under the control of the intracellular concentration of free calcium. The ability of calcium to activate the gelsolin-like protein, and as shown elsewhere to alter fodrin localization, provides a mechanism for gel-sol transition that might be essential for granule movement and membrane-membrane interactions involved in the secretory process.

scholarly journals Coordinated inhibition of actin-induced platelet aggregation by plasma gelsolin and vitamin D-binding protein

Blood ◽  
1993 ◽  
Vol 82 (12) ◽  
pp. 3648-3657 ◽  
Author(s):  
CA Vasconcellos ◽  
SE Lind
Keyword(s):  
Vitamin D ◽  
Platelet Aggregation ◽  
Actin Filaments ◽  
Binding Proteins ◽  
Binding Protein ◽  
Adenine Nucleotides ◽  
Actin Binding ◽  
Vitamin D Binding Protein ◽  
Actin Binding Proteins ◽  
Induced Aggregation

Actin is an abundant intracellular protein that is released into the blood during tissue injury and its injection into rats causes microthrombi to form in the vasculature. This report and others have shown that actin filaments are able to aggregate platelets in an adenosine diphosphate (ADP)-dependent manner. The effects on this process of two plasma actin-binding proteins, vitamin D-binding protein (DBP) and gelsolin, were examined separately and together. The addition of DBP, a monomer-binding protein, to actin filaments did not affect their ability to induce platelet aggregation. However, severing of actin filaments with gelsolin resulted in an increased degree of platelet aggregation. Preincubation of F-actin with both gelsolin and DBP resulted in a significant inhibition of aggregation. The effects of DBP and gelsolin on actin-induced aggregation paralleled their effects on exchange of actin-bound adenine nucleotides. DBP inhibited 1, N6- ethenoadenosine 5′ triphosphate (epsilon-ATP) exchange with G-actin but not with F-actin. Gelsolin increased epsilon-ATP exchange with F-actin, which was largely abrogated by the addition of DBP. These results suggest that gelsolin's severing (and subsequent capping) of actin filaments not only results in an increase in the number of pointed filament ends but also in the dissociation of actin monomers containing ADP. Phalloidin, which stabilizes actin filaments while decreasing both monomer and nucleotide exchange, inhibited actin-induced aggregation, as well, indicating that depolymerization of actin filaments is not required to inhibit aggregation. Platelet activation by either G- or F- actin may thus be regulated by the local concentrations of the plasma actin-binding proteins gelsolin and DBP. Together, these proteins inhibit platelet aggregation in a manner that can be explained by their effects on actin's filament structure and the accessibility of its bound ADP. Depletion of DBP or gelsolin may allow actin released from injured tissues to stimulate purinergic receptors on platelets, and perhaps other cells, via its bound adenine nucleotides.

scholarly journals Yeast actin-binding proteins: evidence for a role in morphogenesis.

1988 ◽  
Vol 107 (6) ◽  
pp. 2551-2561 ◽  
Author(s):  
D G Drubin ◽  
K G Miller ◽  
D Botstein
Keyword(s):  
Yeast Cell ◽  
Myosin Heavy Chain ◽  
Heavy Chain ◽  
Actin Filaments ◽  
Binding Proteins ◽  
Binding Protein ◽  
Actin Binding ◽  
Cortical Actin ◽  
Actin Binding Protein

Three yeast actin-binding proteins were identified using yeast actin filaments as an affinity matrix. One protein appears to be a yeast myosin heavy chain; it is dissociated from actin filaments by ATP, it is similar in size (200 kD) to other myosins, and antibodies directed against Dictyostelium myosin heavy chain bind to it. Immunofluorescence experiments show that a second actin-binding protein (67 kD) colocalizes in vivo with both cytoplasmic actin cables and cortical actin patches, the only identifiable actin structures in yeast. The cortical actin patches are concentrated at growing surfaces of the yeast cell where they might play a role in membrane and cell wall insertion, and the third actin-binding protein (85 kD) is only detected in association with these structures. This 85-kD protein is therefore a candidate for a determinant of growth sites. The in vivo role of this protein was tested by overproduction; this overproduction causes a reorganization of the actin cytoskeleton which in turn dramatically affects the budding pattern and spatial growth organization of the yeast cell.

scholarly journals A high-throughput fluorescence lifetime-based assay to detect binding of myosin-binding protein C to F-actin

2021 ◽  
Vol 153 (3) ◽  
Author(s):  
Thomas A. Bunch ◽  
Victoria C. Lepak ◽  
Kellan M. Bortz ◽  
Brett A. Colson
Keyword(s):  
Fluorescence Lifetime ◽  
Protein C ◽  
Binding Proteins ◽  
Binding Capacity ◽  
Binding Protein ◽  
Actin Binding ◽  
Actin Binding Proteins ◽  
Myosin Binding Protein ◽  
Myosin Binding Protein C ◽  
Myosin Binding

Binding properties of actin-binding proteins are typically evaluated by cosedimentation assays. However, this method is time-consuming, involves multiple steps, and has a limited throughput. These shortcomings preclude its use in screening for drugs that modulate actin-binding proteins relevant to human disease. To develop a simple, quantitative, and scalable F-actin–binding assay, we attached fluorescent probes to actin's Cys-374 and assessed changes in fluorescence lifetime upon binding to the N-terminal region (domains C0–C2) of human cardiac myosin-binding protein C (cMyBP-C). The lifetime of all five probes tested decreased upon incubation with cMyBP-C C0–C2, as measured by time-resolved fluorescence (TR-F), with IAEDANS being the most sensitive probe that yielded the smallest errors. The TR-F assay was compared with cosedimentation to evaluate in vitro changes in binding to actin and actin–tropomyosin arising from cMyBP-C mutations associated with hypertrophic cardiomyopathy (HCM) and tropomyosin binding. Lifetime changes of labeled actin with added C0–C2 were consistent with cosedimentation results. The HCM mutation L352P was confirmed to enhance actin binding, whereas PKA phosphorylation reduced binding. The HCM mutation R282W, predicted to disrupt a PKA recognition sequence, led to deficits in C0–C2 phosphorylation and altered binding. Lastly, C0–C2 binding was found to be enhanced by tropomyosin and binding capacity to be altered by mutations in a tropomyosin-binding region. These findings suggest that the TR-F assay is suitable for rapidly and accurately determining quantitative binding and for screening physiological conditions and compounds that affect cMyBP-C binding to F-actin for therapeutic discovery.

Presence of Clathrin at Adhesion Sites in Phagocytosing Macrophages

1982 ◽  
Vol 40 ◽  
pp. 114-117
Author(s):  
J. Aggeler ◽  
J.E. Heuser ◽  
Z. Werb
Keyword(s):  
Plasma Membrane ◽  
Actin Filaments ◽  
Binding Proteins ◽  
Binding Protein ◽  
Cell Spreading ◽  
Actin Binding ◽  
Dense Network ◽  
Actin Binding Protein ◽  
Adhesion Sites ◽  
Cytoplasmic Surface

Phagocytosis of particles by macrophages may be similar to cell spreading on a substratum, in that a dense network of actin filaments appears beneath the plasma membrane in both cases. When viewed in broken-open or detergent- extracted cells, cytoskeletal filaments are observed to form focal attachments to the plasma membrane and to the cytoplasmic surface of phagosomes. Hartwig et al. have presented a model of phagocytosis in which an actin-binding protein alters the organization of subplasmalemma1 actin filaments in such a way that the plasma membrane is forced up over the particle to form the phagosome. Their evidence indicates that similar actin-binding proteins may function during cell spreading.

scholarly journals Coordinated inhibition of actin-induced platelet aggregation by plasma gelsolin and vitamin D-binding protein

Blood ◽  
1993 ◽  
Vol 82 (12) ◽  
pp. 3648-3657 ◽  
Author(s):  
CA Vasconcellos ◽  
SE Lind
Keyword(s):  
Vitamin D ◽  
Platelet Aggregation ◽  
Actin Filaments ◽  
Binding Proteins ◽  
Binding Protein ◽  
Adenine Nucleotides ◽  
Actin Binding ◽  
Vitamin D Binding Protein ◽  
Actin Binding Proteins ◽  
Induced Aggregation

Abstract Actin is an abundant intracellular protein that is released into the blood during tissue injury and its injection into rats causes microthrombi to form in the vasculature. This report and others have shown that actin filaments are able to aggregate platelets in an adenosine diphosphate (ADP)-dependent manner. The effects on this process of two plasma actin-binding proteins, vitamin D-binding protein (DBP) and gelsolin, were examined separately and together. The addition of DBP, a monomer-binding protein, to actin filaments did not affect their ability to induce platelet aggregation. However, severing of actin filaments with gelsolin resulted in an increased degree of platelet aggregation. Preincubation of F-actin with both gelsolin and DBP resulted in a significant inhibition of aggregation. The effects of DBP and gelsolin on actin-induced aggregation paralleled their effects on exchange of actin-bound adenine nucleotides. DBP inhibited 1, N6- ethenoadenosine 5′ triphosphate (epsilon-ATP) exchange with G-actin but not with F-actin. Gelsolin increased epsilon-ATP exchange with F-actin, which was largely abrogated by the addition of DBP. These results suggest that gelsolin's severing (and subsequent capping) of actin filaments not only results in an increase in the number of pointed filament ends but also in the dissociation of actin monomers containing ADP. Phalloidin, which stabilizes actin filaments while decreasing both monomer and nucleotide exchange, inhibited actin-induced aggregation, as well, indicating that depolymerization of actin filaments is not required to inhibit aggregation. Platelet activation by either G- or F- actin may thus be regulated by the local concentrations of the plasma actin-binding proteins gelsolin and DBP. Together, these proteins inhibit platelet aggregation in a manner that can be explained by their effects on actin's filament structure and the accessibility of its bound ADP. Depletion of DBP or gelsolin may allow actin released from injured tissues to stimulate purinergic receptors on platelets, and perhaps other cells, via its bound adenine nucleotides.

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