scholarly journals Essential and unique roles of PIP5K-γ and -α in Fcγ receptor-mediated phagocytosis

2009 ◽  
Vol 184 (2) ◽  
pp. 281-296 ◽  
Author(s):  
Yuntao S. Mao ◽  
Masaki Yamaga ◽  
Xiaohui Zhu ◽  
Yongjie Wei ◽  
Hui-Qiao Sun ◽  
...  
Keyword(s):  
Actin Cytoskeleton ◽  
Actin Polymerization ◽  
Type I ◽  
Dependent Manner ◽  
Fcγ Receptor ◽  
Protein Activation ◽  
Phosphatidylinositol 4 ◽  
Syndrome Protein ◽  
Pharmacological Manipulations

The actin cytoskeleton is dynamically remodeled during Fcγ receptor (FcγR)-mediated phagocytosis in a phosphatidylinositol (4,5)-bisphosphate (PIP2)-dependent manner. We investigated the role of type I phosphatidylinositol 4-phosphate 5-kinase (PIP5K) γ and α isoforms, which synthesize PIP2, during phagocytosis. PIP5K-γ−/− bone marrow–derived macrophages (BMM) have a highly polymerized actin cytoskeleton and are defective in attachment to IgG-opsonized particles and FcγR clustering. Delivery of exogenous PIP2 rescued these defects. PIP5K-γ knockout BMM also have more RhoA and less Rac1 activation, and pharmacological manipulations establish that they contribute to the abnormal phenotype. Likewise, depletion of PIP5K-γ by RNA interference inhibits particle attachment. In contrast, PIP5K-α knockout or silencing has no effect on attachment but inhibits ingestion by decreasing Wiskott-Aldrich syndrome protein activation, and hence actin polymerization, in the nascent phagocytic cup. In addition, PIP5K-γ but not PIP5K-α is transiently activated by spleen tyrosine kinase–mediated phosphorylation. We propose that PIP5K-γ acts upstream of Rac/Rho and that the differential regulation of PIP5K-γ and -α allows them to work in tandem to modulate the actin cytoskeleton during the attachment and ingestion phases of phagocytosis.

scholarly journals ADictyosteliumHomologue of WASP Is Required for Polarized F-Actin Assembly during Chemotaxis

2005 ◽  
Vol 16 (5) ◽  
pp. 2191-2206 ◽  
Author(s):  
Scott A. Myers ◽  
Ji W. Han ◽  
Yoonsung Lee ◽  
Richard A. Firtel ◽  
Chang Y. Chung
Keyword(s):  
Actin Cytoskeleton ◽  
Actin Polymerization ◽  
Essential Role ◽  
Leading Edge ◽  
Actin Assembly ◽  
Temporal Control ◽  
Wiskott Aldrich Syndrome ◽  
Low Levels ◽  
Syndrome Protein

The actin cytoskeleton controls the overall structure of cells and is highly polarized in chemotaxing cells, with F-actin assembled predominantly in the anterior leading edge and to a lesser degree in the cell's posterior. Wiskott-Aldrich syndrome protein (WASP) has emerged as a central player in controlling actin polymerization. We have investigated WASP function and its regulation in chemotaxing Dictyostelium cells and demonstrated the specific and essential role of WASP in organizing polarized F-actin assembly in chemotaxing cells. Cells expressing very low levels of WASP show reduced F-actin levels and significant defects in polarized F-actin assembly, resulting in an inability to establish axial polarity during chemotaxis. GFP-WASP preferentially localizes at the leading edge and uropod of chemotaxing cells and the B domain of WASP is required for the localization of WASP. We demonstrated that the B domain binds to PI(4,5)P2and PI(3,4,5)P3with similar affinities. The interaction between the B domain and PI(3,4,5)P3plays an important role for the localization of WASP to the leading edge in chemotaxing cells. Our results suggest that the spatial and temporal control of WASP localization and activation is essential for the regulation of directional motility.

scholarly journals PACAP-mediated sperm–cumulus cell interaction promotes fertilization

Reproduction ◽  
2011 ◽  
Vol 141 (2) ◽  
pp. 163-171 ◽  
Author(s):  
Ichiro Tanii ◽  
Tadashi Aradate ◽  
Kouhei Matsuda ◽  
Akira Komiya ◽  
Hideki Fuse
Keyword(s):  
Sperm Motility ◽  
Acrosome Reaction ◽  
Cumulus Cell ◽  
Cumulus Cells ◽  
Cell Interaction ◽  
Fertilization Rate ◽  
Type I ◽  
Dependent Manner ◽  
Sperm Penetration

The developing acrosome in spermatids contains pituitary adenylate cyclase-activating polypeptide (PACAP). However, the role of the acrosomal PACAP remains unclear because it has not been detected in mature spermatids and sperm. We reinvestigated whether the sperm acrosome contains PACAP. An antiserum produced against PACAP reacted to the anterior acrosome in epididymal sperm fixed under mild conditions, suggesting that PACAP acts on oocytes and/or cumulus cells at the site of fertilization. Immunolabeling and RT-PCR demonstrated the presence of PACAP type I receptor, a PACAP-specific receptor, in postovulatory cumulus cells. To investigate the role of PACAP in fertilization, we pretreated cumulus–oocyte complexes with the polypeptide. At a low concentration of sperm, the fertilization rate was significantly enhanced by PACAP in a dose-dependent manner. Sperm penetration through the oocyte investment, cumulus layer, and zona pellucida was also enhanced by PACAP. The enhancement was probably due to an enhancement in sperm motility and the zona-induced acrosome reaction, which were stimulated by a cumulus cell-releasing factor. Indeed, PACAP treatment increased the secretion of progesterone from the cumulus–oocyte complexes. These results strongly suggest that in response to PACAP, cumulus cells release a soluble factor that probably stimulates sperm motility and the acrosome reaction, thereby promoting fertilization.

scholarly journals Role of Hypoxia-Mediated Autophagy in Tumor Cell Death and Survival

Cancers ◽  
2021 ◽  
Vol 13 (3) ◽  
pp. 533
Author(s):  
Rania F. Zaarour ◽  
Bilal Azakir ◽  
Edries Y. Hajam ◽  
Husam Nawafleh ◽  
Nagwa A. Zeinelabdin ◽  
...  
Keyword(s):  
Cell Death ◽  
Type I ◽  
Dependent Manner ◽  
Destruction Process ◽  
Tumor Cell Death ◽  
Complex Regulation ◽  
Cancer Immunotherapies ◽  
The Relationship ◽  
Shed Light

Programmed cell death or type I apoptosis has been extensively studied and its contribution to the pathogenesis of disease is well established. However, autophagy functions together with apoptosis to determine the overall fate of the cell. The cross talk between this active self-destruction process and apoptosis is quite complex and contradictory as well, but it is unquestionably decisive for cell survival or cell death. Autophagy can promote tumor suppression but also tumor growth by inducing cancer-cell development and proliferation. In this review, we will discuss how autophagy reprograms tumor cells in the context of tumor hypoxic stress. We will illustrate how autophagy acts as both a suppressor and a driver of tumorigenesis through tuning survival in a context dependent manner. We also shed light on the relationship between autophagy and immune response in this complex regulation. A better understanding of the autophagy mechanisms and pathways will undoubtedly ameliorate the design of therapeutics aimed at targeting autophagy for future cancer immunotherapies.

Fibrinogen binding to the integrin αIIbβ3 modulates store-mediated calcium entry in human platelets

Blood ◽  
2001 ◽  
Vol 97 (9) ◽  
pp. 2648-2656 ◽  
Author(s):  
Juan A. Rosado ◽  
Else M. Y. Meijer ◽  
Karly Hamulyak ◽  
Irena Novakova ◽  
Johan W. M. Heemskerk ◽  
...  
Keyword(s):  
Actin Cytoskeleton ◽  
Actin Polymerization ◽  
Adenosine Diphosphate ◽  
Inhibitory Effect ◽  
Human Platelets ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Integrin Αiibβ3 ◽  
Fibrinogen Binding

Abstract Effects of the occupation of integrin αIIbβ3 by fibrinogen on Ca++signaling in fura-2–loaded human platelets were investigated. Adding fibrinogen to washed platelet suspensions inhibited increases in cytosolic [Ca++] concentrations ([Ca++]i) evoked by adenosine diphosphate (ADP) and thrombin in a concentration-dependent manner in the presence of external Ca++ but not in the absence of external Ca++ or in the presence of the nonselective cation channel blocker SKF96365, indicating selective inhibition of Ca++entry. Fibrinogen also inhibited store-mediated Ca++ entry (SMCE) activated after Ca++ store depletion using thapsigargin. The inhibitory effect of fibrinogen was reversed if fibrinogen binding to αIIbβ3 was blocked using RDGS or abciximab and was absent in platelets from patients homozygous for Glanzmann thrombasthenia. Fibrinogen was without effect on SMCE once activated. Activation of SMCE in platelets occurs through conformational coupling between the intracellular stores and the plasma membrane and requires remodeling of the actin cytoskeleton. Fibrinogen inhibited actin polymerization evoked by ADP or thapsigargin in control cells and in cells loaded with the Ca++ chelator dimethyl BAPTA. It also inhibited the translocation of the tyrosine kinase p60src to the cytoskeleton. These results indicate that the binding of fibrinogen to integrin αIIbβ3 inhibits the activation of SMCE in platelets by a mechanism that may involve modulation of the reorganization of the actin cytoskeleton and the cytoskeletal association of p60src. This action may be important in intrinsic negative feedback to prevent the further activation of platelets subjected to low-level stimuli in vivo.

Role of common cytokine receptor γ chain (γc)– and Jak3-dependent signaling in the proliferation and survival of murine mast cells

Blood ◽  
2000 ◽  
Vol 96 (6) ◽  
pp. 2172-2180 ◽  
Author(s):  
Kotaro Suzuki ◽  
Hiroshi Nakajima ◽  
Norihiko Watanabe ◽  
Shin-ichiro Kagami ◽  
Akira Suto ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Mast Cells ◽  
Cytokine Receptor ◽  
Type I ◽  
Dependent Manner ◽  
Type Ii ◽  
Wild Type ◽  
Peritoneal Mast Cells ◽  
The Common

Abstract The regulatory roles of the common cytokine receptor γ chain (γc)– and Jak3-dependent signaling in the proliferation and survival of mast cells were determined using γc-deficient (γc−) and Jak3-deficient (Jak3−) mice. Although the mast cells in γc− and Jak3− mice were morphologically indistinguishable from those in wild-type mice, the number of peritoneal mast cells was decreased in γc− and Jak3− mice as compared with that in wild-type mice. Among γc-related cytokines, interleukin (IL)-4 and IL-9, but not IL-2, IL-7, or IL-15, enhanced the proliferation and survival of bone marrow–derived mast cells (BMMCs) from wild-type mice. However, the effects of IL-4 and IL-9 were absent in BMMCs from γc− and Jak3−mice. In addition, IL-4Rα, γc, and Jak3, but not IL-2Rβ or IL-7Rα, were expressed in BMMCs. In contrast, IL-13 did not significantly induce the proliferation and survival of BMMCs even from wild-type mice, and IL-13Rα1 was not expressed in BMMCs. Furthermore, IL-4 phosphorylated the 65-kd isoform of Stat6 in BMMCs from wild-type mice but not from γc− and Jak3− mice. These results indicate that γc- and Jak3-dependent signaling is essential for IL-4– and IL-9–induced proliferation and survival of murine mast cells, that the effects of IL-4 are mediated by type I IL-4R and that type II IL-4R is absent on mast cells, and that IL-4 phosphorylates the 65-kd isoform of Stat6 in mast cells in a γc- and Jak3-dependent manner.

scholarly journals Type I phosphatidylinositol 4-phosphate 5-kinase controls neutrophil polarity and directional movement

2007 ◽  
Vol 179 (7) ◽  
pp. 1539-1553 ◽  
Author(s):  
Rosa Ana Lacalle ◽  
Rosa M. Peregil ◽  
Juan Pablo Albar ◽  
Ernesto Merino ◽  
Carlos Martínez-A ◽  
...  
Keyword(s):  
Actin Polymerization ◽  
Cell Movement ◽  
Leading Edge ◽  
Cell Polarization ◽  
Type I ◽  
Lipid Kinase ◽  
Erm Proteins ◽  
Chemical Gradients ◽  
C Terminus ◽  
Phosphatidylinositol 4

Directional cell movement in response to external chemical gradients requires establishment of front–rear asymmetry, which distinguishes an up-gradient protrusive leading edge, where Rac-induced F-actin polymerization takes place, and a down-gradient retractile tail (uropod in leukocytes), where RhoA-mediated actomyosin contraction occurs. The signals that govern this spatial and functional asymmetry are not entirely understood. We show that the human type I phosphatidylinositol 4-phosphate 5-kinase isoform β (PIPKIβ) has a role in organizing signaling at the cell rear. We found that PIPKIβ polarized at the uropod of neutrophil-differentiated HL60 cells. PIPKIβ localization was independent of its lipid kinase activity, but required the 83 C-terminal amino acids, which are not homologous to other PIPKI isoforms. The PIPKIβ C terminus interacted with EBP50 (4.1-ezrin-radixin-moesin (ERM)-binding phosphoprotein 50), which enabled further interactions with ERM proteins and the Rho-GDP dissociation inhibitor (RhoGDI). Knockdown of PIPKIβ with siRNA inhibited cell polarization and impaired cell directionality during dHL60 chemotaxis, suggesting a role for PIPKIβ in these processes.

scholarly journals Binding of Brucella protein, Bp26, to select extracellular matrix molecules

2019 ◽  
Vol 20 (1) ◽  
Author(s):  
Yasmin ElTahir ◽  
Amna Al-Araimi ◽  
Remya R. Nair ◽  
Kaija J. Autio ◽  
Hongmin Tu ◽  
...  
Keyword(s):  
Extracellular Matrix ◽  
Type I Collagen ◽  
Host Cells ◽  
Mouse Serum ◽  
Enzyme Linked Immunosorbent Assay ◽  
Target Cells ◽  
Type I ◽  
Dependent Manner ◽  
Biolayer Interferometry

Abstract Background Brucella is a facultative intracellular pathogen responsible for zoonotic disease brucellosis. Little is known about the molecular basis of Brucella adherence to host cells. In the present study, the possible role of Bp26 protein as an adhesin was explored. The ability of Brucella protein Bp26 to bind to extracellular matrix (ECM) proteins was determined by enzyme-linked immunosorbent assay (ELISA) and biolayer interferometry (BLI). Results ELISA experiments showed that Bp26 bound in a dose-dependent manner to both immobilized type I collagen and vitronectin. Bp26 bound weakly to soluble fibronectin but did not bind to immobilized fibronectin. No binding to laminin was detected. Biolayer interferometry showed high binding affinity of Bp26 to immobilized type I collagen and no binding to fibronectin or laminin. Mapping of Bp26 antigenic epitopes by biotinylated overlapping peptides spanning the entire sequence of Bp26 using anti Bp26 mouse serum led to the identification of five linear epitopes. Collagen and vitronectin bound to peptides from several regions of Bp26, with many of the binding sites for the ligands overlapping. The strongest binding for anti-Bp26 mouse serum, collagen and vitronectin was to the peptides at the C-terminus of Bp26. Fibronectin did not bind to any of the peptides, although it bound to the whole Bp26 protein. Conclusions Our results highlight the possible role of Bp26 protein in the adhesion process of Brucella to host cells through ECM components. This study revealed that Bp26 binds to both immobilized and soluble type I collagen and vitronectin. It also binds to soluble but not immobilized fibronectin. However, Bp26 does not bind to laminin. These are novel findings that offer insight into understanding the interplay between Brucella and host target cells, which may aid in future identification of a new target for diagnosis and/or vaccine development and prevention of brucellosis.

scholarly journals Farnesylcysteine analogues inhibit store-regulated Ca2+ entry in human platelets: evidence for involvement of small GTP-binding proteins and actin cytoskeleton

2000 ◽  
Vol 347 (1) ◽  
pp. 183-192 ◽  
Author(s):  
Juan A. ROSADO ◽  
Stewart O. SAGE
Keyword(s):  
Actin Cytoskeleton ◽  
Actin Polymerization ◽  
Binding Proteins ◽  
Selective Inhibition ◽  
Human Platelets ◽  
Cytochalasin D ◽  
Dependent Manner ◽  
Gtp Binding Proteins ◽  
Gtp Binding ◽  
Prenylated Proteins

We have investigated the mechanism of Ca2+ entry into fura-2-loaded human platelets by preventing the prenylation of proteins such as small GTP-binding proteins. The farnesylcysteine analogues farnesylthioacetic acid (FTA) and N-acetyl-S-geranylgeranyl-L-cysteine (AGGC), which are inhibitors of the methylation of prenylated and geranylgeranylated proteins respectively, significantly decreased thrombin-evoked increases in intracellular free Ca2+ concentration ([Ca2+]i) in the presence, but not in the absence, of external Ca2+, suggesting a relatively selective inhibition of Ca2+ entry over internal release. Both these compounds and N-acetyl-S-farnesyl-L-cysteine, which had similar effects to those of FTA, also decreased Ca2+ entry evoked by the depletion of intracellular Ca2+ stores with thapsigargin. The inactive control N-acetyl-S-geranyl-L-cysteine was without effect. Patulin, an inhibitor of prenylation that is inert with respect to methyltransferases, also decreased store-regulated Ca2+ entry. Cytochalasin D, an inhibitor of actin polymerization, significantly decreased store-regulated Ca2+ entry in a time-dependent manner. Both cytochalasin D and the farnesylcysteine analogues FTA and AGGC inhibited actin polymerization; however, when evoking the same extent of decrease in actin filament formation, FTA and AGGC showed greater inhibitory effects on Ca2+ entry, indicating a cytoskeleton-independent component in the regulation of Ca2+ entry by small GTP-binding-protein. These findings suggest that prenylated proteins such as small GTP-binding proteins are involved in store-regulated Ca2+ entry through actin cytoskeleton-dependent and cytoskeleton-independent mechanisms in human platelets.

scholarly journals Actin-dependent Lamellipodia Formation and Microtubule-dependent Tail Retraction Control-directed Cell Migration

2000 ◽  
Vol 11 (9) ◽  
pp. 2999-3012 ◽  
Author(s):  
Christoph Ballestrem ◽  
Bernhard Wehrle-Haller ◽  
Boris Hinz ◽  
Beat A. Imhof
Keyword(s):  
Cell Migration ◽  
Cell Body ◽  
Actin Polymerization ◽  
Green Fluorescence Protein ◽  
Time Lapse ◽  
Dependent Manner ◽  
Directed Cell Migration ◽  
Cell Front ◽  
Fluorescence Protein

Migrating cells are polarized with a protrusive lamella at the cell front followed by the main cell body and a retractable tail at the rear of the cell. The lamella terminates in ruffling lamellipodia that face the direction of migration. Although the role of actin in the formation of lamellipodia is well established, it remains unclear to what degree microtubules contribute to this process. Herein, we have studied the contribution of microtubules to cell motility by time-lapse video microscopy on green flourescence protein-actin- and tubulin-green fluorescence protein–transfected melanoma cells. Treatment of cells with either the microtubule-disrupting agent nocodazole or with the stabilizing agent taxol showed decreased ruffling and lamellipodium formation. However, this was not due to an intrinsic inability to form ruffles and lamellipodia because both were restored by stimulation of cells with phorbol 12-myristate 13-acetate in a Rac-dependent manner, and by stem cell factor in melanoblasts expressing the receptor tyrosine kinase c-kit. Although ruffling and lamellipodia were formed without microtubules, the microtubular network was needed for advancement of the cell body and the subsequent retraction of the tail. In conclusion, we demonstrate that the formation of lamellipodia can occur via actin polymerization independently of microtubules, but that microtubules are required for cell migration, tail retraction, and modulation of cell adhesion.

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