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Membrane Skeleton (membrane + skeleton)
Selected AbstractsTwo different unique cardiac isoforms of protein 4.1R in zebrafish, Danio rerio, and insights into their cardiac functions as related to their unique structuresDEVELOPMENT GROWTH & DIFFERENTIATION, Issue 7 2010Kenji Murata Protein 4.1R (4.1R) has been identified as the major component of the human erythrocyte membrane skeleton. The members of the protein 4.1 gene family are expressed in a tissue-specific alternative splicing manner that increases their functions in each tissue; however, the exact roles of cardiac 4.1R in the developing myocardium are poorly understood. In zebrafish (ZF), we identified two heart-specific 4.1R isoforms, ZF4.1RH2 and ZF4.1RH3, encoding N-terminal 30 kDa (FERM) domain and spectrin-actin binding domain (SABD) and C-terminal domain (CTD), separately. Applying immunohistochemistry using specific antibodies for 30 kDa domain and CTD separately, the gene product of ZF4.1RH2 and ZF4.1RH3 appeared only in the ventricle and in the atrium, respectively, in mature hearts. During embryogenesis, both gene expressions are expressed starting 24 h post-fertilization (hpf). Following whole-mount in situ hybridization, ZF4.1RH3 gene expression was detected in the atrium of 37 hpf embryos. These results indicate that the gene product of ZF4.1RH3 is essential for normal morphological shape of the developing heart and to support the repetitive cycles of its muscle contraction and relaxation. [source] Targeted deletion of the ,-adducin gene (Add3) in mice reveals differences in ,-adducin interactions in erythroid and nonerythroid cells,AMERICAN JOURNAL OF HEMATOLOGY, Issue 6 2009Kenneth E. Sahr In red blood cells (RBCs) adducin heterotetramers localize to the spectrin-actin junction of the peripheral membrane skeleton. We previously reported that deletion of ,-adducin results in osmotically fragile, microcytic RBCs and a phenotype of hereditary spherocytosis (HS). Notably, ,-adducin was significantly reduced, while ,-adducin, normally present in limited amounts, was increased ,5-fold, suggesting that ,-adducin requires a heterologous binding partner for stability and function, and that ,-adducin can partially substitute for the absence of ,-adducin. To test these assumptions we generated ,-adducin null mice. ,-adducin null RBCs appear normal on Wright's stained peripheral blood smears and by scanning electron microscopy. All membrane skeleton proteins examined are present in normal amounts, and all hematological parameters measured are normal. Despite a loss of ,70% of ,-adducin in ,-adducin null platelets, no bleeding defect is observed and platelet structure appears normal. Moreover, systemic blood pressure and pulse are normal in ,-adducin null mice. ,- and ,-adducin null mice were intercrossed to generate double null mice. Loss of ,-adducin does not exacerbate the ,-adducin null HS phenotype although the amount ,-adducin is reduced to barely detectable levels. The stability of ,-adducin in the absence of a heterologous binding partner varies considerably in various tissues. The amount of ,-adducin is modestly reduced (,15%) in the kidney, while in the spleen and brain is reduced by ,50% with the loss of a heterologous ,- or ,-adducin binding partner. These results suggest that the structural properties of adducin differ significantly between erythroid and various nonerythroid cell types. Am. J. Hematol., 2009. © 2009 Wiley-Liss, Inc. [source] Subproteome analysis of the neutrophil cytoskeletonPROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 7 2009Ping Xu Abstract Neutrophils play a key role in the early host-defense mechanisms due to their capacity to migrate into inflamed tissues and phagocytose microorganisms. The cytoskeleton has an essential role in these neutrophil functions, however, its composition is still poorly understood. We separately analyzed different cytoskeletal compartments: cytosolic skeleton, phagosome membrane skeleton, and plasma membrane skeleton. Using a proteomic approach, 138 nonredundant proteins were identified. Proteins not previously known to associate with the skeleton were: n -acetylglucosamine kinase, phosphoglycerate mutase 1, prohibitin, ficolin-1, phosphogluconate dehydrogenase, glucosidase, transketolase, major vault protein, valosin-containing protein, aldehyde dehydrogenase, and lung cancer-related protein-8 (LCRP8). The majority of these proteins can be classified as energy metabolism enzymes. Such a finding was interesting because neutrophil energy metabolism is unusual, mainly relying on glycolysis. The enrichment of phosphoglycerate mutase in cytosolic skeleton was additionally indicated by the use of Western blotting. This is the broadest subcellular investigation to date of the neutrophil cytoskeletal proteome and the first proteomic analysis in any cell type of the phagosome skeleton. The association of metabolic enzymes with cytoskeleton is suggestive of the importance of their localized enrichment and macromolecular organization in neutrophils. [source] The Epiplasm Gene EPC1 Influences Cell Shape and Cortical Pattern in Tetrahymena thermophila,THE JOURNAL OF EUKARYOTIC MICROBIOLOGY, Issue 2 2004NORMAN E. WILLIAMS ABSTRACT. The cortical protein Epclp is the most abundant protein in the membrane skeleton, or epiplasm, of Tetrahymena thermophila. A partial sequence of the EPC1 gene was obtained and used to obtain a knockout construct that was successful in transforming Tetrahymena thermophila cells. The results support the conclusion that Epclp influences cell shape and the fidelity of cortical development. It was further observed that this protein is transferred from plus to minus cells during conjugation, and that the imported protein is assembled into the epiplasm of the recipient cell in a discreet series of steps. [source] WASP plays a novel role in regulating platelet responses dependent on ,IIb,3 integrin outside-in signallingBRITISH JOURNAL OF HAEMATOLOGY, Issue 3 2010Anna Shcherbina Summary The most consistent feature of Wiskott Aldrich syndrome (WAS) is profound thrombocytopenia with small platelets. The responsible gene encodes WAS protein (WASP), which functions in leucocytes as an actin filament nucleating agent ,yet, actin filament nucleation proceeds normally in patient platelets regarding shape change, filopodia and lamellipodia generation. Because WASP localizes in the platelet membrane skeleton and is mobilized by ,IIb,3 integrin outside-in signalling, we questioned whether its function might be linked to integrin. Agonist-induced ,IIb,3 activation (PAC-1 binding) was normal for patient platelets, indicating normal integrin inside-out signalling. Inside-out signalling (fibrinogen, JON/A binding) was also normal for wasp-deficient murine platelets. However, adherence/spreading on immobilized fibrinogen was decreased for patient platelets and wasp-deficient murine platelets, indicating decreased integrin outside-in responses. Another integrin outside-in dependent response, fibrin clot retraction, involving contraction of the post-aggregation actin cytoskeleton, was also decreased for patient platelets and wasp-deficient murine platelets. Rebleeding from tail cuts was more frequent for wasp-deficient mice, suggesting decreased stabilisation of the primary platelet plug. In contrast, phosphatidylserine exposure, a pro-coagulant response, was enhanced for WASP-deficient patient and murine platelets. The collective results reveal a novel function for WASP in regulating pro-aggregatory and pro-coagulant responses downstream of integrin outside-in signalling. [source] Phenylhydrazine as a partial model for ,-thalassaemia red blood cell hemodynamic propertiesBRITISH JOURNAL OF HAEMATOLOGY, Issue 6 2008Yuval Ramot Summary ,-Thalassaemia is a congenital haemoglobinopathy, associated with red blood cells (RBC) anomalies, leading to impairment of their flow-affecting properties, namely, RBC deformability, self-aggregability, and adherence to endothelial cells (EC). Treatment of normal RBC with phenylhydrazine (PHZ) causes selective association of oxidized ,-globin chains with the membrane skeleton, leading to reduced RBC deformability, characteristic of ,-thalassaemia. PHZ has thus been used to mimic phenotypes of ,-thalassaemia RBC. The present study was undertaken to further elucidate the suitability of PHZ-treated RBC as a model for ,-thalassemic RBC, by comparing the aggregability and adhesiveness of PHZ-treated RBC to those of RBC from thalassaemia intermedia (TI) patients, using image analysis of RBC under flow. In addition, the externalization of phosphatidylserine (PS), a mediator of RBC/EC interaction, was determined. It was found that PHZ caused enhanced RBC adhesiveness to extracellular matrix, similar to TI-RBC. Furthermore, in both conditions, the enhanced adhesiveness was mediated by PS translocated to the RBC surface. In contrast, PHZ treatment completely abolished RBC aggregability, while TI-RBC aggregability was slightly elevated. It is proposed that PHZ-treated RBC resemble ,-thalassaemia RBC in their deformability and adhesiveness, but not in their aggregability, and thus can be used as a limited model for ,-thalassaemia RBC phenotypes. [source] | |||||||||||||