SEARCH · Search PubMed
Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.
Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.
Explore the source record for details and available documents.
Explore the source record for details and available documents.
Explore the source record for details and available documents.
The membrane fraction isolated from the crab nerve was treated by ultrasonic irradiation to obtain closed osmotically active vesicles. The liposomes from brain lipids were modified by a cytoplasmic fraction to assay the tetrodotoxin influence on the 22Na influx into such liposomes. The 22Na influx into vesicles and modified liposomes was studied in the presence and absence of substances which change sodium conductance of excitable membranes. It is shown that tetrodotoxin at concentrations blocking electrical excitation in vivo decreased the sodium influx into membrane vesicles and modified liposomes.
The cytoplasmic domain of band 3 was released from spectrin-depleted, acetic acid-stripped erythrocyte membrane vesicles by mild chymotryptic digestion. After purification by ion exchange and gel filtration chromatography, the fragment preparation was found to be greater than 90% pure on polyacrylamide disc gels in the presence of 0.2% sodium dodecyl sulfate. The subunit Mr from the electrophoretic procedure was estimated at approximately 40,000. The isolated cytoplasmic fragment ws judged to be a dimer, since (i) the unmodified fragment and its disulfide-cross-linked (dimeric) counterpart eluted in the same peak fraction from a Sephacryl S-200 gel filtration column, and (ii) the sedimentation velocity molecular weight of the native fragment was calculated to be approximately 95,000. No evidence of either larger or smaller aggregates was obtained. The frictional ratio of the fragment was measured at 1.6, suggesting a highly elongated morphology. The circular dichroism spectrum of the fragment corresponded to approximately 37% alpha helix. Titration of the cytoplasmic fragment over the physiological pH range gave rise to a reversible 2-fold increase in the intrinsic fluorescence quantum yield (lambda ex, 290 nm; lambda em, 335 nm) between pH 6 and 9. Computer analysis of the data yielded a temperature-dependent apparent pKa of 7.8 at 37 degrees C and 8.1 at 20 degrees C, both with Hill coefficients less than or equal to 1. Calorimetric experiments revealed a similar sensitivity to pH, where the denaturation temperature of the fragment titrated from 74 degrees C at pH 6 to 59 degrees C at pH 8.5, with an apparent pKa of 7.3 and a Hill coefficient less than 1. The enthalpies and widths at half-height of the transitions were also exquisitely sensitive to pH. The fluorescence and calorimetric data could all be described by the titration of a single ionizable group of apparent pKa of 7.8 at 37 degrees C and delta pKa/degrees C of -0.018. The ionization of this critical group is suggested to exert significant control over the structure/stability of the cytoplasmic domain of band 3.
Double-stranded segments are found in poly(A)+hnRNA as well as in poly(A)-hnRNA, but in the second case they have partly intermolecular character and may originate from an interaction between primary transcripts and snRNA. Fragments of hnRNA containing double-stranded segments are preferentially removed from the nucleus. In cytoplasmic poly(A)+RNA double-stranded segments longer than 20 base pairs represent about 2%. In cytoplasmic poly(A)-RNA intermolecular double-stranded segments derived from rRNA prevail. Double-stranded fragments from nuclear RNP and free cytoplasmic mRNP exhibit similar affinity to hydroxyapatite but their base compositions are different: low content of adenine and high of guanosine in nuclear in contrast to nearly symmetrical base composition in cytoplasmic fragments.
Integrin alphaIIbbeta3 is a heterodimeric (alpha/beta) cell surface receptor critical for platelet aggregation, and its dysfunction is linked to thrombosis and a number of other vascular diseases. Upon agonist stimulation, which leads to platelet aggregation, alphaIIbbeta3 is activated via a distinct inside-out signaling pathway, i.e., the short alphaIIb/beta3 cytoplasmic tails receive intracellular signals, which trigger the conformational change of the extracellular domain for the high-affinity ligand binding. The structural basis for how the alphaIIbbeta3 cytoplasmic face regulates the inside-out activation of the receptor has been extensively studied over the past decade. We have recently used nuclear magnetic resonance (NMR) spectroscopy to characterize and determine the structural features of the alphaIIbbeta3 cytoplasmic domain. This chapter describes detailed practical procedures for performing these NMR studies, which have provided key atomic insights into the mechanism of the alphaIIbbeta3 function, especially its inside-out signaling.
In Treponema denticola, a ribbon-like structure of cytoplasmic filaments spans the cytoplasm at all stages of the cell division process. Insertional inactivation was used as a first step to determine the function of the cytoplasmic filaments. A suicide plasmid was constructed that contained part of cfpA and a nonpolar erythromycin resistance cassette (ermF and ermAM) inserted near the beginning of the gene. The plasmid was electroporated into T. denticola, and double-crossover recombinants which had the chromosomal copy of cfpA insertionally inactivated were selected. Immunoblotting and electron microscopy confirmed the lack of cytoplasmic filaments. The mutant was further analyzed by dark-field microscopy to determine cell morphology and by the binding of two fluorescent dyes to DNA to assess the distribution of cellular nucleic acids. The cytoplasmic filament protein-deficient mutant exhibited pleiotropic defects, including highly condensed chromosomal DNA, compared to the homogeneous distribution of the DNA throughout the cytoplasm in a wild-type cell. Moreover, chains of cells are formed by the cytoplasmic filament-deficient mutant, and those cells show reduced spreading in agarose, which may be due to the abnormal cell length. The chains of cells and the highly condensed chromosomal DNA suggest that the cytoplasmic filaments may be involved in chromosome structure, segregation, or the cell division process in Treponema.
BACKGROUND: DNA structure checkpoints are conserved eukaryotic signal transduction pathways that help preserve genomic integrity. Upon detecting checkpoint signals such as stalled replication forks or double-stranded DNA breaks, these pathways coordinate appropriate stress responses. Members of the PI-3 kinase related kinase (PIKK) family are essential elements of DNA structure checkpoints. In fission yeast, the Rad3 PIKK and its regulatory subunit Rad26 coordinate the detection of checkpoint signals with pathway outputs. RESULTS: We found that untreated rad26Delta cells were defective for two microtubule-dependent processes: chromosome segregation and morphogenesis. Interestingly, cytoplasmic accumulation of Rad26-GFP occurred following treatment with microtubule destabilizing drugs, but not during treatment with the genotoxic agent Phleomycin. Cytoplasmic accumulation of Rad26-GFP depended on Rad24, a 14-3-3 protein also required for DNA structure checkpoints and morphogenesis. Results of over expression and epistasis experiments confirm that Rad26 and Rad24 define a response to microtubule destabilizing conditions. CONCLUSION: Two DNA structure checkpoint proteins with roles in morphogenesis define a response to microtubule destabilizing conditions.
Scanning electron microscopy (SEM) has had a shorter time course in biology than conventional transmission electron microscopy (TEM) but has nevertheless produced a wealth of images that have significantly complemented our perception of biological structure and function from TEM information. By its nature, SEM is a surface imaging technology, and its impact at the subcellular level has been restricted by the considerably reduced resolution in conventional SEM in comparison to TEM. This restriction has been removed by the recent advent of high-brightness sources used in lensfield emission instruments (FEISEM) which have produced resolution of around 1 nanometre, which is not usually a limiting figure for biological material. This communication reviews our findings in the use of FEISEM in the imaging of nuclear surfaces, then associated structures, such as nuclear pore complexes, and the relationships of these structures with cytoplasmic and nucleoplasmic elements. High resolution SEM allows the structurally orientated cell biologist to visualise, directly and in three dimensions, subcellular structure and its modulation with a view to understanding, its functional significance. Clearly, intracellular surfaces require separation from surrounding structural elements in vivo to allow surface imaging, and we review a combination of biochemical and mechanical isolation methods for nuclear surfaces.
Functional significances of the cytoskeleton and the plasmalemmal undercoat are discussed with special reference to their ultrastructural aspects. The cytoskeleton is mainly composed of three distinctive cytoplasmic fibrous structures, namely, microtubules, actin filaments and intermediate filaments. Closely associated with such fibrous structures are a variety of proteins, which have been shown not only to control the formation or breakdown of the fibrous structures but also to involve the construction of a higher order of fibrous architectures such as bundles and networks. Thus, these cytoskeletal fibrous components may form the respective systems, though such systems often interact with each other for the cytoplasmic integrity. The fundamental function of the cytoskeleton is to provide structural frameworks for the cytoplasm. Through such frameworks the cytoskeleton is involved in various cellular organizations and activities such as cell shape, distribution of cell organelles, cell motility, membrane phenomena, and metabolism. Much interest has also been drawn to the interaction between the plasmalemma and the cytoskeleton. Part of such interaction can be assigned to the plasmalemmal undercoat, though it indeed constitutes a part of the cytoskeleton. The significance of the plasmalemmal undercoat may be to primarily provide structural supports for the plasmalemma, playing various roles in cell physiology.
A 57-yr-old woman presented to the National Cancer Institute (NCI) with a history of nephrectomy for a clear cell renal cell carcinoma (RCC), Fuhrman grade 3 of 4 diagnosed 1 yr prior to admission to the NCI. A CT scan done upon admission revealed multiple bilateral lung masses. A CT-guided fine-needle aspiration (FNA) of one of the lung masses revealed a cellular specimen composed primarily of follicular structures surrounding dense hyalinized central cores. The cells in the follicular structures displayed bland nuclei and had granular to vacuolated cytoplasm. Papillary structures were also appreciated. Immunocytochemical studies showed tumor cells that were strongly vimentin and TFE3 positive. Focal staining for AE1/AE3 and CD10 was observed, as was negative staining for EMA. A surgical biopsy specimen reflected the FNA findings and demonstrated a similar immunoprofile. These findings correspond to the recently described Xp11.2 translocation/TFE3 fusion renal cell carcinoma. To our knowledge, this is the first report describing the cytologic features of an Xp11.2 translocation/TFE3 fusion RCC.
Round eosinophilic hyaline inclusion bodies with halos in the somata of anterior horn cells from a case of sporadic lower motor neuron disease (MND) were intensely immunostained with the monoclonal anti-ubiquitin antibody (DF2). A few similar, DF2-positive inclusions were also observed in the nerve cell processes of anterior horn cell or in the neuropil. Most inclusions showed intense homogeneous staining of the entire inclusion, whereas a few had intense staining of their periphery with no or pale staining of the central areas. Other DF2-positive structures in the somata of anterior horn cells included cytoplasmic granular structures, eosinophilic thread-like or reticular structures, and small eosinophilic profiles different from Bunina bodies. The DF2-staining intensity of Bunina bodies and spheroids did not exceed the background level. These results suggest that ubiquitination is associated with a pathological process of anterior horn cell degeneration in this MND case.
We aimed at understanding of formation and function of the "Nieuwkoop Centre" in embryonic pattern formation. Discussed are data on genesis of cytoplasmic localizations in ovarian oocytes, transient modifications of cytoskeletal structures creating cytoplasmic asymmetries in fertilized eggs, the axis determining "vegetal cortical rotation" and fate of distinct cells, as shown by injection of specific molecular markers into particular blastomeres at specific times. Egg rotation and centrifugation suggested that sperm that gravity cooperate in symmetrization of the axially symmetrical anuran egg. After fertilization in space or in a fast rotating clinostate, axis formation and embryonic development were normal although the blastocoel was transiently abnormal. Normal tadpoles came back on Earth after ovulation, fertilization and culture in space. They metamorphosed normally and got healthy Earth-born F1 offspring. We conclude that neither sperm nor gravity are required for determination of the bilateral symmetry in the embryo of Xenopus laevis. In normal development sperm and gravity, either alone or in collaboration, may overrule an initial bilaterality inherent to, the full-grown oocyte, residing in some still unidentified component(s)/or mechanisms.
Oncocytic carcinoma arising in the submandibular gland is a very rare tumor that has only previously been reported in nine cases. This paper describes an additional case of oncocytic carcinoma in the right submandibular gland. The patient was a 55-year-old Japanese male who was presented with a 3 cm right submandibular mass. Both radical resection of the tumor and right supraomohyoid neck dissection were performed. Histologically, the tumor cells exhibited an abundant eosinophilic cytoplasm, which appeared to be finely granular, and invaded the surrounding tissues. In addition, perineural invasion was also observed. Electron microscopy demonstrated the presence of an abundance mitochondria in the cytoplasm of the tumor cells. However, since the fine structure of cytoplasm was destroyed by fixation and embedding of the tissue for the light microscopy, the cytoplasm of the tumor could not be observed clearly. Immunohistochemically, anti-mitochondrial antibody positivity was evident in the cytoplasm of the tumor cells. Consequently, we diagnosed the mass as oncocytic carcinoma. Usually, electron microscopy is necessary to diagnose oncocytic lesions, but the paraffin-embedded tissue was not suitable for electron microscopy. Immunohistochemistry using an anti-mitochondrial antibody was found to be useful and helpful for the diagnosis of oncocytic lesions.
Human embryos from the 2-cell to the morula stage developing in vitro after monospermic fertilization were incubated with [3H]thymidine or [3H]uridine and those possessing multinucleated blastomeres were examined by conventional transmission electron microscopy and by light- and electron-microscope autoradiography. Labelled DNA was present in all nuclei showing normal ultrastructural appearance, in pseudonuclei lacking nucleoli and their precursors and often demonstrating an unusual chromatin organization, free in the cytoplasm in structures resembling aggregates of dense chromatin and in small cytoplasmic vesicles in the close vicinity of these aggregates. The labelling with [3H]thymidine was not detected in about 50% of the cytoplasmic chromatin aggregates, suggesting that this extranuclear DNA was no longer replicated. Signs of extrusion of pseudonuclei to the extracellular space were occasionally observed. RNA synthesis could not be detected on free cytoplasmic chromatin, while [3H]uridine was incorporated into all nuclei and most pseudonuclei of multinucleated blastomeres of embryos at the 8-cell and morula stages. However, the major outburst of transcriptional activity and the ultrastructural cytoplasmic changes typical of the assumption of embryonic genome expression, occurring normally at the 8-cell stage, were observed only exceptionally.
The patient reported had a fulminant, refractory diarrhea of unknown etiology producing about 81 of diarrheal fluid daily which required continuously large volumes of intravenous fluid therapy. No protozoon other than Blastocystis hominis was present. Bacterial counts were very low in the fecal material because of continuous antibiotic therapy. Blastocystis hominis was present in large numbers, averaging for a 5-day period, 8.3 X 10(6)/ml of diarrheal fluid. Treatment with metronidazole for 12 days had no effect on either the diarrhea or the numbers of B. hominis present. The patient died from aspiration pneumonia without a firm diagnosis of his underlying disease. Identification of an unusual form of B. hominis seen only in diarrheal fluid was confirmed through cultivation, specific fluorescent antibody staining, as well as by transmission electron microscopy and light microscopy using hematoxylin staining and Nomarski interference contrast. The most significant result of this study is the description of ultrastructure of the in vivo trophozoite form of B. hominis. The persistence of certain morphologic features of the in vivo and in vitro forms of B. hominis is described. In vivo, B. hominis has a larger nucleus than in vitro, more prominent, well structured nucleoli, and hundreds of mitochondria with numerous delicate saccular cristae in a clear electron-lucent matrix and complex internal structure. Many cytoplasmic vesicles lined with ribosomes are present. The in vivo structure differs from the granular culture form of B. hominis, which has a smaller nucleus (nuclei), no distinct nucleoli, mitochondria characterized by a uniform structureless electron dense matrix, and few cytoplasmic, ribosome-lined vesicles. A feature retained in both in vivo and in vitro B. hominis is a distinct crescentic band of nuclear chromatin.
Ultrastructural changes associated with ascospore formation in Saccharomyces cerevisiae were investigated by using freeze-etching and thin-sectioning techniques. The first nuclear division (meiosis I) is indicated by the appearance of spindle fibers within the nucleus. The nucleus subsequently elongates and eventually assumes a barbell shape; the second nuclear division (meiosis II) occurs before nuclear separation. The spindle fibers involved in meiosis II appear to be oriented perpendicular to those observed in meiosis I. A discrete bilaminar structure (forespore wall) progressively delineates each ascospore nucleus and encloses cytoplasmic material including mitochondria and endoplasmic reticulum. The forespores then elongate, close off, and become separated from the ascus cytoplasm by membranes. The ascospores assume a spherical shape as spore coat material is laid down; the latter stages of ascospore formation are characterized by thickening of the ascospore wall and disintegration of the ascus cytoplasm. No structures which could be identified as chromosomes were observed.