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At least 1,261 records · Page 70Linked to original sources
PubMed1991

Abrogation of tumorigenicity by MHC class II antigen expression requires the cytoplasmic domain of the class II molecule.

Transfection of syngeneic MHC class II genes into the lethal mouse SaI tumor abrogates the malignancy of the tumor in the autologous host, and protects the host against subsequent challenges with the wild type class II- tumor. We have hypothesized that the transfectants induce protective immunity by functioning as APC for tumor peptides, and stimulating tumor-specific Th cells. Recent in vitro studies suggest that Ag presentation by class II-restricted APC requires the cytoplasmic domain of the class II molecule, and may involve intracellular signaling via the cytoplasmic domain. To determine if the class II cytoplasmic domain is required for enhanced tumor-specific immunity, SaI mouse sarcoma cells were transfected with syngeneic Aak and Abk genes with truncated cytoplasmic domains. These transfectants are as malignant as wild type class II- SaI cells in autologous A/J mice. Stimulation of tumor-specific immunity by class II+ tumor cells is therefore dependent on the class II cytoplasmic region, and may involve intracellular signaling events.

Animals↗
PubMed1990

Identification of the major cytoplasmic regions of the Neurospora crassa plasma membrane H(+)-ATPase using protein chemical techniques.

The transmembrane topography of the Neurospora crassa plasma membrane H(+)-ATPase has been investigated using purified, reconstituted components and direct protein chemical techniques. Reconstituted proteoliposomes containing H(+)-ATPase molecules oriented predominantly with their cytoplasmic surface facing outward were treated with trypsin to liberate peptides present on the cytoplasmic surface of the H(+)-ATPase as recently described (Hennessey, J.P., Jr., and Scarborough, G. (1990) J. Biol. Chem. 265, 532-537. The released peptides were then separated from the proteoliposomes by gel filtration chromatography and further purified by high performance liquid chromatography. Fourteen such peptides were identified by NH2-terminal amino acid sequence analysis, directly defining these parts of the molecule as present on the cytoplasmic surface of the membrane. Moreover, this information identified several additional flanking stretches as likely to be cytoplasmically located by virtue of the fact that they are too short to cross the membrane and return. These results and the results of other recent experiments establish 417 residues of the 919 present in the ATPase molecule, at positions 2-100, 186-256, 441-663, and 897-920, as cytoplasmically located. Taken together with the results of our preliminary investigations of the membrane embedded sectors of the ATPase, this information allows the formulation of a reasonably detailed model for the transmembrane topography of the ATPase polypeptide chain.

Amino Acid Sequence↗
PubMed1990

A positively charged region is a determinant of the orientation of cytoplasmic membrane proteins in Escherichia coli.

Basic amino acid residues were introduced into an extracellular (periplasmic) domain, preceding a membrane-spanning hydrophobic domain, of SecY, an integral cytoplasmic membrane protein. The localization of the domain was monitored as to the alkaline phosphatase activity of TnPhoA fused adjacent to the domain. The alkaline phosphatase activity of such Escherichia coli cells drastically decreased when positive charges were introduced, indicating that on the introduction the SecY domain showed a change in localization from the periplasm to the cytoplasm. In another experiment, positive charges were introduced to the same periplasmic domain of another SecY-PhoA fusion protein, in which PhoA is fused to the cytoplasmic domain of SecY following the particular hydrophobic domain. The alkaline phosphatase activity increased drastically when positive charges were introduced, indicating that the SecY domain fused to PhoA showed a change in localization from the cytoplasm to the periplasm. In both experiments, the removal of a large amino-terminal portion of the SecY domain did not alter the effect of the positive charge introduction. Changes in localization of SecY domains thus demonstrated were also supported by a protease accessibility test on spheroplasts. It is proposed that a positively charged region adjacent to a membrane-embedded hydrophobic region tends to be stabilized on the cytoplasmic surface of the membrane, which in turn endows the hydrophobic region with the ability to act as a stop-transfer sequence or a signal sequence and consequently determines the orientation of the hydrophobic region in the membrane.

Amino Acid Sequence↗
PubMed1990

Deletions in the cytoplasmic domain of the polymeric immunoglobulin receptor differentially affect endocytotic rate and postendocytotic traffic.

We have examined the function of the cytoplasmic domain of the polymeric immunoglobulin receptor (pIg-R) by producing two separate deletions in the cytoplasmic domain of the pIg-R, expressing the mutant receptors in polarized MDCK cells, and analyzing each for their effects on receptor and ligand traffic. Deletion of the C-terminal 30 amino acids (726-755) reduces the rate of internalization of receptor-bound ligand from the basolateral surface. However, this mutation has no effect on delivery of receptor from the Golgi to the basolateral surface or the post-endocytotic traffic of receptor and ligand. Mutation of a tyrosine at position 734 to serine produces a receptor with a similar phenotype. If residues 670-707 are deleted from the middle of the cytoplasmic domain, both basolateral delivery and internalization are unaffected. However, unlike wild type, after endocytosis from the basolateral surface, both receptor and ligand are largely degraded. We reported previously that deletion of the entire cytoplasmic domain prevents the basolateral delivery of newly synthesized receptor (Mostov, K.E., de Bruyn Kops, A., and Deitcher, D.L. (1986) Cell 47, 359-364). In contrast, the mutants reported here are delivered to the basolateral surface, suggesting that only residues 653-669 and/or 708-725 are necessary for basolateral delivery. Thus, different deletions in the cytoplasmic domain of the pIg-R can produce mutant receptors which alter different aspects of receptor traffic.

Animals↗
PubMed1986

Acute effect of 5-fluorouracil on cytoplasmic and nuclear dihydrofolate reductase messenger RNA metabolism.

Studies were completed in C3-L5178Y cells, in which the DNA-coding region for dihydrofolate reductase messenger RNA (DHFR-mRNA) is amplified, to determine the acute effect of 5-fluorouracil (FUra) on DHFR-mRNA metabolism. There was minimal to no effect of 100 microM FUra on total cytoplasmic DHFR-mRNA levels by 6 and 12 h and only a 25% reduction by 24 h. These results contrasted with the nuclear DHFR-mRNA levels which by 6 h following exposure to FUra increased by 80% in a dose-dependent manner. Furthermore, some of the increased nuclear DHFR-mRNA was found to be in a non-polyadenylated form. Under conditions to examine only RNA synthesized during the drug exposure, FUra was found to markedly enhance the level of newly synthesized nuclear DHFR-mRNA in a dose-dependent manner, while also producing an apparent dose-dependent reduction in the cytoplasmic DHFR-mRNA. RNA fractionated by 1.5% agarose-urea gel electrophoresis revealed two major cytoplasmic DHFR-mRNA species approximately 1.8 and 0.8 kilobases in size. Following a 24-h FUra exposure, a dose-dependent loss of the 0.8-kilobase DHFR-mRNA was observed. The combined results of these experiments indicate that FUra treatment reduces the ability of nascent DHFR-mRNA to relocate to the cytoplasm, suggesting either an inhibition of mRNA processing or nuclear-cytoplasmic transport.

Animals↗
PubMed1989

[Effect of active oxygen on cytoplasmic Ca2+ sequestration mechanism].

Effects of the active oxygen on the extrusion mechanism of once-increased cytoplasmic Ca2+, which causes various physiological phenomena, were investigated using different kinds of culture cells. First we found that, in response to stimulation with vitamin K (VK), various culture cells showed a decrease in cytoplasmic Ca2+ concentration. On the presumption that this phenomenon might be related to the oxidizing action of VK, we performed the same experiments using oxidizing agents such as H2O2 or KO2. They also showed a decrease in cytoplasmic Ca2+ concentration. Furthermore, they suppressed the increase of cytoplasmic Ca2+ by vasopressin. It would be inferred from these results that the active oxygen may act upon some site of the cellular signal transduction system of cell membrane to lower the cytoplasmic Ca2+ level.

Animals↗
PubMed1989

Leucine induces initial lowering of cytoplasmic Ca2+ in pancreatic beta-cells without concomitant inhibition of insulin release.

The early effects of glucose and leucine on cytoplasmic Ca2+ and insulin release were compared in suspensions of cells prepared by dispersal of the beta-cell-rich pancreatic islets of ob/ob-mice. Adequate temporal resolution was achieved by continuously recording the 340/380 nm fluorescence excitation ratio from cells loaded with the Ca2+ indicator fura-2 and measuring insulin in the perifusate from cells mixed with polyacrylamide beads. Raising the glucose concentration from 3 to 20 mM resulted in concomitant reductions of cytoplasmic Ca2+ and insulin release during the first minute. Whereas 10 mM leucine was as efficient as glucose in inducing temporary lowering of cytoplasmic Ca2+, this amino acid did not depress insulin release. It is concluded that the initial decrease of cytoplasmic Ca2+ is a phenomenon coupled to stimulation of the metabolism. The leucine-induced lowering of Ca2+ may essentially reflect changes in cytoplasmic pools other than in a peripheral one regulating insulin release.

Animals↗
PubMed1977

[Study of the cytostatic action of the cytoplasm of oocytes and mature ova of the common frog and sturgeon (Acipenser stellatus)].

The presence of the cytostatic factor found by Masui and Markert (1971) in the cytoplasm of oocytes and eggs of Rana pipiens was verified in the cytoplasm and mature inactivated eggs of the common frog Rana temporaria and sevryuga Acipenser stellatus. The cytoplasm was injected to the embryos of the same species at different phases of the first cleavage division in the animal region of one of the two blastomeres. The injection of cytoplasm did not cause the arrest of cleavage divisions. Many embryos proceeded to gastrulation. The cytoplasm of maturing oocyte and mature egg in the common frog and sevryuga, unlike in R. pipiens, has no cytostatic effect.

Animals↗
PubMed1988

In vitro regulatory mechanisms for cytoplasmic maturation of murine megakaryocytes derived from colony-forming units megakaryocyte (CFU-M).

We studied the effects of recombinant erythropoietin (r-Epo) and thrombocytopenic serum (TS) on the cytoplasmic maturation of megakaryocytes derived from colony-forming units megakaryocyte (CFU-M). Serotonin content, ATP content, acetylcholinesterase (Ach-E) activity per megakaryocyte, and electron microscopic analysis were selected as markers of cytoplasmic maturation. Megakaryocytes induced by pokeweed mitogen-stimulated spleen cell-conditioned medium (PWM-SCM) alone showed low levels of ATP content and Ach-E activity, which did not increase during culture, as well as a low level of serotonin content, which gradually accumulated during day 7 of culture. When r-EPo was added to the culture system on day 3 after megakaryocytic colony formation with PWM-SCM, the serotonin content in megakaryocytes increased markedly but ATP content and Ach-E activity did not increase significantly. In contrast, TS caused an increase in ATP content and Ach-E activity, but did not cause an increase in serotonin content. Electron microscopic analysis showed that the demarcation membrane system (DMS) developed defectively only in local areas with the addition of r-Epo and PWM-SCM, whereas the DMS developed normally and dense granules were generated to near normal with the addition of TS. Recombinant Epo may act on the early stage of cytoplasmic maturation, whereas TS may act on the late stage of cytoplasmic maturation. The results shown herein suggest that at least two different factors may be necessary for full in vitro cytoplasmic maturation of megakaryocytes derived from CFU-M.

Acetylcholinesterase↗
PubMed1986

Generation of cells expressing cytoplasmic and/or surface T-cell receptor beta chains during the development of mouse fetal thymus.

We have used a double immunofluorescence technique to identify newly formed cells expressing cytoplasmic and/or surface T-cell receptor beta chains in mouse embryo thymus. In addition, we have localized these cells to the developing thymic cortex by immunoperoxidase labelling of frozen sections. The first positive cells appear in small numbers in the thymus at Day 15 of gestation and express cytoplasmic but not surface beta chains (C beta+). At Day 16 of gestation, cells appear that express small capped areas of surface beta chains as well as cytoplasmic beta chains (C beta+S beta+ cells). Just prior to birth, a cohort of cells appears that express surface beta chains but no cytoplasmic beta chains detectable by the methods employed (S beta+ cells). These results and vincristine mitosis-blocking studies suggest that a proliferating population of cortical thymocytes accumulate cytoplasmic beta chains before giving rise to T cells that express surface beta chains (presumably in association with alpha chains). When a monoclonal antibody to beta chains is present in organ cultures of embryonic thymus, C beta+ thymocytes continue to be generated but there is an absence of S beta+ cells. The latter appear after the cultures are transferred to fresh medium not containing the antibody.

Animals↗
PubMed1986

Regulation of development in the fully grown mouse oocyte: chromosome-mediated temporal and spatial differentiation of the cytoplasm and plasma membrane.

The relationship between nuclear maturation and the differentiation of the cytoplasm and plasma membrane during resumption of arrested meiosis was investigated by culture of GV-and MII-stage mouse oocytes in the presence and absence of nocodazole. Culture in the presence of nocodazole was associated with dispersal of MI and MII chromosomes throughout the subplasmalemmal cytoplasm. A progression of cortical (thickening of actin filaments) and plasma membrane changes (denudation of microvilli, reduction in cell surface glycoproteins, formation of chromosome-containing evaginations) that normally occurs in proximity to chromosomes associated with intact MI or MII spindles took place only in those regions of the cortical cytoplasm containing the dispersed subplasmalemmal chromosomes. The dispersion and migration of the chromosomes occurred in an apparently random fashion. Fluorescent probe analysis of normal and treated oocytes indicated a stage-specific association between the spatial distribution of chromosomes and mitochondria. Transfer of individual bivalent chromosomes to untreated oocytes at different stages of maturation and to cytoplasts derived from oocytes anucleated prior to GVB demonstrated the necessity of chromosomes for cytoplasmic and plasma membrane differentiation, and that the capacity of the cytoplasm and plasma membrane to differentiate in response to the presence of a chromosome is acquired prior to GVB.

Animals↗
PubMed1986

Simultaneous measurement of stimulus-induced changes in cytoplasmic Ca2+ and in membrane potential of human neutrophils.

The activation of human neutrophils by chemotactic peptides evokes a rapid change in membrane potential and an increase in cytoplasmic Ca2+ levels. These events are followed up to a minute later by detectable levels of microbicidal agents formed by the oxidative burst. Except for the latter, the sequence of events has remained unclear. We report here that a new fluorescent Ca2+ indicator developed by R. Tsien, Indo-1, has allowed us to resolve the temporal relationship between the rapid and transient cytoplasmic Ca2+ rise and the membrane potential change and to do so on very small samples by using a fluorescence-activated cell sorter. We have adapted a FACS 440 for simultaneous single cell membrane depolarization and cytoplasmic [Ca2+] detection in human neutrophils upon stimulation with formyl-methionyl-leucyl-phenylalanine (fMLP). A membrane potential probe, dipentyloxacarbocyanine, allows us to determine that the membrane potential change is fMLP dose-dependent and apparently biphasic. The depolarization is maximal 40 s after stimulation. In contrast, cytosolic [Ca2+], while fMLP-dose dependent, is maximal at 10 s and already decreasing rapidly when the cell has reached its lowest potential. It can be measured with Indo-1 which has a fluorescence emission (lambda ex = 357 nm) maximum at 485 nm when Ca2+-free and 405 nm when Ca2+-liganded. The ratio of these fluorescences may then be calibrated in terms of cytoplasmic Ca2+ levels. Thus, Ca2+ release into the cytoplasm becomes the earliest evidence of neutrophil stimulation by fMLP and occurs in close association with an apparent membrane hyperpolarization.

Calcium↗
PubMed1985

Morphologic demonstration of cytoplasmic ASSAM-related antigenic substance (CASSAM) by an immunoperoxidase technique.

Murine senile amyloid protein identified in the senescence-accelerated mouse (SAM) was called ASSAM, and the ASSAM-related antigenic substance was detected in the cytoplasm of hepatocytes, columnar epithelia of the small intestine, and epithelia of the proximal convoluted tubules of the kidney, with the use of an immunoperoxidase method. This substance, called CASSAM (cytoplasmic ASSAM-related antigenic substance), did not stain positively with Congo red nor fibril structure, as determined under an electron microscope. As the ASSAM (senile amyloid) deposition increased with advancing age, CASSAM observed in hepatocytes and columnar epithelia decreased both in SAM-P and SAM-R strains. In the liver of the SAM-P strain in particular, the incidence and intensity in deposition of ASSAM increased rapidly from 5 months of age; on the other hand, CASSAM observed in the hepatocytes decreased rapidly at about the same time. Cycloheximide-treated animals showed a significantly low concentration of SASSAM (serum ASSAM-related antigenic substance) and also a low incidence and intensity of CASSAM observed in the cytoplasm of the hepatocytes and epithelia of the small intestine. In colchicine-treated animals, SASSAM concentration was slightly lower, and the severity of CASSAM observed in the cytoplasm was slightly higher, in the liver and kidney, as compared with control values. CASSAM is assumed to be synthesized in the cytoplasm of the cell and to be secreted alone or in the lipoprotein form into the serum. This CASSAM or lipoprotein including CASSAM is perhaps a constituent of SASSAM (CASSAM is assumed to include apoSASSAM) and the hepatocytes and intestinal mucosal epithelia are possible production sites of apoSASSAM.

Aging↗
PubMed1974

[The distribution of nuclear and cytoplasmic proteins in the blastoderm of the loach, Misgurnus fossilis L].

The concentration of dry substance (protein) and the dry weight of nuclei, cytoplasm and cells from different blastoderm regions at the early blastula and midgastrula stages were determined by interferentional microscopy. It was shown that at the early blastula stage the dry weight of cells in the basal layer is higher than that in the outer layer. Although the protein concentration in the basal layer cells appears to be somewhat higher, differences in their dry weight are due primarily to the big volume of cytoplasm of the basal layer cells. By the midgastrula stage, the total (nucleus + cytoplasm) protein concentration increases (by 17% in the basal layer cells and by 9% in the outer layer cells) due to the increase of nuclear protein concentration. At the same time dry weight of these cells markedly decreases due to the decrease of their volumes in the process of cell divisions. At the midgastrula stage the epiblast cells have the highest dry weight due to the highest protein concentration in the cytoplasm and the biggest cell volume. The results obtained are discussed with respect to the data on the pattern of accumulation of newly synthesized protein in nuclei and cytoplasm with special reference to the duration of individual cell cycle phases.

Animals↗
PubMed1979

The estimation of velocity distribution profile of Paramecium cytoplasmic streaming.

Cytoplasmic streaming of Paramecium bursaria and Paramecium tetraurelia was investigated by cinematographic techniques. Analysis of the records reveals the paraboidal character of the velocity distribution profiles in all arbitrarily chosen zones along the whole route of moving cytoplasm in the cell. According to the date obtained from cytoplasmic streaming analysis and food vacuole path, the geometry of the "channel" was described in terms of ellipsoid axes. Total volume changes of cytoplasm flowing through a given cross section in a given time unit were computed and no significant differences were found. The participation of a pressure gradient in motive force generation in cytoplasmic streaming is discussed.

Animals↗
PubMed1979

Cytoplasmic fragments causing spurious platelet counts in the leukemic phase of poorly differentiated lymphocytic lymphoma.

A patient with a leukemic phase of poorly differentiated lymphocytic lymphoma had a spuriously high automated platelet count because of cytoplasmic fragments. The number and relative percentage of cytoplasmic fragments increased during chemotherapy. The cytoplasmic fragments did not interfere with platelet aggregation using adenosine diphosphate, collagen, and epinephrine even though they were found in platelet-rich plasma. The ultrastructure of the cytoplasmic fragments is discussed. Cytoplasmic fragments as a cause of spuriously high automated platelet counts should be considered in cases of leukemic patients.

Adult↗
PubMed1982

Tissue distribution, molecular profile, and shedding of a cytoplasmic antigen identified by the monoclonal antibody 465.12S to human melanoma cells.

The mouse immunoglobulin G2 monoclonal antibody (MoAb) 465.12S reacts with a cytoplasmic antigen present in human melanoma cells but not detectable in melanocytes. Indirect immunofluorescent staining of a large number of surgically removed normal adult and fetal tissues with the MoAb 465.12S detected the cytoplasmic antigen in epithelial cells from several organs. The intensity of staining was greater in adult tissues than in the corresponding fetal tissues. Furthermore, the MoAb 465.12S stained nearly all of the surgically removed tumors tested but did not stain many of the normal tissues from which they originated. In almost all cases, the intensity and frequency of staining wa greater for tumor cells than for corresponding normal tissues. From cultured carcinoma and melanoma cells, the MoAb 465.12S immunoprecipitated four glycopolypeptides with molecular weights of 94,000, 75,000, 70,000, and 25,000. Incorporation of 3H-labeled sugars into the various components of the cytoplasmic antigen revealed that the M.W. 75,000 component was unusual in that it contained only glucosamine and mannose. The antigenic determinant defined by the MoAb 465.12S appears to be protein rather than carbohydrate in nature since it is heat sensitive and is expressed on the antigens synthesized by cells in presence of tunicamycin. Analysis of the spent culture medium of carcinoma and melanoma cell lines revealed that the cytoplasmic antigen is readily shed by these cells and consists of a major M.W. 94,000 and a minor M.W. 72,000 component. Treatment of cultured melanoma cells with the antibiotic tunicamycin showed that glycosylation of the cytoplasmic antigen is required for its shedding and/or stability in the spent culture medium.

Antibodies, Monoclonal↗
PubMedDate not supplied

[Nuclear and cytoplasmic RNA showing hybridization with repetitive sequences B1 and B2 of the mouse genome].

Heavy nuclear RNA contains high amounts of transcripts from repetitive sequences B1 and B2. Cytoplasmic poly(A+)RNA and in particular, polysomal poly(A)+RNA (mRNA) also contain these sequences though their content is much lower. An abundant 2 kb mRNA of mouse liver was found to contain B2 sequence. A novel class of RNA is described namely as low molecular weight heterogeneous (200-400 nucleotide long) poly(A)+RNA. These RNAs contain B1 and B2 sequences and are located mostly in the cytoplasm but not in polysomes. Tumor cells contain high amounts of small B2+RNAs. The content of B1+ small cytoplasmic RNA is much lower. The hybridization tests confirm the existence of extended homology between B1 sequence and 4.5 S small nuclear RNA which has been earlier deduced from comparison of their base sequences. Homology of B1 and small cytoplasmic 7S RNA was also detected. No hybridization of B2 to small nuclear or small cytoplasmic RNAs could be detected.

Animals↗