Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “RNA Aggregation”

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.

At least 1,207 records · Page 67Linked to original sources
PubMed1993

Expression of Wnt-1 in PC12 cells results in modulation of plakoglobin and E-cadherin and increased cellular adhesion.

The Wnt-1 gene plays an essential role in fetal brain development and encodes a secreted protein whose signaling mechanism is presently unknown. In this report we have investigated intracellular mechanisms by which the Wnt-1 gene induces morphological changes in PC12 pheochromocytoma cells. PC12 cells expressing Wnt-1 show increased steady-state levels of the adhesive junction protein plakoglobin, and an altered distribution of this protein within the cell. This effect appears similar to a modulation of the plakoglobin homolog, Armadillo, that occurs in Drosophila embryos in response to the Wnt-1 homolog, wingless (Riggleman, B., P. Schedl, and E. Wieschaus. 1990. Cell. 63:549-560). In addition, PC12/Wnt-1 cells show elevated expression of E-cadherin and increased calcium-dependent cell-cell adhesion. These results imply evolutionary conservation of cellular responses to Wnt-1/wingless and indicate that in certain cell types Wnt-1 may act to modulate cell adhesion mechanisms.

Animals↗
PubMed1983

Role of laminin in epithelium formation by F9 aggregates.

The formation and maturation of the outer epithelial layer is essential for maximal alphafetoprotein (AFP) production during differentiation of F9 embryoid bodies in the presence of 5 X 10(-8) M retinoic acid (Grover et al., 1983. J. Cell Biol. 96:1690-1696). The critical phase is between the third and the fourth day when the components of the extracellular matrix organize into a basement membrane. The role of some of these components in the process of epithelium formation and maturation is analyzed in this paper. The role of laminin was investigated by testing the effect of exogenous laminin and antilaminin in cultures of differentiating F9 aggregates. Tests included growth rates, morphological changes, AFP production, determination of AFP mRNA levels, and fluorescent staining for basement membrane components and for epithelial markers. At concentrations greater than 5 micrograms/ml, exogenous laminin inhibited the production of AFP and prevented AFP gene transcription. On the basis of immunofluorescence tests, exogenous laminin appeared to act by preventing the accumulation of a basement membrane and by disrupting the organization of the outer layer into an epithelium. No such effects were produced by fibronectin or collagens type I or IV. Aggregates cultured in the presence of antilaminin also failed to organize an epithelium and did not produce AFP, whereas those in normal rabbit serum differentiated normally. Therefore, endogenous laminin plays a key role not only as a basement membrane structural component but also in organizing the epithelial layer of endoderm cells and hence (indirectly) in gene expression.

Animals↗
PubMed1995

Interaction of LHRH with growth hormone-releasing factor-dependent and -independent postnatal development of somatotrophs in rat pituitary cell aggregates.

Addition of LHRH for 40 h to aggregate cell cultures of 14-day-old rat pituitary significantly decreased the number of [3H]thymidine ([3H]T)-incorporating cells which immunostained for GH protein as well as the number of [3H]T-labelled cells expressing GH mRNA detectable by in situ hybridization with a digoxigenin-labelled riboprobe. The effect at the level of GH protein was seen at a dose of 1 nM LHRH. However, the effect at the GH mRNA level required a higher dose of LHRH (10 nM) or a longer incubation time (64 h). Treatment of the cultures for 40 h with 0.1 nM GH-releasing factor (GRF) provoked a 54% increase in the number of [3H]T-labelled cells containing GH mRNA and a 30% increase in the number of cells immunostained for GH protein. The latter effects of GRF were completely blocked by simultaneous addition of LHRH (1 nM) to the cultures. In the absence of GRF, LHRH (1 nM) also had an inhibitory effect on the total number of cells containing GH mRNA and a comparable effect on the total number of cells stained for GH protein. The present data show that LHRH is capable of inhibiting the GRF-independent as well as the GRF-dependent development of somatotrophs in postnatal rat pituitary in culture.

Animals↗
PubMed2000

Relative abundance of different cadherins defines differentiation of mesenchymal precursors into osteogenic, myogenic, or adipogenic pathways.

Cadherins, a family of cell-cell adhesion molecules, provide recognition signals that are important for cell sorting and aggregation during tissue development. This study was performed to determine whether distinct cadherin repertoires define tissue-specific lineages during differentiation of immature C3H10T1/2 and C2C12 mesenchymal cells. Both cell lines expressed mRNA for N-cadherin (N-cad), cadherin-11 (C11), and R-cadherin (R-cad). After induction of osteogenesis by recombinant human BMP-2 (rhBMP-2) treatment, steady state N-cad mRNA slightly increased in C3H10T1/2 cells. Likewise, the abundance of C11 mRNA increased in both cell lines, although the changes were more remarkable in C2C12 cells. By contrast, R-cad expression was almost shut off by rhBMP-2. The immature but committed osteoblastic MC3T3-E1 cells exhibited only minor changes in N-cad and C11 mRNA abundance after rhBMP-2 treatment. Whereas adipogenic differentiation was associated with a net decrease of N-cad and C11 expression in C3H10T1/2 cells, induction of myogenesis in C2C12 cells resulted in up-regulation of N-cad, while R-cad mRNA became undetectable in either case. Similarly, the adipocytic 3T3-L1 cells expressed very low levels of all cadherins when fully differentiated. Therefore, the repertoire of cadherins present in undifferentiated mesenchymal cells undergoes distinct changes during transition to mature cell phenotypes. Although neither N-cad nor C11 represent strict tissue-specific markers, the relative abundance of these mesenchymal cadherins defines lineage-specific signatures, perhaps providing recognition signals for aggregation and differentiation of committed precursors.

3T3 Cells↗
PubMed1993

Characterization of a third ras gene, rasB, that is expressed throughout the growth and development of Dictyostelium discoideum.

Previous reports have indicated that the cellular slime mold Dictyostelium discoideum possesses two ras genes (rasG and rasD) and one rap gene (rap1). All three genes are developmentally regulated, with each showing a different pattern of transcription during the Dictyostelium life cycle. To establish whether there are additional ras or rap genes in Dictyostelium, we used degenerate oligonucleotide primers to the highly conserved GTP-binding domains and both ras- and rap-unique sequences to amplify products from cDNA using the polymerase chain reaction (PCR). No additional rap genes were amplified, but a fragment whose nucleotide sequence predicted a novel ras gene was isolated. Using this PCR product as a probe, a full-length cDNA clone was isolated and sequenced. Its deduced amino acid sequence predicted a 197 amino acid protein that is 71% and 68% identical to RasG and RasD respectively. The new ras gene contains the conserved Ras-specific effector domain, the conserved binding site for the Ras-specific Y13-259 monoclonal antibody, and shows greater sequence similarity to the human H-Ras protein than to any other mammalian Ras protein. In view of this high level of identity to the ras gene subfamily, we have designated this gene rasB. Northern blot analysis has shown that rasB is developmentally regulated with maximum levels of a single 950-bp message detected during vegetative growth and the first 8 h of development.

Amino Acid Sequence↗
PubMed2003

A Balbiani body and the fusome mediate mitochondrial inheritance during Drosophila oogenesis.

Maternally inherited mitochondria and other cytoplasmic organelles play essential roles supporting the development of early embryos and their germ cells. Using methods that resolve individual organelles, we studied the origin of oocyte and germ plasm-associated mitochondria during Drosophila oogenesis. Mitochondria partition equally on the spindle during germline stem cell and cystocyte divisions. Subsequently, a fraction of cyst mitochondria and Golgi vesicles associates with the fusome, moves through the ring canals, and enters the oocyte in a large mass that resembles the Balbiani bodies of Xenopus, humans and diverse other species. Some mRNAs, including oskar RNA, specifically associate with the oocyte fusome and a region of the Balbiani body prior to becoming localized. Balbiani body development requires an intact fusome and microtubule cytoskeleton as it is blocked by mutations in hu-li tai shao, while egalitarian mutant follicles accumulate a large mitochondrial aggregate in all 16 cyst cells. Initially, the Balbiani body supplies virtually all the mitochondria of the oocyte, including those used to form germ plasm, because the oocyte ring canals specifically block inward mitochondrial transport until the time of nurse cell dumping. Our findings reveal new similarities between oogenesis in Drosophila and vertebrates, and support our hypothesis that developing oocytes contain specific mechanisms to ensure that germ plasm is endowed with highly functional organelles.

Animals↗
PubMed1998

Repressive effect on oligonucleosome transcription of the core histone tail domains.

Histone-DNA templates for bacteriophage T7 RNA polymerase were assembled from histone octamers and three different DNA species, two circular (pGEMEX-1 and pT207-18) and one linear (T7-207-18). pGEMEX is devoid of nucleosome positioning sequences, while in pT207-18 and T7-207-18 the region downstream of the promoter contains 18 tandem repeats of a 207 bp positioning sequence derived from the 5S RNA gene of the sea urchin Lytechinus variegatus. Elimination of the histone tails in the assembled oligonucleosomes by trypsin digestion is accompanied, in all three DNA species, by substantial increases in transcription efficiency, assayed at different KCl and MgCl2 concentrations, after allowing for the aggregation observed under certain conditions. In the absence of KCl and at low MgCl2 concentration, the presence of 2 mM spermidine causes substantial aggregation of the intact oligonucleosomes but has a much smaller effect on those trypsin digested. The untreated histone-DNA templates, assembled on pGEMEX-1 and T7-207-18, give transcription products significantly shorter than those obtained with the corresponding free DNA. With oligonucleosome templates lacking histone tails, the transcripts have an average length intermediate between those corresponding to free DNA and intact histone-DNA, which indicates a partial elimination of the elongation restrictions imposed by intact histone octamers. The absence of histone terminal domains facilitates both transcriptional initiation and elongation. Apparently, the interaction of the histone tails with DNA at the nucleosomal level is responsible, at least in part, for their repressive effect on transcription.

Bacteriophage T7↗
PubMed1999

Polymorphism of turnip yellow mosaic virus empty shells and evidence for conformational changes occurring after release of the viral RNA. A differential scanning calorimetric study.

Turnip yellow mosaic virus (TYMV) is a small isometric plant virus which decapsidates by releasing its RNA through a hole in the capsid, leaving behind an empty shell [R. E. F. Matthews and J. Witz, (1985) Virology 144, 318-327]. Similar empty shells (artificial top component, ATC) can be obtained by submitting the virions to various treatments in vitro. We have used differential scanning calorimetry, analytical sedimentation, and electron microscopy to investigate the thermodenaturation of natural empty shells (NTC, natural top component) present in purified virus suspensions, and of several types of ATCs. ATCs divided in two major classes. Those obtained by alkaline titration, by the action of urea or butanol behaved as NTC: their thermograms contained only one peak corresponding to the irreversible dissociation of the shells and the denaturation of the coat protein. The temperature of this unique transition varied significantly with pH, from 71 degrees C at pH 4.5 to 84 degrees C at pH 8.5. The thermograms of ATCs obtained by freezing and thawing, or by the action of high pressure, contained two peaks: shells dissociated first into smaller protein aggregates at 57 degrees C (at pH 5.0) to 61 degrees C (at pH 8.5), which denatured at the temperature of the unique transition of NTC. Shells obtained by heating virions to 55 degrees C at pH 7.6, changed conformation after the release of the viral RNA, as upon continuous heating to 95 degrees C, their thermograms were similar to those of the shells obtained by freezing and thawing, whereas after purification they behaved like NTC. Structural implications of these observations are discussed.

Calorimetry, Differential Scanning↗
PubMed1993

Platelet activation releases megakaryocyte-synthesized apolipoprotein J, a highly abundant protein in atheromatous lesions.

Apolipoprotein J (apoJ) is an abundant glycoprotein in many biological fluids, and its constitutive high level synthesis is characteristic of many epithelial cells exposed to harsh fluids such as urine, bile, and gastric secretions. In addition, dramatic induction of apoJ occurs in cells surrounding several kinds of pathological lesions. Because platelets and circulating inflammatory cells represent critical elements in numerous pathological processes, we evaluated bone marrow cells for the presence of apoJ. Based upon messenger RNA in situ hybridization and immunofluorescent protein detection, high-level apoJ gene expression and protein accumulation occurred exclusively in mature megakaryocytes. Our results indicate that apoJ is stored in platelet granules and is released into extracellular fluid following platelet activation. Because atheromatous plaque development involves platelet aggregation and activation, we looked for and found abundant apoJ protein in advanced human atheromatous lesions. Thus, platelet sequestration and activation may lead to the rapid deployment of apoJ into sites of vascular injury. We hypothesize that platelet-derived apoJ participates in both short-term wound repair processes and chronic pathogenic processes at vascular interfaces.

Animals↗
PubMed2000

A pAD1-encoded small RNA molecule, mD, negatively regulates Enterococcus faecalis pheromone response by enhancing transcription termination.

pAD1 is a 60-kb hemolysin-bacteriocin plasmid in Enterococcus faecalis that encodes a conjugative mating response to a peptide sex pheromone, cAD1, secreted by plasmid-free bacteria. The pheromone response is regulated by two proteins: TraE1, which positively regulates all or most conjugative structural genes, and TraA, which negatively regulates traE1. TraA binds to pAD1 DNA at the iad (encoding the inhibitor peptide iAD1) promoter but is released upon binding to imported pheromone. This leads to enhanced transcription through two closely spaced downstream terminators (t1 and t2) into traE1. TraE1 is believed to then upregulate itself from a site located within t2; thus, a small amount of transcription through t1-t2 could lead to overall induction. It is important therefore that the t1-t2 terminators be tightly controlled to keep the response shut down in the absence of pheromone. A small (200-nucleotide) RNA molecule designated mD is encoded just upstream of t1 by a determinant (traD) oriented in the direction opposite to that of transcripts utilizing t1. mD is expressed at high levels in the uninduced state, but it decreases significantly upon induction. Here we present results of genetic studies relating to the activity of t1-t2 and show that mD strongly enhances transcriptional termination at t1. The mD activity is shown to influence transcription well downstream and can affect the determinant for aggregation substance asa1. The phenomenon is specific in that there is no effect of mD on the unrelated pheromone-responding plasmids pPD1 and pCF10.

Bacterial Proteins↗
PubMed1993

RNA-stimulated NTPase activity associated with yellow fever virus NS3 protein expressed in bacteria.

The nonstructural protein NS3 of the prototypic flavivirus, yellow fever virus, was investigated for possession of an NTPase activity. The entire NS3 protein coding sequence and an amino-terminal truncated version thereof were engineered into Escherichia coli expression plasmids. Bacteria harboring these plasmids produced the expected polypeptides, which upon cell disruption were found in an insoluble aggregated material considerably enriched for the NS3-related polypeptides. Solubilization and renaturation of these materials, followed by examination of their ability to hydrolyze ATP, revealed an ATPase activity present in both the full-length and amino-terminal truncated NS3 preparations but not in a similarly prepared fraction from E. coli cells engineered to express an unrelated polypeptide. The amino-terminal truncated NS3 polypeptide was further enriched to greater than 95% purity by ion-exchange and affinity chromatography. Throughout the purification scheme, the ATPase activity cochromatographed with the recombinant NS3 polypeptide. The enzymatic activity of the purified material was shown to be a general NTPase and was dramatically stimulated by the presence of particular single-stranded polyribonucleotides. These results are discussed in view of similar activities identified for proteins of other positive-strand RNA viruses.

Adenosine Triphosphatases↗
PubMed1990

Isolation and characterization of spore coat protein (sp96) gene of Dictyostelium discoideum.

A cDNA library was constructed from poly(A)+ RNA isolated from slug cells of Dictyostelium discoideum, using lambda gt11 phage, and screened with an antiserum specific for the spore coat protein sp96. A positive clone was obtained and the gene product was identified as sp96. The sp96 mRNA is 2.2 kb in size, and it starts to accumulate at the tipped aggregate stage only in prespore cells. Southern analysis using nuclear DNA established that the sp96 gene is unique. Two genomic clones containing the sp96 gene were isolated and the sequence of the gene established. The coding region contains a long open reading frame interrupted by a single intron.

Amino Acid Sequence↗
PubMed2002

Persistent and transient replication of full-length hepatitis C virus genomes in cell culture.

The recently developed subgenomic hepatitis C virus (HCV) replicons were limited by the fact that the sequence encoding the structural proteins was missing. Therefore, important information about a possible influence of these proteins on replication and pathogenesis and about the mechanism of virus formation could not be obtained. Taking advantage of three cell culture-adaptive mutations that enhance RNA replication synergistically, we generated selectable full-length HCV genomes that amplify to high levels in the human hepatoma cell line Huh-7 and can be stably propagated for more than 6 months. The structural proteins are efficiently expressed, with the viral glycoproteins E1 and E2 forming heterodimers which are stable under nondenaturing conditions. No disulfide-linked glycoprotein aggregates were observed, suggesting that the envelope proteins fold productively. Electron microscopy studies indicate that cell lines harboring these full-length HCV RNAs contain lipid droplets. The majority of the core protein was found on the surfaces of these structures, whereas the glycoproteins appear to localize to the endoplasmic reticulum and cis-Golgi compartments. In agreement with this distribution, no endoglycosidase H-resistant forms of these proteins were detectable. In a search for the production of viral particles, we noticed that these cells release substantial amounts of nuclease-resistant HCV RNA-containing structures with a buoyant density of 1.04 to 1.1 g/ml in iodixanol gradients. The same observation was made in transient-replication assays using an authentic highly adapted full-length HCV genome that lacks heterologous sequences. However, the fact that comparable amounts of such RNA-containing structures were found in the supernatant of cells carrying subgenomic replicons demonstrates a nonspecific release independent of the presence of the structural proteins. These results suggest that Huh-7 cells lack host cell factors that are important for virus particle assembly and/or release.

Culture Media, Conditioned↗
PubMed1993

An immunohistologic study of granulomatous inflammation in SIV-infected rhesus monkeys.

We studied granulomatous inflammation in simian AIDS using histologic, immunohistologic, and in situ hybridization techniques. Complete Freund's adjuvant was used to induce granulomas in two control animals and two macaques infected with simian immunodeficiency virus (SIV) and having low peripheral CD4+ T cell counts. Control animals developed large (> 2 cm diameter) epithelioid granulomas containing CD68+ macrophages (m phi s), epithelioid m phi s and multinucleated giant cells (MNGCs), CD4+ and CD8+ T cells, and small perivascular collections of CD20+ B cells. Lymphocytes rarely expressed proliferating cell nuclear antigen (Ki-67), and only rare endothelial cells expressed vascular cell adhesion molecule 1 (VCAM-1). In contrast, SIV+ animals had smaller (< 0.5 cm diameter) epithelioid granulomas characterized by numerous large, dense CD8+, CD20+ lymphocyte aggregates with prominent local division (Ki-67+). Despite low blood CD4+ T cell numbers, there was a substantial CD4+ T cell infiltrate, accompanied by enhanced endothelial VCAM-1 expression. These granulomas contained no detectable SIV antigen or RNA. Thus, in simian AIDS, experimentally induced granulomatous responses are grossly attenuated, yet associated with increased local endothelial-leukocyte signaling and lymphocyte division.

Animals↗
PubMed1974

Differences in membrane fluidity and structure in contact-inhibited and transformed cells.

Studies on contact-inhibited mouse embryo fibroblast 3T3 cells and 3T3 cells transformed by oncogenic RNA and DNA viruses and by a chemical carcinogen have demonstrated differences in plasma membrane architecture. Spin-label and freeze-fracture ultrastructural studies have shown that contact-inhibited cells have ordered membrane lipids and aggregated intramembranous particles, whereas transformed cells have fluid membrane lipids and randomly distributed intramembranous particles. These findings suggest a model for how changes in the cell membrane may account for some of the characteristic differences observed between contact-inhibited and transformed cells.

Animals↗
PubMed2001

Accumulation of mutant huntingtin fragments in aggresome-like inclusion bodies as a result of insufficient protein degradation.

The huntingtin exon 1 proteins with a polyglutamine repeat in the pathological range (51 or 83 glutamines), but not with a polyglutamine tract in the normal range (20 glutamines), form aggresome-like perinuclear inclusions in human 293 Tet-Off cells. These structures contain aggregated, ubiquitinated huntingtin exon 1 protein with a characteristic fibrillar morphology. Inclusion bodies with truncated huntingtin protein are formed at centrosomes and are surrounded by vimentin filaments. Inhibition of proteasome activity resulted in a twofold increase in the amount of ubiquitinated, SDS-resistant aggregates, indicating that inclusion bodies accumulate when the capacity of the ubiquitin-proteasome system to degrade aggregation-prone huntingtin protein is exhausted. Immunofluorescence and electron microscopy with immunogold labeling revealed that the 20S, 19S, and 11S subunits of the 26S proteasome, the molecular chaperones BiP/GRP78, Hsp70, and Hsp40, as well as the RNA-binding protein TIA-1, the potential chaperone 14-3-3, and alpha-synuclein colocalize with the perinuclear inclusions. In 293 Tet-Off cells, inclusion body formation also resulted in cell toxicity and dramatic ultrastructural changes such as indentations and disruption of the nuclear envelope. Concentration of mitochondria around the inclusions and cytoplasmic vacuolation were also observed. Together these findings support the hypothesis that the ATP-dependent ubiquitin-proteasome system is a potential target for therapeutic interventions in glutamine repeat disorders.

14-3-3 Proteins↗
PubMed2004

A plant type 2 metallothionein (MT) from cork tissue responds to oxidative stress.

Expression of plant metallothionein genes has been reported in a variety of senescing tissues, such as leaves and stems, ripening fruits, and wounded tissues, and has been proposed to function in both metal chaperoning and scavenging of reactive oxygen species. In this work, it is shown that MT is also associated with suberization, after identifying a gene actively transcribed in Quercus suber cork cells as a novel MT. This cDNA, isolated from a phellem cDNA library, encodes a MT that belongs to type 2 plant MTs (QsMT). Expression of the QsMT cDNA in E. coli grown in media supplemented with Zn, Cd, or Cu has yielded recombinant QsMT. Characterization of the respective metal aggregates agrees well with a copper-related biological role, consistent with the capacity of QsMT to restore copper tolerance to a MT-deficient, copper-sensitive yeast mutant. Furthermore, in situ hybridization results demonstrate that RNA expression of QsMT is mainly observed under conditions related to oxidative stress, either endogenous, as found in cork or in actively proliferating tissues, or exogenous, for example, in response to H(2)O(2) or paraquat treatments. The putative role of QsMT in oxidative stress, both as a free radical scavenger via its sulphydryl groups or as a copper chelator is discussed.

Amino Acid Sequence↗