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 775 records · Page 43Linked to original sources
PubMed1998

Specific inclusion bodies are associated with replication of lettuce infectious yellows virus RNAs in Nicotiana benthamiana protoplasts.

Nicotiana benthamiana mesophyll protoplasts, either mock-inoculated or inoculated using in vitro transcripts derived from lettuce infectious yellows virus (LIYV) RNA 1- and/or RNA 2-cloned cDNAs were analysed by transmission electron microscopy (TEM) and, in some cases, also by immunogold labelling. TEM revealed the main cytopathological effects of LIYV infections in N. benthamiana protoplasts infected with RNAs 1 and 2: (a) typical closterovirus-induced (beet yellows virus-type) accumulations of vesiculated cytoplasmic membranes as inclusion bodies, sometimes with associated virions; (b) scattered aggregations of virions within the cytoplasm; and (c) electron-dense plasmalemma deposits. These were not seen in mock-inoculated protoplasts. Protoplasts inoculated only with LIYV RNA 1 contained vesiculated cytoplasmic inclusion bodies, but not virions or plasmalemma deposits. Thus, infection by only LIYV RNA 1 is sufficient to induce characteristic closterovirus vesiculated cytoplasmic inclusion bodies. However, both LIYV RNAs 1 and 2 are needed for production of virions and plasmalemma deposits.

Closterovirus↗
PubMed1984

Structural characterization of virion proteins and genomic RNA of human parainfluenza virus 3.

The virion proteins and genomic RNA of human parainfluenza virus 3 have been characterized. The virion contains seven major and two minor proteins. Three proteins of 195 X 10(3) molecular weight (195K), 87K, and 67K are associated with the nucleocapsid of the virion and have been designated L, P, and NP, respectively. Three proteins can be labeled with [14C]glucosamine and have molecular weights of 69K, 60K, and 46K. We have designated these proteins as HN, F0, and F1, respectively. HN protein has interchain disulfide bonds, but does not participate in disulfide bonding to form homomultimeric forms. F1 appears to be derived from a complex, F1,2, that has an electrophoretic mobility similar to that of F0 under nonreducing conditions. A protein of 35K is associated with the envelope components of the virion and aggregates under low-salt conditions; this protein has been designated M. The genome of human parainfluenza virus 3 is a linear RNA molecule with a molecular weight of approximately 4.6 X 10(6).

Capsid↗
PubMed1994

Cloning and biological function of laminin in Hydra vulgaris.

The Cnidarian, hydra, lends itself to studies related to the role of extracellular matrix (ECM) components in development because of its high regenerative capacity and its simple structure, which is organized as an epithelial bilayer with an intervening ECM termed the mesoglea. Previous immunocytochemical and biochemical studies have established that hydra mesoglea contains many of the major matrix components (e.g., fibronectin, laminin, type IV collagen, and heparan sulfate proteoglycan) associated with the ECM of vertebrate and more complex invertebrate species. Additional studies have also established that ECM components have a critical role in hydra development as monitored during head regeneration and morphogenesis of hydra cell aggregates. In the present study a monoclonal antibody (mAb52) raised to isolated hydra mesoglea was used as a probe in additional functional studies and to screen a cDNA expression library made from poly(A)+ RNA isolated from Hydra vulgaris. Immunofluorescent analysis indicated that mAb52 was localized along the entire longitudinal axis of adult polyps in what is termed the subepithelial zones of hydra mesoglea. Cytochemical studies found these subepithelial zones to be rich in anionic sites. Previous studies have shown that mAb52 blocks hydra cell aggregate development and experiments in the current study have shown that mAb52 also blocks in vivo interstitial cell (I-cell) migration in hydra grafts. Sequence analysis of cDNA clones isolated using mAb52 indicated that the protein encoded by these clones had structural homology with mammalian and Drosophila laminin B1 chain and hybridized to a single 6.75-kb band on Northern blots of total hydra RNA. One interesting difference in hydra laminin B1 was the presence of a FTGTQ amino acid sequence in place of the vertebrate YIGSR cell binding domain. Under nonreducing conditions, polyclonal antibodies against FTGTQ bound to the same > 200-kDa band on Western blots of mesoglea as mAb52 and also immunolocalized to the subepithelial zones. Under reducing conditions, anti-FTGTQ antibodies bound to a single band with a mass of approximately 200 kDa. In addition, FTGTQ peptide inhibited adhesion of dissociated hydra cells to mesoglea and anti-FTGTQ antibodies inhibited hydra cell binding to substrates coated with mesoglea or FTGTQ peptide. Anti-FTGTQ antibodies also inhibited in vivo I-cell migration in hydra grafts. Given the early divergence of Cnidarians during evolution, these studies indicate the highly conserved nature of laminin and provide additional information regarding the critical role of ECM components during hydra development.

Amino Acid Sequence↗
PubMed1992

Characterization of Escherichia coli RNase PH.

We have previously shown that the orfE gene of Escherichia coli encodes RNase PH. Here we show that the OrfE protein (purified as described in the accompanying paper) (Jensen, K. F., Andersen, J. T., and Poulsen, P. (1992) J. Biol. Chem. 267, 17147-17152) has both the degradative and synthetic activities of RNase PH. This highly purified protein was used to characterize the enzymatic and structural properties of RNase PH. The enzyme requires a divalent cation and phosphate for activity, the latter property indicating that RNase PH is exclusively a phosphorolytic enzyme. Among tRNA-type substrates, the enzyme is most active against synthetic tRNA precursors containing extra residues following the -CCA sequence, and it can act on these molecules to generate mature tRNA with amino acid acceptor activity; 3'-phosphoryl-terminated molecules are not active as substrates. The equilibrium constant for RNase PH is near unity, suggesting that at the phosphate concentration present in vivo, the enzyme would participate in RNA degradation. The synthetic reaction of RNase PH displays a nonlinear response to increasing enzyme concentrations, and this may be due to self-aggregation of the protein. Higher order multimers of RNase PH could be detected by gel filtration at higher protein concentrations and by protein cross-linking. The possible role of RNase PH in tRNA processing is discussed.

Base Sequence↗
PubMed2000

Confocal raman microspectroscopy and imaging study of theraphthal in living cancer cells.

Binary systems combining a transition metal complex and ascorbate have been proposed recently for catalytic therapy of malignant tumors. The killing effect on tumor cells is achieved by production of free radicals in the course of accelerated oxidation of ascorbate by dioxygen in the presence of transition metal complexes. Further progress in the development of binary catalytic systems (BCSs) requires a special method for their investigation in cells and tissues, because neither component of BCSs fluoresces. Here a resonance Raman confocal spectral imaging (RR CSI) technique was introduced as a unique approach to monitor quantitatively the transition metal complexes within living cells. Intracellular accumulation, localization, and retention of theraphthal (TP), a catalyst of the advanced TP/ascorbate BCS, were investigated in A549 cells with the RR CSI technique. The cellular analysis was complemented with the detailed study of molecular interactions of TP in solution and environmental factors affecting the RR spectrum of TP. TP does not penetrate into membranes, it binds very weakly to DNA and RNA, but it readily forms complexes with proteins. Binding with Ca(2+) cations and decreasing pH below 6 induce aggregation of TP. By analyzing RR spectra recorded from every point within a TP-treated cell, three states of the agent were discriminated, namely, monomeric TP in polar environment, TP bound to proteins, and aggregated TP. Their cytoplasmic and nuclear distributions were mapped at different stages of uptake and efflux. By introducing organelle-selective fluorescent probes into drug-treated cells and measuring intracellular localization of both the probe and the drug, compartmentation of TP was revealed. Cell growth suppression by the TP/ascorbate system was measured, and probable molecular and organelle targets of radical damage were characterized.

Adenocarcinoma↗
PubMed1989

An outbreak of gastroenteritis associated with acute rotaviral infection in schoolchildren.

In April 1988 a large outbreak of group C rotavirus infection associated with acute gastroenteritis occurred among schoolchildren and their teachers simultaneously at seven elementary schools in Fukui city, Japan. Of 3,102, 675 (21.8%) became ill. Clinical symptoms were mild, predominantly abdominal pain and vomiting, with diarrhea reported in only 27.6%. The outbreak subsided within 2 d. No pathogenic bacteria were found in fecal specimens; the virus particles detected by electron microscopy were morphologically indistinguishable from conventional infantile rotavirus. Immune electron microscopy showed that these virions formed large aggregates with convalescent serum and with the reference serum specific to group C rotavirus. Polyacrylamide gel electrophoresis showed similar RNA patterns for virus from this outbreak and typical group C rotavirus.

Acute Disease↗
PubMed1989

Structure and function of conjugative pili: inducible synthesis of functional F pili by Escherichia coli K-12 containing a lac-tra operon fusion.

In vivo and in vitro recombination methods were used to construct the recombinant plasmid pTG801, in which the F-plasmid DNA transfer (tra) genes required for the formation of functional F pili were placed under the lac transcriptional control sequences of pUC19. The 20 kilobases of cloned F DNA includes genes traA through the 5'-terminal part of traG; the plasmid lacks the positive regulatory gene traJ and all of the known tra genes required for the DNA transfer stage of conjugation. pTG801 transformants were sensitive to the donor-specific bacteriophages Q beta and f1, as measured by the formation of infectious centers. They were relatively insensitive to bacteriophage R17, as expected from the absence of traD. In the presence of a lacIq allele, sensitivity of pTG801 transformants to f1 and Q beta depended on the concentration of inducer (isopropyl-beta-D-thiogalactopyranoside [IPTG]). Viewed by electron microscopy, pTG801 transformants elaborated 7- to 10-nm-diameter filaments that could be laterally decorated with RNA bacteriophage particles, consistent with the formation of F pili. In stationary-phase cultures, these filaments formed massive aggregates and could be seen to adhere lengthwise to the cell surface; few pili accumulated in the medium as single filaments.

Cloning, Molecular↗
PubMed2005

New perspectives on rickettsial evolution from new genome sequences of rickettsia, particularly R. canadensis, and Orientia tsutsugamushi.

The complete genome sequences available for eight species of Rickettsia and information for other near relatives in the Rickettsiales including Orientia and species of Anaplasmataceae are a rich resource for comparative analyses of the evolution of these obligate intracellular bacteria. Differences in these organisms have permitted them to colonize varied intracellular compartments, arthropod vectors, and vertebrate reservoirs in both pathogenic and symbiotic relationships. We summarize some comparative aspects of the genomes of these organisms, paying particular attention to the recently completed sequence for R. canadensis McKiel strain and an estimated two-thirds of the genome sequence for a Thailand patient isolate of Orientia tsutsugamushi. The Rickettsia genomes exhibit a high degree of synteny punctuated by distinctive chromosome inversions and consistent phylogenetic relationships regardless of whether protein coding sequences or RNA genes, concatenated open reading frames or gene regions, or whole genomes are used to construct phylogenetic trees. The aggregate characteristics (number, length, composition, repeat identity) of tandem repeat sequences of Rickettsia, which often exhibit recent and rapid divergence between closely related strains and species of bacteria, are also very conserved in Rickettsia but differed significantly in Orientia. O. tsutsugamushi shared no significant synteny to species of Rickettsia or Anaplasmataceae, supporting its placement in a unique genus. Like Rickettsia felis, Orientia has many transposases and ankyrin and tetratricopeptide repeat domains. Orientia shares the important ATP/ADP translocase and proline-betaine transporter multigene families with Rickettsia, but has more gene families that may be involved in regulatory and transporter responses to environmental stimuli.

Animals↗
PubMed1998

The RNP protein, RNPS1, associates with specific isoforms of the p34cdc2-related PITSLRE protein kinase in vivo.

The PITSLRE protein kinases are members of the p34cdc2 superfamily, with >20 different isoforms expressed from two linked genes in humans. PITSLRE homologues have been identified in mouse, chicken, Drosophila, Xenopus, and possibly Plasmodium falciparum, suggesting that their function may be well conserved. A possible role for a caspase processed PITSLRE isoform has been suggested by studies of Fas- and TNF-induced cell death. However, the function of these kinases in proliferating cells is still unknown. Here we demonstrate that the 110 kDa PITSLRE isoforms (p110) are localized to both the nucleoplasm and nuclear speckles, and that these isoforms specifically interact in vitro and in vivo with the RNA-binding protein RNPS1. RNPS1 is also localized to nuclear speckles, and its over expression disrupts normal nuclear speckle organization by causing the aggregation of many nuclear speckles into approximately 6 'mega' speckles. This type of nuclear speckle aggregation closely resembles what occurs when cells are treated with several transcriptional inhibitors. These data indicate that the PITSLRE p110 isoforms interact with RNPS1 in vivo, and that these proteins may in turn influence some aspect of transcriptional and/or splicing regulation.

Animals↗
PubMed1995

In vitro transcription/translation assay for the screening of hMLH1 and hMSH2 mutations in familial colon cancer.

BACKGROUND & AIMS: Hereditary nonpolyposis colorectal cancer (HNPCC) has been linked recently to a defect in repairing mismatched nucleotides in DNA. The aim of this study was to screen for germline mutations that result in prematurely truncated proteins in two of the mismatch repair genes identified at this time, hMLH1 and hMSH2, in a consecutive series of patients belonging to familial aggregations of colorectal cancer. METHODS: Nineteen individuals with colorectal cancer from 19 families were consecutively referred because of a strong positive family history of colorectal cancer. Premature truncation mutations in hMLH1 and hMSH2 were sought from lymphocyte RNA by using an in vitro transcription/translation (IVTT) assay. RESULTS: Protein truncating mutations in the hMLH1 or hMSH2 genes were found in 50% of families with HNPCC (6 of 12) but were not observed in any of the remaining familial aggregations that did not fulfill the standard criteria for HNPCC. In some of the IVTT-positive samples, the mutations were characterized by genomic sequencing. CONCLUSIONS: IVTT may be a practical method to accomplish primary screening of germline mutations in DNA mismatch pair genes in HNPCC; however, a broader approach is necessary to obtain a more complete picture of the mutational spectrum in HNPCC and other familial aggregations of colorectal cancer.

Base Sequence↗
PubMed2002

Expression of the active human and duck hepatitis B virus polymerases in heterologous system of Pichia methanolica.

We expressed the Hepatitis B virus polymerase (HBV P protein) using a recently introduced yeast system, Pichia methanolica. HBV (1-680 amino acids) and Duck Hepatitis B virus (DHBV, 1-780 amino acids) polymerase were expressed and showed DNA dependent DNA polymerase (DDDP). The DHBV polymerase had RNA dependent DNA polymerase (RDDP) and RNase H activities. We present a new simplified way of obtaining active viral P protein using the yeast expression system. The viral P proteins proved to be stable and were not aggregated in the yeast system.

Animals↗
PubMed1986

[Isolation and fractionation of the nuclear matrix from cultured cells].

A new method for the isolation of tissue culture cell nuclei is presented which involves incubation of the nuclei in the presence of Cu2+- or Zn2+-ions. This method eliminates the danger of nuclear aggregation and permits nuclear matrix isolation and subsequent fractionation. Stabilization of the inner matrix by Cu2-ions permits analysis of the role of nucleic acids in the maintenance of the matrix structure. It is shown that solubilization of more than 95% of matrix-bound DNA and more than 90% of matrix-bound RNA did not cause any significant changes in the nuclear matrix structure.

Animals↗
PubMed1981

The quaternary structure of DNA-dependent RNA polymerase.

The crystals of RNA polymerase of T. thermophilus were examined by electron microscopic observation of the negatively stained and sectioned materials. Three types of crystals were observed: ordered aggregates (Type I), cylindrical duplei (Type II), and plane (Type III) forms. It was deduced mainly from sectioned images that Type I crystal is a precursor or a premature form of Type II crystal. The III corresponds to the flattened layer of type II. In Type II crystal the enzyme molecules are arranged in an orderly two-dimensional lattice and thus we could analyze the molecular structure by optical filtering of the negatively stained images. On the basis of these results, a quaternary structure is proposed for RNA polymerase.

Bacteria↗
PubMed2000

A critical role for Xdazl, a germ plasm-localized RNA, in the differentiation of primordial germ cells in Xenopus.

Xdazl is an RNA component of Xenopus germ plasm and encodes an RNA-binding protein that can act as a functional homologue of Drosophila boule. boule is required for entry into meiotic cell division during fly spermatogenesis. Both Xdazl and boule are related to the human DAZ and DAZL, and murine Dazl genes, which are also involved in gamete differentiation. As suggested from its germ plasm localization, we show here that Xdazl is critically involved in PGC development in Xenopus. Xdazl protein is expressed in the cytoplasm, specifically in the germ plasm, from blastula to early tailbud stages. Specific depletion of maternal Xdazl RNA results in tadpoles lacking, or severely deficient in, primordial germ cells (PGCs). In the absence of Xdazl, PGCs do not successfully migrate from the ventral to the dorsal endoderm and do not reach the dorsal mesentery. Germ plasm aggregation and intracellular movements are normal indicating that the defect occurs after PGC formation. We propose that Xdazl is required for early PGC differentiation and is indirectly necessary for the migration of PGCs through the endoderm. As an RNA-binding protein, Xdazl may regulate translation or expression of factors that mediate migration of PGCs.

Animals↗
PubMed1975

RNA-directed DNA polymerase from human leukemic blood cells and from primate type-C virus-producing cells: high- and low-molecular-weight forms with variant biochemical and immunological properties.

RNA-directed DNA polymerase (reverse transcriptase) from leukocytes of individual leukemic patients can be grouped by velocity gradient analyses into two distinct classes, a low-molecular-weight (LMW) class of approximately 70,000 and a high-molecular-weight (HMW) class of 130,000 to 140,000. The reverse transcriptases from mammalian type-C viruses have with one exception (see text) been isolated as enzymes with molecular weights of 70,000. In this study, the reverse transcriptase from extracellular gibbon ape leukemia virus was also isolated only as the LMW class. However, the enzyme from gibbon virus-producing cells was isolated partially in the HMW form; this form was converted completely to the LMW form by treatment with 0.5 M KC1 and 0.5% Triton X-100 and could be re-converted to the HMW form by lowering the KC1 and Triton X-100 concentrations. A similar conversion from a HMW form to a LMW form was demonstrated with enzyme from human leukemic cells. The LMW form of the human and gibbon ape cellular enzymes utilized synthetic primer-templates in a similar fashion to viral enzyme, and this form was strongly inhibited by antisera (IgG) to reverse transcriptase from simian (woolly monkey) type-C virus. The HMW form of both enzymes utilized synthetic primer-templates less efficiently than the LMW form, and was resistant to inhibition by antipolymerase IgG of simian type-C virus. The HMW form of the cellular reverse transcriptases transcribed viral 70S RNA in the absence of synthetic primer relatively more efficiently than did the extracellular viral form. These data suggest that the HMW form is due in part to aggregation of the LMW form and in part to a cellular factor(s) which may affect both the form and function of intracellular reverse transciptase.

Animals↗
PubMed1990

Modified subcellular localization of interferon-induced p68 kinase during encephalomyocarditis virus infection.

The double-stranded (ds) RNA-activated protein kinase from human cells is a 68,000 Mr protein (p68 kinase) induced by interferon. When autophosphorylated, p68 kinase catalyzes the phosphorylation of the protein synthesis eukaryotic initiation factor-2, thus mediating inhibition of protein synthesis. The level of p68 kinase is dramatically reduced in nonionic detergent NP-40 extracts, obtained from interferon-treated cells during infection with encephalomyocarditis virus (EMCV) (A. G. Hovanessian, J. Galabru, E. Meurs, C. Buffet-Janvresse, J. Svab and N. Robert, Virology 159, 126-136, 1987). Here we show that such reduction of p68 kinase is in fact due to its reduced NP-40 solubility occurring during EMCV infection. However, p68 kinase can be recovered by extraction with an ionic detergent. Reduced NP-40 extractibility of p68 kinase is dependent on the multiplicity of virus infection and seems to be specific, since other cellular proteins as well as the 100-kDa 2',5'-oligoadenylate synthetase also induced by interferon are not modified. Immunofluorescence studies using specific antibodies demonstrated that p68 kinase which is distributed evenly in the cytoplasm of HeLa cells becomes concentrated around the nuclei after EMCV infection. As a consequence of aggregating around the nuclei, p68 kinase might then resist extraction by NP-40. The aggregated kinase is found to be already activated probably due to binding to the replicative form and/or to replicative intermediates of EMCV RNA. Through this process, the functioning of p68 kinase might be guaranteed by a localized activation in the replication complexes of EMCV.

2',5'-Oligoadenylate Synthetase↗
PubMed2004

Polyelectrolyte nanoparticles mediate vascular gene delivery.

PURPOSE: The purpose is to develop a non-viral gene delivery system that meets the requirements of colloidal stability of DNA complexes expressed in terms of no particle aggregation under physiologic conditions. The system should be used to transfect cardiovascular tissues. METHODS: We used a strategy based on the formation of polyelectrolyte nanoparticles by deposition of alternatively charged polyelectrolytes onto a DNA core. Polyelectrolytes were transfer RNA as well as the synthetic polyanion, polyvinyl sulfate (PVS), and the polycation polyethylenimine (PEI). The PEI/DNA complex formed the DNA core. RESULTS: We observed that the DNA is condensed by polycations and further packaged by association with a polyanion. These nanoparticles exhibited negative surface charge and low aggregation tendency. In vivo rat carotid artery experiments revealed high transfection efficiency, not only with the reporter gene but also with the gene encoding human urokinase plasminogen activator (Hu-uPA). Hu-uPA is one of the proteins involved in the recovery of the blood vessels after balloon catheter injury and therefore clinically relevant. CONCLUSIONS: A strategy for in vivo gene transfer is proposed that uses the incorporation of polyanions as RNA or PVS into PEI/DNA complexes in order to overcome colloidal instability and to generate a negative surface charge. The particles proved to be transfectionally active in vascular gene transfer.

Animals↗