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microwave-assisted rapid enzymatic synthesis of nucleic acids.herein we report microwave-induced enhancement of the reactions catalyzed by escherichia coli dna polymerase i and avian myeloblastosis virus-reverse transcriptase. the reactions induced by microwaves result in a highly selective synthesis of nucleic acids in 10-50 seconds. in contrast, same reactions failed to give desired reaction products when carried out in the same time periods, but without microwave irradiation. each of the reactions was carried out for different duration of microwave expo ...201627159147
kl-6 regulated the expression of hgf, collagen and myofibroblast differentiation.kl-6 is a pulmonary epithelial-derived mucin which is secreted mainly by type ii alveolar epithelial cells. the level of kl-6 in serum is closely correlated to the clinical activity of various interstitial lung diseases (ild) and acts as a prognostic factor for ild patients. previous studies have showed that kl-6 promoted chemotaxis, proliferation and inhibited the apoptosis of human lung fibroblasts. however, the underlying mechanism remains unknown. in this study, we investigated the function ...201324302189
base-pairing preferences, physicochemical properties and mutational behaviour of the dna lesion 8-nitroguanine.8-nitro-2'-deoxyguanosine (8-nitrodg) is a relatively unstable, mutagenic lesion of dna that is increasingly believed to be associated with tissue inflammation. due to the lability of the glycosidic bond, 8-nitrodg cannot be incorporated into oligodeoxynucleotides (odns) by chemical dna synthesis and thus very little is known about its physicochemical properties and base-pairing preferences. here we describe the synthesis of 8-nitro-2'-o-methylguanosine, a ribonucleoside analogue of this lesion, ...201222965127
viral reverse transcriptases show selective high affinity binding to dna-dna primer-templates that resemble the polypurine tract.previous results using a selex (systematic evolution of ligands by exponential enrichment)-based approach that selected dna primer-template duplexes binding with high affinity to hiv reverse transcriptase (rt) showed that primers mimicking the 3' end, and in particular the six nt terminal g tract, of the rna polypurine tract (ppt; hiv ppt: 5'-aaaagaaaagggggg-3') were preferentially selected. in this report, two viral (moloney murine leukemia virus (mulv) and avian myeloblastosis virus (amv)) and ...201222848574
improving the thermal stability of avian myeloblastosis virus reverse transcriptase α-subunit by site-directed mutagenesis.avian myeloblastosis virus reverse transcriptase (amv rt) is a heterodimer consisting of a 63 kda α-subunit and a 95 kda β subunit. moloney murine leukaemia virus reverse transcriptase (mmlv rt) is a 75 kda monomer. these two rts are the most extensively used for conversion of rna to dna. we previously developed several mutations that increase the thermostability of mmlv rt and generated a highly stable mmlv rt variant e286r/e302k/l435r/d524a by combining three of them (glu286→arg, glu302→lys, a ...201222426840
complete nucleotide sequence of a spanish isolate of alfalfa mosaic virus: evidence for additional genetic variability.alfalfa mosaic virus (amv) is a plant virus that is distributed worldwide and can induce necrosis and/or yellow mosaic on a large variety of plant species, including commercially important crops. it is the only virus of the genus alfamovirus in the family bromoviridae. amv isolates can be clustered into two genetic groups that correlate with their geographic origin. here, we report for the first time the complete nucleotide sequence of a spanish isolate of amv found infecting cape honeysuckle (t ...201121327783
comparison of the thermal stabilities of the αβ heterodimer and the α subunit of avian myeloblastosis virus reverse transcriptase.avian myeloblastosis virus reverse transcriptase (amv rt) is a heterodimer consisting of a 63-kda α subunit and a 95-kda β subunit. in this study, we explored the role of the interaction between the α and β subunits on amv rt stability. the recombinant amv rt α subunit was expressed in insect cells and purified. it exhibited lower thermal stability than the native amv rt αβ heterodimer. unlike the αβ heterodimer, the α subunit was not stabilized by template-primer. these results suggest that int ...201121821920
Detection of Murine Leukemia Virus or Mouse DNA in Commercial RT-PCR Reagents and Human DNAs.The xenotropic murine leukemia virus (MLV)-related viruses (XMRV) have been reported in persons with prostate cancer, chronic fatigue syndrome, and less frequently in blood donors. Polytropic MLVs have also been described in persons with CFS and blood donors. However, many studies have failed to confirm these findings, raising the possibility of contamination as a source of the positive results. One PCR reagent, Platinum Taq polymerase (pol) has been reported to contain mouse DNA that produces f ...201122205995
Status of tobacco viruses in Serbia and molecular characterization of tomato spotted wilt virus isolates. In a four-year survey to determine the presence and distribution of viruses in tobacco crops at 17 localities of the Vojvodina Province and Central Serbia, 380 samples were collected and analyzed by DAS-ELISA. Out of the seven viruses tested, tomato spotted wilt virus (TSWV), potato virus Y (PVY), tobacco mosaic virus (TMV), cucumber mosaic virus (CMV), and alfalfa mosaic virus (AMV) were detected in 37.9, 33.4, 28.7, 23.9, and 15.5% of the total tested samples, respectively. TSWV was the most ...201122149499
mechanism of hiv reverse transcriptase inhibition by zinc: formation of a highly stable enzyme-(primer-template) complex with profoundly diminished catalytic activity.several physiologically relevant cations including ca(2+), mn(2+), and zn(2+) have been shown to inhibit hiv reverse transcriptase (rt), presumably by competitively displacing one or more mg(2+) ions bound to rt. we analyzed the effects of zn(2+) on reverse transcription and compared them to ca(2+) and mn(2+). using nucleotide extension efficiency as a readout, zn(2+) showed significant inhibition of reactions with 2 mm mg(2+), even when present at only ∼5 μm. mn(2+) and ca(2+) were also inhibit ...201121953456
Molecular breeding of transgenic white clover (Trifolium repens L.) with field resistance to Alfalfa mosaic virus through the expression of its coat protein gene.Viral diseases, such as Alfalfa mosaic virus (AMV), cause significant reductions in the productivity and vegetative persistence of white clover plants in the field. Transgenic white clover plants ectopically expressing the viral coat protein gene encoded by the sub-genomic RNA4 of AMV were generated. Lines carrying a single copy of the transgene were analysed at the molecular, biochemical and phenotypic level under glasshouse and field conditions. Field resistance to AMV infection, as well as mi ...201121947755
Rapid detection of wheat yellow mosaic virus by reverse transcription loop-mediated isothermal amplification.ABSTRACT: For the detection of wheat yellow mosaic virus (WYMV), we established a reverse transcription loop-mediated isothermal amplification (RT-LAMP) method. Using Primer Explorer software, four sets of primers were designed and RT-LAMP assay reaction conditions were optimized. The RT-LAMP was performed at different times by four primer sets. Agarose gel analysis showed that WYMV could be detected after 30 min with the primer set III and after 45 min with the other three primer sets, both und ...201122185375
effects of organic solvents on the reverse transcription reaction catalyzed by reverse transcriptases from avian myeloblastosis virus and moloney murine leukemia virus.the use of certain organic chemicals has been found to improve yields and specificity in pcr. in this study, we examined the effects of dimethyl sulfoxide (dmso), formamide, and glycerol on the reverse transcription reaction catalyzed by reverse transcriptases (rts) from avian myeloblastosis virus (amv) and moloney murine leukemia virus (mmlv). at 42 °c, dmso at 24% v/v and formamide at 12-14% inhibited the cdna synthesis reaction, but dmso at 12% and formamide at 6-8% improved the efficiency of ...201020834159
in vitro and in vivo studies of the rna conformational switch in alfalfa mosaic virus.the 3' termini of alfalfa mosaic virus (amv) rnas adopt two mutually exclusive conformations, a coat protein binding (cpb) and a trna-like (tl) conformer, which consist of a linear array of stem-loop structures and a pseudoknot structure, respectively. previously, switching between cpb and tl conformers has been proposed as a mechanism to regulate the competing processes of translation and replication of the viral rna (r. c. l. olsthoorn et al., embo j. 18:4856-4864, 1999). in the present study, ...201019923185
polymerase incorporation and miscoding properties of 5-chlorouracil.inflammation-mediated hypochlorous acid (hocl) can damage dna, dna precursors, and other biological molecules, thereby producing an array of damage products such as 5-chlorouracil (clu). in this study, we prepared and studied 5-chloro-2'-deoxyuridine (cldu) and clu-containing oligonucleotide templates. we demonstrate that human k-562 cells grown in culture with 10 mum cldu incorporate substantial amounts of cldu without significant toxicity. when in the template, clu residues pair with datp but ...201020104909
caulimoviridae tubule-guided transport is dictated by movement protein properties.plant viruses move through plasmodesmata (pd) either as nucleoprotein complexes (npcs) or as tubule-guided encapsidated particles with the help of movement proteins (mps). to explore how and why mps specialize in one mechanism or the other, we tested the exchangeability of mps encoded by dna and rna virus genomes by means of an engineered alfalfa mosaic virus (amv) system. we show that caulimoviridae (dna genome virus) mps are competent for rna virus particle transport but are unable to mediate ...201020130061
duplex structural differences and not 2'-hydroxyls explain the more stable binding of hiv-reverse transcriptase to rna-dna versus dna-dna.human immunodeficiency virus reverse transcriptase (hiv-rt) binds more stably in binary complexes with rna-dna versus dna-dna. current results indicate that only the -2 and -4 rna nucleotides (-1 hybridized to the 3' recessed dna base) are required for stable binding to rna-dna, and even a single rna nucleotide conferred significantly greater stability than dna-dna. replacing 2'- hydroxyls on pivotal rna bases with 2'-o-methyls did not affect stability, indicating that interactions between hydro ...201020338878
optimisation of reverse transcription can improve the sensitivity of rt-pcr for detection of classical swine fever virus.classical swine fever is a highly contagious, notifiable disease of pigs and wild boars listed by the world organisation for animal health (oie). therefore, methods employed in the diagnosis of csf should be fast, sensitive and specific. the aim of this study was optimisation of the reverse transcription reaction to increase the sensitivity of real-time rt-pcr for the detection of classical swine fever virus, the aetiological agent of the disease. the efficiency of reverse transcription reaction ...201020460224
complete genome amplification of equine influenza virus subtype 2.this work reports a method for rapid amplification of the complete genome of equine influenza virus subtype 2 (h3n8). a thermoscript reverse transcriptase instead of the avian myeloblastosis virus reverse transcriptase or moloney murine leukemia virus reverse transcriptase was used. this enzyme has demonstrated higher thermal stability and is described as suitable to make long cdna with a complex secondary structure. the product obtained by this method can be cloned, used in later sequencing rea ...200920085182
characterization of moloney murine leukaemia virus/avian myeloblastosis virus chimeric reverse transcriptases.reverse transcriptases (rts) from moloney murine leukaemia virus (mmlv) and avian myeloblastosis virus (amv) contain all the fingers, palm, thumb, connection and rnase h domains. the fingers, palm and thumb domains are thought to be involved in the reverse transcription activity, and the rnase h domain is in the rnase h activity. in this study, we characterized four chimeric rts which comprise one of the fingers, palm, thumb and rnase h domains originated from amv rt and the other three and conn ...200919060310
transient protein expression in three pisum sativum (green pea) varieties.the expression of proteins in plants both transiently and via permanently transformed lines has been demonstrated by a number of groups. transient plant expression systems, due to high expression levels and speed of production, show greater promise for the manufacturing of biopharmaceuticals when compared to permanent transformants. expression vectors based on a tobacco mosaic virus (tmv) are the most commonly utilized and the primary plant used, nicotiana benthamiana, has demonstrated the abili ...200919156736
evaluation of different rt enzyme standards for quantitation of retroviruses using the single-tube fluorescent product-enhanced reverse transcriptase assay.pcr-based reverse transcriptase (rt) assays are highly sensitive for broad detection of retroviruses. these assays are currently used for demonstrating the absence of retroviral contamination in vaccines and can also be applied to clinical and laboratory research to investigate low-virus replication. a single-tube fluorescent product-enhanced reverse transcriptase assay (stf-pert) has been published that was highly sensitive for retrovirus detection (<10 virions), with enhanced reproducibility a ...200919186191
explorations into neolignan biosynthesis: concise total syntheses of helicterin b, helisorin, and helisterculin a from a common intermediate.helicterins a and b (1 and 2), helisorin (3), and helisterculin a (4) are structurally unique natural products with the ability to combat the avian myeloblastosis virus. biogenetically, their architectures are considered to be products of seemingly straightforward diels-alder, radical-based, or acid-induced dimerizations of common, simpler precursors. yet, the pursuit of such blueprints in the laboratory has failed thus far in enabling their successful synthesis. herein, we describe the first to ...200919191699
evidence for variation in the optimal translation initiation complex: plant eif4b, eif4f, and eif(iso)4f differentially promote translation of mrnas.eukaryotic initiation factor (eif) 4b is known to interact with multiple initiation factors, mrna, rrna, and poly(a) binding protein (pabp). to gain a better understanding of the function of eif4b, the two isoforms from arabidopsis (arabidopsis thaliana) were expressed and analyzed using biophysical and biochemical methods. plant eif4b was found by ultracentrifugation and light scattering analysis to most likely be a monomer with an extended structure. an extended structure would facilitate the ...200919493973
higher random oligo concentration improves reverse transcription yield of cdna from bioptic tissues and quantitative rt-pcr reliability.real time quantitative reverse transcription-pcr (qrt-pcr) is the most sensitive technique for detection and quantification of mrna targets. reliable quantification of gene expression in formalin-fixed, paraffin-embedded tissues (ffpe), however, has been subjected to serious limitations so far, mainly due to the fragmentation of rna transcripts. we tried to improve the sensitivity and reliability of mrna quantification in ffpe by boosting the reverse transcription (rt) step, that is neglected in ...200919619529
myb-induced chromatin remodeling at a dual enhancer/promoter element involves non-coding rna transcription and is disrupted by oncogenic mutations of v-myb.the oncogene v-myb of avian myeloblastosis virus (amv) encodes a transcription factor (v-myb) that transforms myelomonocytic cells by deregulating the expression of specific target genes. v-myb has acquired its oncogenic potential by truncation as well as by a number of point mutations of its cellular progenitor c-myb. as a result of these changes, the target gene spectrum v-myb differs from that of c-myb. we recently showed that the chicken mim-1 gene, a c-myb target gene that is not activated ...200919841477
amino acid preferences of retroviral proteases for amino-terminal positions in a type 1 cleavage site.the specificities of the proteases of 11 retroviruses were studied using a series of oligopeptides with amino acid substitutions in the p1, p3, and p4 positions of a naturally occurring type 1 cleavage site (val-ser-gln-asn-tyr downward arrowpro-ile-val-gln) in human immunodeficiency virus type 1 (hiv-1). previously, the substrate specificity of the p2 site was studied for the same representative set of retroviral proteases, which included at least one member from each of the seven genera of the ...200818701588
fbxw7 acts as an e3 ubiquitin ligase that targets c-myb for nemo-like kinase (nlk)-induced degradation.the c-myb proto-oncogene product (c-myb) is degraded in response to wnt-1 signaling via a pathway involving tak1 (transforming growth factor-beta-activated kinase 1), hipk2 (homeodomain-interacting protein kinase 2), and nlk (nemo-like kinase). nlk directly binds to c-myb, which results in the phosphorylation of c-myb at multiple sites, and induces its ubiquitination and proteasome-dependent degradation. here, we report that fbxw7, the f-box protein of an scf complex, targets c-myb for degradati ...200818765672
a dual activation mechanism for myb-responsive genes in myelomonocytic cells.the retroviral oncogene v-myb encodes a transcription factor (v-myb) which is responsible for the transformation of myelomonocytic cells by avian myeloblastosis virus (amv). v-myb is thought to exert its biological effects by deregulating the expression of specific target genes. here we have used dnasei hypersensitive site mapping and reporter gene assays to study the activation of three myb target genes--mim-1, the lysozyme gene and the c/ebpbeta gene--all of which are activated by myb in myelo ...200817950008
comparison of the thermal stabilities of reverse transcriptases from avian myeloblastosis virus and moloney murine leukaemia virus.reverse transcriptases (rts) from avian myeloblastosis virus (amv) and moloney murine leukaemia virus (mmlv) have been most extensively used as a tool for conversion of rna to dna. in this study, we compared the thermal stabilities of amv rt and mmlv rt by observing their irreversible thermal inactivation. the temperatures reducing initial activity by 50% in 10-min incubation, t(50), of amv rt were 47 degrees c without the template-primer (t/p), poly(ra)-p(dt)(12-18), and 52 degrees c with the t ...200818006517
novel fret-based assay to detect reverse transcriptase activity using modified dutp analogues.we have developed a novel continuous assay to measure reverse transcriptase (rt) polymerase activity. the assay uses fluorescence energy transfer measurements to detect the incorporation of complementary pairs of fluorescently labeled deoxyuridine into cdna product. the fluorescently labeled dutp substrates were prepared using commercially available reagents with a simple coupling reaction. the fluorescent dye pairs have significant spectral overlap which allows fret interaction between dyes inc ...200818163534
v-myb suppresses phorbol ester- and modifies retinoic acid-induced differentiation of human promonocytic u937 cells.the c-myb protooncogene as well as its transforming derivate, the v-myb oncogene code for transcription factors. they regulate transcription of specific target genes thus controlling proliferation, differentiation and apoptosis of hematopoietic cells. up-regulation of the c-myb expression or rearrangement/amplification of the myb locus are often involved in leukemogenesis. enforced myb expression blocks differentiation of various leukemic cell lines. human promonocytes u937 can be induced to dif ...200818505338
avian myeoloblastosis virus (amv): only one side of the coin.for many years, scientists and suppliers have refered to amv-rt as the reverse transcriptase produced by the avian myelobalstosis virus. this manuscript briefly reviews the molecular basis for biological dependence of amv for the envelope and rt proteins that are produced by its natural helper the myeloblastosis associated virus (mav). because the wide use of the term <<amv rt>> obscures scientific facts, it is worthwhile to clarify this issue for the scientific community, especially for younger ...200818558010
rna-dependent dna polymerases.reverse transcriptases (rts) are multifunctional enzymes, but are mainly used as rna-directed dna polymerases in first-strand cdna synthesis. specifically, oligodeoxynucleotides are used as primers for extension on rna templates. the dna synthesized from an rna template is referred to as complementary dna (cdna) and is often used as a template for pcr or converted to dsdna for cloning. this unit describes appropriate reaction conditions for rts from moloney murine leukemia virus (mmlv) and avian ...200818972389
oncogenic point mutations in the myb dna-binding domain alter the dna-binding properties of myb at a physiological target gene.the oncoprotein v-myb of avian myeloblastosis virus (amv) transforms myelomonocytic cells by deregulating specific target genes. previous work has shown that the oncogenic potential of v-myb was activated by truncation of n- and c-terminal sequences of c-myb and was further increased by amino acid substitutions in the dna-binding domain and other parts of the protein. we have analyzed the activation of the chicken lysozyme gene which is strongly activated by c-myb but not by its oncogenic counte ...200717959653
alfalfa mosaic virus coat protein bridges rna and rna-dependent rna polymerase in vitro.alfalfa mosaic virus (amv) rna replication requires the viral coat protein (cp). amv cp is an integral component of the viral replicase; moreover, it binds to the viral rna 3'-termini and induces the formation of multiple new base pairs that organize the rna conformation. the results described here suggest that amv coat protein binding defines template selection by organizing the 3'-terminal rna conformation and by positioning the rna-dependent rna polymerase (rdrp) at the initiation site for mi ...200717400272
mn2+ suppressor mutations and biochemical communication between ty1 reverse transcriptase and rnase h domains.ty1 reverse transcriptase/rnase h (rt/rh) is exquisitely sensitive to manganese concentrations. elevated intracellular free mn(2+) inhibits ty1 retrotransposition and in vitro ty1 rt-polymerizing activity. furthermore, mn(2+) inhibition is not limited to the ty1 rt, as this ion similarly inhibits the activities of both avian myeloblastosis virus and human immunodeficiency virus type 1 rts. to further characterize mn(2+) inhibition, we generated rt/rh suppressor mutants capable of increased ty1 t ...200717537863
a proteomic analysis of protein variations during differentiation of v-myb-transformed monoblasts.v-myb oncogene of avian myeloblastosis virus (amv) transforms myelomonocytic cells in vitro and induces acute monoblastic leukemia in vivo. the transforming effect of the v-myb can be suppressed using phorbol ester (tpa) or histone deacetylase inhibitor trichostatin a (tsa), the inducers of cell differentiation that are in clinical trials. in this study, we used proteomics-based approach to identify proteins with variable expression in differentiated bm2 cells. proteome variations induced by tpa ...200716930693
the promoter regions of the myb-regulated adora2b and mcm4 genes co-localize with origins of dna replication.the retroviral oncogene v-myb encodes a transcription factor (v-myb) which is responsible for the transformation of myelomonocytic cells by avian myeloblastosis virus (amv). v-myb is thought to exert its biological effects by deregulating the expression of specific target genes. we have recently demonstrated that the chicken gas41 gene, whose promoter co-localizes with an origin of dna replication, is a bona fide myb target gene. because of this finding we have asked whether other myb-regulated ...200717822556
a chimeric plum pox virus shows reduced spread and cannot compete with its parental wild-type viruses in a mixed infection.the effect of a recombination event in the genomic 3' end on the biological properties and competitiveness of plum pox virus (ppv) was investigated. therefore, a fragment spanning the coat protein (cp) coding region and a part of the 3' non-translated region of a non-aphid-transmissible strain of ppv (ppv-nat) was replaced by the corresponding region of a ppv sour cherry isolate (ppv-soc). the resulting chimera (ppv-nat/soc) caused severe symptoms in nicotiana benthamiana, resembling those of pp ...200717872539
trans-lesion synthesis and rnaseh activity by reverse transcriptases on a true abasic rna template.while much is known about abasic dna, the biological impact of abasic rna is largely unexplored. to test the mutagenic potential of this rna lesion in the context of retroviruses, we synthesized a 31-mer oligoribonucleotide containing an abasic (ras) site and used it as a template for studying dna primer extension by hiv-1, avian myeloblastosis virus (amv) and moloney murine leukemia virus (mmlv) reversed transcriptases (rt). we found that trans-lesion synthesis readily takes place with hiv-1 rt ...200717932068
[study on evaluation of human immunodeficiency virus reverse transcriptase inhibitor by elisa].to establish an elisa method for detecting the activity of reverse transcriptase.200717940565
expression of active hbmp2 in transgenic tobacco plants.bone morphogenetic protein 2 (bmp2) is important for bone tissue repair. the goal of this research is to construct a high level human bmp2 (hbmp2) expression system using transgenic tobacco plants as a bioreactor. cauliflower mosaic virus (camv) 35s promoter, alfalfa mosaic virus (amv) enhancer, tobacco mosaic virus (tmv) enhancer, matrix attachment regions (mars) sequence, and "kozak" sequence were used to construct recombinant expression vectors and the high-expression vectors were screened ou ...200616819603
arabidopsis thaliana is an asymptomatic host of alfalfa mosaic virus.the susceptibility of arabidopsis thaliana ecotypes to infection by alfalfa mosaic virus (amv) was evaluated. thirty-nine ecotypes supported both local and systemic infection, 26 ecotypes supported only local infection, and three ecotypes could not be infected. no obvious symptoms characteristic of virus infection developed on the susceptible ecotypes under standard conditions of culture. parameters of amv infection were characterized in ecotype col-0, which supported systemic infection and accu ...200616875753
synthesis and biological evaluation of phosphonated dihydroisoxazole nucleosides.phosphonated isoxazolinyl nucleosides have been prepared via 1,3-dipolar cycloaddition reaction of nitrile oxides with corresponding vinyl or allyl nucleobases for antiviral studies. the cytotoxicity, the anti-hsv activity and the rt-inhibitory activity of the obtained compounds were evaluated and compared with those of azt and diethyl{(1'sr,4'rs)-1'-[[(5-methyl-2,4-dioxo-3,4-dihydropyrimidin-1(2h)-yl)]-3'-methyl-2'-oxa-3'-azacyclopent-4'-yl]}methylphosphonate, a saturated phosphonated dihydrois ...200616480883
effects of codon modification on human bmp2 gene expression in tobacco plants.bone morphogenetic protein 2 (bmp2) has great potential in therapeutic applications. we are working on generating transgenic plants as a bioreactor to produce bmp2. we have studied the effects of codon optimization on the expression of human bmp2 (hbmp2) in tobacco plants. three modified hbmp2 genes were transformed into tobacco under the control of either cauliflower mosaic virus 35s (camv35s) promoter or double-camv35s promoter plus alfalfa mosaic virus (amv) enhancer. the fused beta-glucuroni ...200616491379
identification of potential human oncogenes by mapping the common viral integration sites in avian nephroblastoma.gene deregulation is a frequent cause of malignant transformation. alteration of the gene structure and/or expression leading to cellular transformation and tumor growth can be experimentally achieved by insertion of the retroviral genome into the host dna. retrovirus-containing host loci found repeatedly in clonal tumors are called common viral integration sites (cvis). cvis are located in genes or chromosomal regions whose alterations participate in cellular transformation. here, we present th ...200616397219
sar and mode of action of novel non-nucleoside inhibitors of hepatitis c ns5b rna polymerase.novel non-nucleoside inhibitors of the hcv rna polymerase (ns5b) with sub-micromolar biochemical potency have been identified which are selective for the inhibition of hcv ns5b over other polymerases. the structures of the complexes formed between several of these inhibitors and hcv ns5b were determined by x-ray crystallography, and the inhibitors were found to bind in an allosteric binding site separate from the active site. structure-activity relationships and structural studies have identifie ...200616451069
evolutionary relationships among members of the bromoviridae deduced from whole proteome analysis.many molecular phylogenies of viruses build upon the analysis of single genes. the study of whole-genomes, however, might yield more reliable information to infer tree topologies and a better approach for drawing the evolutionary history of virus families. in this study, we apply a novel comparative proteomic approach to seek whether incorporating information from the entire proteome would support the actual taxonomy of the family bromoviridae. our results suggest that the current taxonomic clas ...200616172839
oncogenic mutations cause dramatic, qualitative changes in the transcriptional activity of c-myb.the v-myb oncoprotein encoded by avian myeloblastosis virus is highly oncogenic, induces leukemias in chickens and mice and transforms immature hematopoietic cells in vitro. the v-myb protein is a mutated and truncated version of c-myb, a dna-binding transcription factor expressed in many cell types that is essential for normal hematopoiesis. previous studies suggested that two types of differences, dna binding domain mutations and the deletion of a c-terminal negative regulatory domain were imp ...200616205643
cell-to-cell movement of alfalfa mosaic virus can be mediated by the movement proteins of ilar-, bromo-, cucumo-, tobamo- and comoviruses and does not require virion formation.rna 3 of alfalfa mosaic virus (amv) encodes the movement protein (mp) and coat protein (cp). chimeric rna 3 with the amv mp gene replaced by the corresponding mp gene of prunus necrotic ringspot virus, brome mosaic virus, cucumber mosaic virus or cowpea mosaic virus efficiently moved from cell-to-cell only when the expressed mp was extended at its c-terminus with the c-terminal 44 amino acids of amv mp. mp of tobacco mosaic virus supported the movement of the chimeric rna 3 whether or not the mp ...200616316673
occurrence of viruses infecting pea in iran.a survey was conducted to determine the incidence of alfalfa mosaic virus (amv), bean yellow mosaic virus (bymv), broad bean wilt virus-1 (bbwv), pea leafroll virus (plrv), pea enation mosaic virus (pemv), pea seed borne mosaic virus (psbmv), potato virus x(pvx), tomato mosaic virus (tomv), tomato spotted wilt virus (tswv) on pea (pisum sativum) in iran. a total of 1276 random and 684 symptomatic pea samples were collected during the spring and summer of 2002-2004 in tehran province of iran, whe ...200617390891
identification, characterization, and molecular detection of alfalfa mosaic virus in potato.abstract alfalfa mosaic virus (amv) was detected in potato fields in several provinces in canada and characterized by bioassay, enzyme-linked immunosorbent assay, and reverse-transcription polymerase chain reaction (rt-pcr). the identity of eight canadian potato amv isolates was confirmed by sequence analysis of their coat protein (cp) gene. sequence and phylogenetic analysis indicated that these eight amv potato isolates fell into one strain group, whereas a slight difference between ca175 and ...200618943961
effects of diterpenes isolated from the brazilian marine alga dictyota menstrualis on hiv-1 reverse transcriptase.it has been recently demonstrated that hiv-1 reverse transcriptase is the target of two diterpenes, (6 r)-6-hydroxydichotoma-3,14-diene-1,17-dial (compound 1) and (6 r)-6-acetoxydichotoma-3,14-diene-1,17-dial (compound 2), that inhibit hiv-1 replication in vitro. in this work, the effects of both diterpenes on the kinetic properties of the recombinant hiv-1 reverse transcriptase (rt) enzyme were evaluated. rna-dependent dna-polymerase (rddp) activity assays demonstrated that both diterpenes beha ...200516320202
optimization of acidothermus cellulolyticus endoglucanase (e1) production in transgenic tobacco plants by transcriptional, post-transcription and post-translational modification.an attempt was made to obtain a high-level production of intact acidothermus cellulolyticus endoglucanase (e1) in transgenic tobacco plants. the e1 expression was examined under the control of the constitutive and strong mac promoter or light-inducible tomato rubisco small sub-unit (rbcs-3c) promoter with its original or alfalfa mosaic virus (amv) rna4 5'-untranslated leader (utl) and targeted to different sub-cellular compartments via transit peptides. the transit peptides included native e1, e ...200516245154
amino acid preferences for a critical substrate binding subsite of retroviral proteases in type 1 cleavage sites.the specificities of the proteases of 11 retroviruses representing each of the seven genera of the family retroviridae were studied using a series of oligopeptides with amino acid substitutions in the p2 position of a naturally occurring type 1 cleavage site (val-ser-gln-asn-tyr pro-ile-val-gln; the arrow indicates the site of cleavage) in human immunodeficiency virus type 1 (hiv-1). this position was previously found to be one of the most critical in determining the substrate specificity differ ...200515767422
comparison of dna polymerase activities between recombinant feline immunodeficiency and leukemia virus reverse transcriptases.in this study, we present enzymatic differences found between recombinant rts derived from feline leukemia virus and feline immunodeficiency virus. firstly, fiv rt showed low steady state k(m) values for dntps compared to felv rt. consistent with this, fiv rt synthesized dna more efficiently than felv rt at low dntp concentrations. we observed similar concentration-dependent activity differences between other lentiviral (hiv-1 and siv) and non-lentiviral (mulv and amv) rts. second, felv rt showe ...200515823610
evaluation of the conformational switch model for alfalfa mosaic virus rna replication.key elements of the conformational switch model describing regulation of alfalfa mosaic virus (amv) replication (r. c. olsthoorn, s. mertens, f. t. brederode, and j. f. bol, embo j. 18:4856-4864, 1999) have been tested using biochemical assays and functional studies in nontransgenic protoplasts. although comparative sequence analysis suggests that the 3' untranslated regions of amv and ilarvirus rnas have the potential to fold into pseudoknots, we were unable to confirm that a proposed pseudokno ...200515827189
coat protein activation of alfalfa mosaic virus replication is concentration dependent.alfalfa mosaic virus (amv) and ilarvirus rnas are infectious only in the presence of the viral coat protein; therefore, an understanding of coat protein's function is important for defining viral replication mechanisms. based on in vitro replication experiments, the conformational switch model states that amv coat protein blocks minus-strand rna synthesis (r. c. olsthoorn, s. mertens, f. t. brederode, and j. f. bol, embo j. 18:4856-4864, 1999), while another report states that coat protein prese ...200515827190
coat protein enhances translational efficiency of alfalfa mosaic virus rnas and interacts with the eif4g component of initiation factor eif4f.the three plus-strand genomic rnas of alfalfa mosaic virus (amv) and the subgenomic messenger for viral coat protein (cp) contain a 5'-cap structure, but no 3'-poly(a) tail. binding of cp to the 3' end of amv rnas is required for efficient translation of the viral rnas and to initiate infection in plant cells. to study the role of cp in translation, plant protoplasts were transfected with luciferase (luc) transcripts with 3'-terminal sequences consisting of the 3' untranslated region of amv rna ...200515914864
cbp sensitizes v-myb-transformed monoblasts to differentiation inducers.creb-binding protein (cbp) regulates gene expression by binding to certain components of basal transcription machinery and by histone acetylation. in addition, it integrates various cellular signaling pathways through binding to multiple transcription factors, including the myb proteins. we report in this study that cbp can partially suppress function of the v-myb oncoprotein in leukemic cells. although originally described as an activator of v-myb function, we show that cbp can also act as a v- ...200516026544
replication of alfamo- and ilarviruses: role of the coat protein.in the family bromoviridae, a mixture of the three genomic rnas of bromo-, cucumo-, and oleaviruses is infectious as such, whereas the rnas of alfamo- and ilarviruses require binding of a few molecules of coat protein (cp) to the 3' end to initiate infection. most studies on the early function of cp have been done on the alfamovirus alfalfa mosaic virus (amv). the 3' 112 nucleotides of amv rnas can adopt two different conformations. one conformer consists of a trna-like structure that, together ...200516078876
v-myb mediates cooperation of a cell-specific enhancer with the mim-1 promoter.the oncogenic transcription factor v-myb disrupts myelomonocytic differentiation and transforms myelomonocytic cells by deregulating the expression of specific target genes. one of these genes, the chicken mim-1 gene, is activated by myb exclusively in myelomonocytic cells and, therefore, has been an interesting model system to study how myb activates a target in a lineage-specific manner. previous work has suggested that myb activates mim-1 by cooperating with ccaat box/enhancer binding protein ...200515601869
6-[1-(4-fluorophenyl)methyl-1h-pyrrol-2-yl)]-2,4-dioxo-5-hexenoic acid ethyl ester a novel diketo acid derivative which selectively inhibits the hiv-1 viral replication in cell culture and the ribonuclease h activity in vitro.the human immunodeficiency virus-type 1 (hiv-1) reverse transcriptase (rt) is a multifunctional enzyme which displays dna polymerase activity, which recognizes rna and dna templates, and a degradative ribonuclease h (rnase h) activity. while both rt functions are required for retroviral replication, until now only the polymerase function has been widely explored as drug target. we have identified a novel diketo acid derivative, 6-[1-(4-fluorophenyl)methyl-1h-pyrrol-2-yl)]-2,4-dioxo-5-hexenoic ac ...200515708638
detection of alfalfa mosaic virus (amv) in pea field in iran.during the spring and summer, in 2003-2004, pea viruses were identified in twenty pea fields of tehran. some leaf samples were collected randomly from pea fields of tehran. samples were tested by double antibody sandwich enzyme linked immunosorbent assay (das-elisa) technique using polyclonal antiserum of alfalfa mosaic virus (amv), as-0001, dsmz, braunschweig, germany). the samples were extracted in 0.1 m phosphate buffer ph 7 to 7.5 and inoculated on chenopodium amaranticolor, chenopodium quin ...200516637206
mutagenic properties of ribonucleotide analogues in reverse transcription with hiv and amv reverse transcriptases.pyrimidine analogues, n4-hydroxycytosine (c(oh)), n4-methoxycytosine (c(mo)) and 6h, 8h-3,4-dihydropyrimido[4,5-c][1,2]oxazin-7-one (p) can form base pairs with both adenine and guanine. we examined the mutagenic properties of these ribonucleotide analogues in rna in reverse transcription with hiv and amv reverse transcriptases. both reverse transcriptases incorporated datp and dgtp opposite these analogues in rna. the incorporation ratio of dgtp to datp opposite each analogue was measured to es ...200517150651
spider dragline silk proteins in transgenic tobacco leaves: accumulation and field production.spider dragline silk is a unique biomaterial and represents nature's strongest known fibre. as it is almost as strong as many commercial synthetic fibres, it is suitable for use in many industrial and medical applications. the prerequisite for such a widespread use is the cost-effective production in sufficient quantities for commercial fibre manufacturing. agricultural biotechnology and the production of recombinant dragline silk proteins in transgenic plants offer the potential for low-cost, l ...200417168889
experimental and natural weed host-virus relations.weeds, as alternative hosts of plant viruses and nutrient plants of virus vectors play important role in virus ecology and epidemiology. the aim of our study was to discover new weed-virus relations. therefore some weed species were mechanically inoculated with 28 viruses (strains or isolates) maintained in our glasshouse. different weed species with and without visible symptoms were collected from agro-, water ecosystems and wastelands of hungary between 1997 and 2003. virus infections were eva ...200415759395
improvement of extraction and processing of rna from renal biopsies.assessment of mrna levels has the potential to predict renal outcome. the objective of this study was to optimize several steps in the protocol for obtaining cdna from routine clinical kidney biopsy material.200414675040
spatial determinants of the alfalfa mosaic virus coat protein binding site.the biological functions of rna-protein complexes are, for the most part, poorly defined. here, we describe experiments that are aimed at understanding the functional significance of alfalfa mosaic virus rna-coat protein binding, an interaction that parallels the initiation of viral rna replication. peptides representing the rna-binding domain of the viral coat protein are biologically active in initiating replication and bind to a 39-nt 3'-terminal rna with a stoichiometry of two peptides: 1 rn ...200414681584
efficient translation of alfamovirus rnas requires the binding of coat protein dimers to the 3' termini of the viral rnas.the coat protein (cp) of alfalfa mosaic virus (amv) is required to initiate infection by the viral tripartite rna genome whereas infection by the tripartite brome mosaic virus (bmv) genome is independent of cp. amv cp stimulates translation of amv rna in vivo 50- to 100-fold. the 3' untranslated region (utr) of the amv subgenomic cp messenger rna 4 contains at least two cp binding sites. a cp binding site in the 3'-terminal 112 nucleotides of rna 4 was found to be required for efficient translat ...200414718638
structural organization of avian retrovirus integrase in assembled intasomes mediating full-site integration.retrovirus preintegration complexes (pic) purified from virus-infected cells are competent for efficient concerted integration of the linear viral dna ends by integrase (in) into target dna (full-site integration). in this report, we have shown that the assembled complexes (intasomes) formed in vitro with linear 3.6-kbp dna donors possessing 3'-oh-recessed attachment (att) site sequences and avian myeloblastosis virus in (4 nm) were as competent for full-site integration as isolated retrovirus p ...200414990573
similarities and differences between the subgenomic and minus-strand promoters of an rna plant virus.promoter regions required for minus-strand and subgenomic rna synthesis have been mapped for several plus-strand rna viruses. in general, the two types of promoters do not share structural features even though they are recognized by the same viral polymerase. the minus-strand promoter of alfalfa mosaic virus (amv), a plant virus of the family bromoviridae, consists of a triloop hairpin (hpe) which is attached to a 3' trna-like structure (tls). in contrast, the amv subgenomic promoter consists of ...200415047821
quantitation of mlv-based retroviral vectors using real-time rt-pcr.murine leukaemia virus-based vectors quantitation is a time consuming process that can take up to five days. in order to reduce this time a real-time rt-pcr was developed. this method quantifies vectors without an rna extraction step, using amv reverse transcriptase and lightcycler technology. besides a low quantitation time, this method has the advantages of using a plasmid dna standard curve with good reproducibility, and of having a high sensitivity (3 x 10(2) particles/microl) as well as an ...200415158592
detection of piscine nodaviruses by real-time nucleic acid sequence based amplification (nasba).nucleic acid sequence based amplification (nasba) is an isothermal nucleic acid amplification procedure based on target-specific primers and probes, and the co-ordinated activity of 3 enzymes: amv reverse transcriptase, rnase h, and t7 rna polymerase. we have developed a real-time nasba procedure for detection of piscine nodaviruses, which have emerged as major pathogens of marine fish. viral rna was isolated by guanidine thiocyanate lysis followed by purification on silica particles. primers we ...200415212274
degenerate in vitro genetic selection reveals mutations that diminish alfalfa mosaic virus rna replication without affecting coat protein binding.the alfalfa mosaic virus (amv) rnas are infectious only in the presence of the viral coat protein; however, the mechanisms describing coat protein's role during replication are disputed. we reasoned that mechanistic details might be revealed by identifying rna mutations in the 3'-terminal coat protein binding domain that increased or decreased rna replication without affecting coat protein binding. degenerate (doped) in vitro genetic selection, based on a pool of randomized 39-mers, was used to ...200415254175
differential sensitivity of v-myb and c-myb to wnt-1-induced protein degradation.recently we have shown that the c-myb proto-oncogene product (c-myb) is degraded in response to wnt-1 signaling via the pathway involving tak1 (transforming growth factor-beta-activated kinase), hipk2 (homeodomain-interacting protein kinase 2), and nlk (nemo-like kinase). nlk and hipk2 bind directly to c-myb, which results in the phosphorylation of c-myb at multiple sites, followed by its ubiquitination and proteasome-dependent degradation. the v-myb gene carried by avian myeloblastosis virus ha ...200415308626
analysis of mrna transcripts from the nad(p)h oxidase 1 (nox1) gene. evidence against production of the nadph oxidase homolog-1 short (noh-1s) transcript variant.recent reports indicate that nad(p)h oxidase 1 (nox1) mrna undergoes alternative splicing, producing a short transcript (noh-1s) encoding a novel h+ channel. although the h+ transport properties of noh-1s-transfected cells resemble those of many cells, the production of a noh-1s protein was never documented. we characterized nox1 transcripts in colon-derived cells and present evidence that mrna splicing does not produce noh-1s; rather, noh-1s appears to be an artifact of template switching durin ...200415375166
a short insert in the leader sequence of rna 3l, a long variant of alfalfa mosaic virus rna3, introduces two unidentified reading frames.n20-rna 3l, a large form of rna 3 associated with alfalfa mosaic virus (amv) strain n20 comprises 2281 nt and has approximately 97% overall sequence similarity to the longest previously described rna 3 of amv strain ysmv (ysmv-rna 3; 2188 nt). compared with ysmv-rna 3, n20-rna 3l contains an additional 97 nt in the 5' leader upstream of the open reading frames for movement protein (mp) and coat protein (cp). two overlapping unidentified reading frames (urf1 and urf2) result from this modificatio ...200415550770
e26 leukemia virus converts primitive erythroid cells into cycling multilineage progenitors.acute chicken leukemia retroviruses, because of their capacity to readily transform hematopoietic cells in vitro, are ideal models to study the mechanisms governing the cell-type specificity of oncoproteins. here we analyzed the transformation specificity of 2 acute chicken leukemia retroviruses, the myb-ets- encoding e26 virus and the erba/erbb-encoding avian erythroblastosis virus (aev). while cells transformed by e26 are multipotent (designated "mep" cells), those transformed by aev resemble ...200312393697
phosphorylation-dependent down-regulation of c-myb dna binding is abrogated by a point mutation in the v-myb oncogene.the viral myb (v-myb) oncoprotein of the avian myeloblastosis virus (amv) is an activated form of the cellular transcription factor c-myb causing acute monoblastic leukemia in chicken. oncogenic v-myb alterations include n- and c-terminal deletions as well as point mutations. whereas truncations in myb cause loss of various protein modifications, none of the point mutations in v-myb has been directly linked to protein modifications. here we show that the dna-binding domain of c-myb can be phosph ...200312456674
the importance of alfalfa mosaic virus coat protein dimers in the initiation of replication.deletion and substitution mutations affecting the oligomerization of alfalfa mosaic virus (amv) coat protein (cp) were studied in protoplasts to determine their effect on genome activation, an early step in amv replication. the cp mutants that formed dimers, cpdeltac9 and cpc-a(r)f, were highly active in initiating replication with 63-84% of wild-type (wt) cp activity. however, all mutants that did not form dimers, cpdeltac18, cpdeltac19, cpc-wfp, and cpc-w, were much less active with 19-33% of ...200312504539
single-tube fluorescent product-enhanced reverse transcriptase assay with ampliwax (stf-pert) for retrovirus quantitation.a taqman fluorescent probe-based product enhanced reverse transcriptase (rt) assay is described in which the rt and polymerase chain reaction (pcr) steps are set-up in a single tube, in two compartments separated by ampliwax (designated as single-tube fluorescent product-enhanced reverse transcriptase assay (stf-pert)). this simplification of the two-step method resulted in increased assay reproducibility and handling efficiency while maintaining the sensitivity of the pert assay (<10 virions). ...200312565165
an unnatural hydrophobic base pair with shape complementarity between pyrrole-2-carbaldehyde and 9-methylimidazo[(4,5)-b]pyridine.an unnatural hydrophobic base, pyrrole-2-carbaldehyde (denoted as pa), was developed as a specific pairing partner of 9-methylimidazo[(4,5)-b]pyridine (q). the q base is known to pair with 2,4-difluorotoluene (f) as an isostere of the a-t pair, and f also pairs with a efficiently in replication. in contrast, the q-pa pair showed specific selectivity in replication, and the five-membered-ring base pa paired efficiently with q but paired poorly with a. in addition, the interaction of pa with dna p ...200312720441
trichostatin a suppresses transformation by the v-myb oncogene in bm2 cells.bm2 cells are chicken monoblasts transformed by the v-myb oncogene of avian myeloblastosis virus. the constitutively high v-myb expression interferes with the terminal differentiation of bm2 cells, but these cells can be induced to differentiate into macrophage-like cells by phorbol esters. histone acetylation plays an important role in regulation of transcription and is particularly relevant to the regulation and pathology of hematopoiesis. in the present study, we examined the contribution of ...200312804181
expression of a recombinant gag protein from endogenous avian virus and its use in screening for antibody reactivity in recipients of chick-derived vaccines.virions incorporating endogenous avian virus (eav) rna have been identified in chick-derived biological products, including the vaccines used to protect against measles, mumps, and yellow fever. the presence of eav in these vaccines raises safety concerns regarding transmission to vaccine recipients. development of a serologic assay to detect antibodies to eav required the discovery of a diagnostic eav antigen and reactive antiserum. for this purpose, we have identified and expressed an eav caps ...200312922106
deletions within the u3 long terminal repeat alter the tumorigenic potential of myeloblastosis associated virus type 1(n).the molecularly cloned myeloblastosis-associated virus type-1(n) (mav-1(n)) strain induces specifically nephroblastomas in chicken. mav-induced nephroblastoma constitutes a unique animal model of the human wilms' tumor. we have previously shown that the mav-1(n) long terminal repeats (ltr) were necessary and sufficient for nephroblastoma induction. since major determinants for oncogenesis have been mapped in the u3 region of several other retroviruses, we have analyzed the tumorigenic potential ...200314599793
alfalfa mosaic virus: coat protein-dependent initiation of infection.summary taxonomy: alfalfa mosaic virus (amv) is the type species of the genus alfamovirus and belongs to the family bromoviridae. in this family, the tripartite rna genomes of bromo-, cucumo- and probably oleaviruses are infectious as such, whereas infection with the three genomic rnas of alfamo- and ilarviruses requires addition to the inoculum of a few molecules of coat protein (cp) per rna molecule. rnas 1 and 2 encode the replicase proteins p1 and p2, rna 3 encodes the movement protein and c ...200320569357
evidence for virus closely related to avian myeloblastosis-associated virus type 1 in a commercial stock of chickens.a two-round nested polymerase chain reaction assay detected rous associated virus-1 (rav-1), a prototype laboratory strain of avian leukosis virus of subgroup a (alv-a). surprisingly, the test failed to detect three field isolates of alv-a but did detect virus in one commercial stock of chickens (stock f). the sequence analysis of a core of 290 nucleotides of the env gene gave evidence that the virus from stock f was closely related to avian myeloblastosis-associated virus type one (mav-1). othe ...200317585462
translation inhibition of capped and uncapped viral rnas mediated by ribosome-inactivating proteins.abstract ribosome-inactivating proteins (rips) are n-glycosidases that remove specific purine residues from the sarcin/ricin (s/r) loop of the large rrna and arrest protein synthesis at the translocation step. in addition to their enzymatic activity, rips have been reputed to be potent antiviral agents against many plant, animal, and human viruses. we recently showed that pokeweed antiviral protein (pap), an rip from pokeweed, inhibits translation in cell extracts by binding to the cap structure ...200318942981
differential effects of v-jun and c-jun proteins on v-myb-transformed monoblasts.the v-myb oncogene of avian myeloblastosis virus transforms myelomonocytic cells in vitro. the line of v-myb-transformed chicken monoblasts bm2 can be induced to terminal differentiation using phorbol esters. the fact that jun proteins are up-regulated in the phorbol ester-treated bm2 cells prompted us to investigate the role of the jun proteins in regulation of myeloid differentiation. we ectopically expressed v-jun and c-jun in bm2 cells and evaluated their effects on differentiation and proli ...200212475180
purified tomato spotted wilt virus particles support both genome replication and transcription in vitro.purified tomato spotted wilt virus particles were shown to support either genome replication or transcription in vitro, depending on the conditions chosen. transcriptional activity was observed only upon addition of rabbit reticulocyte lysate, indicating a dependence on translation. under these conditions rna molecules of subgenomic length were synthesized that hybridized to strand-specific probes for the n and nss genes. cloning of these transcripts demonstrated the presence of nonviral leader ...200212490389
mechanism of c-myb-c/ebp beta cooperation from separated sites on a promoter.c-myb, but not avian myeloblastosis virus (amv) v-myb, cooperates with c/ebp beta to regulate transcription of myeloid-specific genes. to assess the structural basis for that difference, we determined the crystal structures of complexes comprised of the c-myb or amv v-myb dna-binding domain (dbd), the c/ebp beta dbd, and a promoter dna fragment. within the c-myb complex, a dna-bound c/ebp beta interacts with r2 of c-myb bound to a different dna fragment; point mutations in v-myb r2 eliminate suc ...200211792321
high processivity of the reverse transcriptase from a non-long terminal repeat retrotransposon.r2 is a retrotransposable element that specifically inserts into the 28 s rrna genes of arthropods. the element encodes a single protein with endonuclease activity that cleaves the 28 s gene target site and reverse transcriptase (rt) activity that uses the cleaved dna to prime reverse transcription. here we compare various properties of the r2 rt activity with those of the well characterized retroviral rt, avian myeloblastosis virus (amv). in processivity assays using heterogeneous rna templates ...200212101182
the role of template-primer in protection of reverse transcriptase from thermal inactivation.we compared the thermal stabilities of wild-type recombinant avian myeloblastosis virus (amv) and moloney murine leukemia virus (m-mlv) reverse transcriptase (rt) with those of mutants of the recombinant enzymes lacking rnase h activity. they differed in resistance to thermal inactivation at elevated temperatures in the presence of an rna/dna template-primer. rnase h-minus rts retained the ability to efficiently synthesize cdna at much higher temperatures. we show that the structure of the templ ...200212136094
role of an essential triloop hairpin and flanking structures in the 3' untranslated region of alfalfa mosaic virus rna in in vitro transcription.the minus-strand promoter of alfalfa mosaic virus (amv), a tripartite plant virus belonging to the family bromoviridae, is located within the 3'-terminal 145 nucleotides (nt), which can adopt a trna-like structure (tls). this contrasts with the subgenomic promoter for rna4 synthesis, which requires approximately 40 nt and forms a single triloop hairpin. detailed analysis of the minus-strand promoter now shows that a similar triloop hairpin, hairpin e (hpe), is crucial for minus-strand synthesis. ...200212163595
the viral envelope is a major determinant for the induction of lymphoid and myeloid tumours by avian leukosis virus subgroups a and j, respectively.among the six envelope subgroups of avian leukosis virus (alv) that infect chickens, subgroups a (alv-a) and j (alv-j) are the most pathogenic and widespread among commercial chicken populations. while alv-a is predominantly associated with lymphoid leukosis (ll) and less frequently with erythroblastosis (eb), alv-j mainly induces tumours of the myeloid lineage. in order to examine the basis for the lineage specificity of tumour induction by these two alv subgroups, we constructed two chimeric v ...200212237439
the deoxyribo-mode expression of primase activities of the primase-alpha dna polymerase enzyme complex associated with nucleoprotein complexes harboring an extrachromosomal dna identical with avian myeloblastosis virus core-bound dna: influencing by carbonyldiphosphonate, mimosine and butylphenyl deoxyguanosine-5'-triphosphate.the deoxyribo-mode expression of primase (pr) activities of the pr-alpha dna polymerase (pol) enzyme complex belonging to the naturally occurring nucleoprotein (np) complexes harboring an extrachromosomal dna identical with avian myeloblastosis virus (amv) core-bound dna (ríman and sulová, acta virol. 41, 181-192 (1997)) is similarly influenced by carbonyldiphosphonate (comdp) as the ribo-mode expression of pr activities (ríman, acta virol. 45, 109-124 (2001)). in the presence of all four common ...200212387498
role of the alfalfa mosaic virus methyltransferase-like domain in negative-strand rna synthesis.rnas 1 and 2 of the tripartite genome of alfalfa mosaic virus (amv) encode the replicase proteins p1 and p2, respectively. p1 contains a methyltransferase-like domain in its n-terminal half, which has a putative role in capping the viral rnas. six residues in this domain that are highly conserved in the methyltransferase domains of alphavirus-like viruses were mutated individually in amv p1. none of the mutants was infectious to plants. mutant rna 1 was coexpressed with wild-type (wt) rnas 2 and ...200212388692
rnas 1 and 2 of alfalfa mosaic virus, expressed in transgenic plants, start to replicate only after infection of the plants with rna 3.rnas 1 and 2 of the tripartite genome of alfalfa mosaic virus (amv) encode the two viral replicase subunits. full-length dna copies of rnas 1 and 2 were used to transform tobacco plants (r12 lines). none of the transgenic lines showed resistance to amv infection. in healthy r12 plants, the transcripts of the viral cdnas were copied by the transgenic viral replicase into minus-strand rnas but subsequent steps in replication were blocked. when the r12 plants were inoculated with amv rna 3, this bl ...200111125154
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