Academic literature on the topic 'RNA: DNA hybrides'
Create a spot-on reference in APA, MLA, Chicago, Harvard, and other styles
Consult the lists of relevant articles, books, theses, conference reports, and other scholarly sources on the topic 'RNA: DNA hybrides.'
Next to every source in the list of references, there is an 'Add to bibliography' button. Press on it, and we will generate automatically the bibliographic reference to the chosen work in the citation style you need: APA, MLA, Harvard, Chicago, Vancouver, etc.
You can also download the full text of the academic publication as pdf and read online its abstract whenever available in the metadata.
Journal articles on the topic "RNA: DNA hybrides"
1
Yang, Xuan, Binyuan Zhai, Shunxin Wang, Xiangfei Kong, Yingjin Tan, Lin Liu, Xiao Yang, Taicong Tan, Shuxian Zhang, and Liangran Zhang. "RNA-DNA hybrids regulate meiotic recombination." Cell Reports 37, no. 10 (December 2021): 110097. http://dx.doi.org/10.1016/j.celrep.2021.110097.
Full text
2
Kamath-Loeb, Ashwini S., Amnon Hizi, John Tabone, Marjorie S. Solomon, and Lawrence A. Loeb. "Inefficient Repair of RNA . DNA Hybrids." European Journal of Biochemistry 250, no. 2 (December 1997): 492–501. http://dx.doi.org/10.1111/j.1432-1033.1997.0492a.x.
Full text
3
Waldron, Denise. "RNA–DNA hybrids: double-edged swords." Nature Reviews Genetics 18, no. 1 (November 21, 2016): 3. http://dx.doi.org/10.1038/nrg.2016.153.
Full text
4
Hall, Kathleen B. "NMR spectroscopy of DNA/RNA hybrids." Current Opinion in Structural Biology 3, no. 3 (June 1993): 336–39. http://dx.doi.org/10.1016/s0959-440x(05)80103-4.
Full text
5
Kim, Joung Sug, Junghyun Park, Jang Hyeon Choi, Seungjae Kang, and Nokyoung Park. "RNA–DNA hybrid nano-materials for highly efficient and long lasting RNA interference effect." RSC Advances 13, no. 5 (2023): 3139–46. http://dx.doi.org/10.1039/d2ra06249f.
Full textAbstract:
A new RNAi approach was developed using an X-RDNA and Ri-Dgel. The nanostructured materials of dsRNA–DNA hybrids showed higher efficient and longer lasting RNA interference effect compared with conventional dsRNA.
6
Di, Lin, Yusi Fu, Yue Sun, Jie Li, Lu Liu, Jiacheng Yao, Guanbo Wang, et al. "RNA sequencing by direct tagmentation of RNA/DNA hybrids." Proceedings of the National Academy of Sciences 117, no. 6 (January 27, 2020): 2886–93. http://dx.doi.org/10.1073/pnas.1919800117.
Full textAbstract:
Transcriptome profiling by RNA sequencing (RNA-seq) has been widely used to characterize cellular status, but it relies on second-strand complementary DNA (cDNA) synthesis to generate initial material for library preparation. Here we use bacterial transposase Tn5, which has been increasingly used in various high-throughput DNA analyses, to construct RNA-seq libraries without second-strand synthesis. We show that Tn5 transposome can randomly bind RNA/DNA heteroduplexes and add sequencing adapters onto RNA directly after reverse transcription. This method, Sequencing HEteRo RNA-DNA-hYbrid (SHERRY), is versatile and scalable. SHERRY accepts a wide range of starting materials, from bulk RNA to single cells. SHERRY offers a greatly simplified protocol and produces results with higher reproducibility and GC uniformity compared with prevailing RNA-seq methods.
7
Paull, Tanya T. "RNA–DNA hybrids and the convergence with DNA repair." Critical Reviews in Biochemistry and Molecular Biology 54, no. 4 (July 4, 2019): 371–84. http://dx.doi.org/10.1080/10409238.2019.1670131.
Full text
8
Huang, Yuegao, Congju Chen, and Irina M. Russu. "Structural Energetics of Two RNA-DNA Hybrids." Biophysical Journal 96, no. 3 (February 2009): 578a. http://dx.doi.org/10.1016/j.bpj.2008.12.3022.
Full text
9
Vydzhak, Olga, Brian Luke, and Natalie Schindler. "Non-coding RNAs at the Eukaryotic rDNA Locus: RNA–DNA Hybrids and Beyond." Journal of Molecular Biology 432, no. 15 (July 2020): 4287–304. http://dx.doi.org/10.1016/j.jmb.2020.05.011.
Full text
10
Aguilera, Andrés, and Belén Gómez-González. "DNA–RNA hybrids: the risks of DNA breakage during transcription." Nature Structural & Molecular Biology 24, no. 5 (May 2017): 439–43. http://dx.doi.org/10.1038/nsmb.3395.
Full textDissertations / Theses on the topic "RNA: DNA hybrides"
1
Cohen, Sarah. "Le rôle de senataxine dans la résolution des hybrides ARN : ADN aux cassures double brins de l'ADN." Thesis, Toulouse 3, 2019. http://www.theses.fr/2019TOU30125.
Full textAbstract:
Les gènes transcriptionellement actifs peuvent être la source de l'instabilité du génome via de nombreux mécanismes. Ces gènes sont caractérisés par la formation de structures secondaires telles que les hybrides ADN : ARN. Ils se forment lorsque l'ARN sortant l'ARN polymérase II s'hybride au simple brin d'ADN. De nombreuses études ont montrées que l'accumulation de ces hybrides peut mener à la création de dommages à l'ADN. Parmi ces dommages, les Cassures Double Brins (CDB) sont les plus dangereuses pour la cellule puisqu'elles peuvent produire des mutations et des réarrangements chromosomiques. Il existe deux mécanismes de réparation majeurs dans la cellule : la Jonction Non-Homologue des Extrémités (NHEJ) et la Recombinaison Homologue (HR). Mon équipe a récemment montré que les CDB localisées dans les gènes transcrits sont préférentiellement réparés par HR. De plus, de nombreuses études ont montrées une interaction entre transcription et réparation des CDB. Au vue de ces résultats, nous avons donc émis l'hypothèse que les gènes transcriptionellement actifs pourraient être réparés par un mécanisme spécifique nécessitant l'activité de protéines associées à la transcription : "Réparation couplée à la transcription". Durant ma thèse, je me suis intéressée au rôle de deux protéines dans la réparation des régions transcrites en utilisant la lignée cellulaire DIvA (DSB Induction via AsiSI) qui permet l'induction de cassures annotées sur tout le génome. Premièrement, nous avons montré que la réparation des CDB dans des loci transcrits nécessitent une hélicase ADN : ARN connue : sénataxine (SETX). Après induction d'une cassure dans un gène, SETX est recrutée ce qui permet la résolution d'hybride ADN : ARN (cartographié par DRIP-seq). Nous avons aussi montré que SETX permet le recrutement de RAD51 et limite les jonctions illégitimes des CDB et par conséquent promeut la survie des cellules après induction des cassures. Cette étude montre que les CDB dans les loci transcrits requièrent la résolution spécifique des hybrides ADN : ARN par SETX pour permettre une réparation précise et est absolument indispensable pour la survie cellulaire. Deuxièmement, nous avons montré une interaction entre SETX et Bloom (BLM) une G4 DNA hélicase dans la réparation des CDB dans les régions transcrites. Nous avons montré que BLM est aussi recrutée au CDB dans les loci transcrits où elle est nécessaire à la résection et à la fidélité de réparation. De façon importante, nous avons montré que la déplétion de BLM restaure le défaut de survie cellulaire observé dans les cellules déplétées pour SETX après induction des CDB. La déplétion d'autres hélicases G4 (RTEL1, FANCJ) promeut aussi la survie des cellules déplétées pour SETX après dommages. Ces résultats suggèrent une interaction entre les hélicases G4 et la résolution des hybrides ADN : ARN dans la réparation des gènes actifs. En conclusion, ces études permettent une meilleure compréhension de la spécificité de la réparation des régions transcrites du génome, et notamment l'identification de protéines impliquées dans la "Réparation couplée à la Transcription"Actively transcribed genes can be the source of genome instability through numerous mechanisms. Those genes are characterized by the formation of secondary structures such as RNA-DNA hybrids. They are formed when nascent RNA exiting RNA polymerase II hybridizes single stranded DNA. Numerous studies have shown that RNA-DNA hybrids accumulation can lead to DNA damages. Among those damages, DNA double strand breaks (DSB) are the most deleterious for cells since they can generate mutations and chromosomal rearrangements. Two major repair mechanisms exist in the cell: Non-Homologous End-Joining (NHEJ) and Homologous recombination (HR). My lab showed recently that DSB occurring in transcribed genes are preferentially repaired by HR. Moreover, multiple studies have shown a cross talk between transcription and DSB repair. Those results led us to propose that actively transcribed genes could be repaired by a specific mechanism implicating proteins associated with transcription: "Transcription-coupled DSB repair". During my PhD, using the DIvA (DSB Induction via AsiSI) cell line allowing the induction of annotated DSB through the genome, I worked on 2 projects focusing on DSB repair in transcribed genes. First, we showed that DSB repair in transcribed loci requires a known RNA: DNA helicase: senataxin (SETX). After DSB induction in an active gene, SETX is recruited which allows RNA-DNA hybrid resolution (mapped by DRIP-seq). We also showed that SETX activity allows RAD51 loading and limits DSB illegitimate rejoining and consequently promotes cell survival after DSB induction. This study shows that DSB in transcribed loci require specific RNA-DNA hybrids removal by SETX for accurate repair. Second, we showed an interplay between SETX and Bloom (BLM) a G4 DNA helicase in DSB repair induced in transcribed loci. We showed that BLM is also recruited at DSB in transcribed loci where it promotes resection and repair fidelity. Strikingly, we showed that BLM depletion rescued the survival defects observed in SETX depleted cells following DSB induction. Knock down of other G4-helicases (RTEL1, FANCJ) also promoted cell survival in SETX depleted cells upon damage. Those data suggest an interplay between G4 helicases and RNA: DNA resolution for DSB repair in active genes. Altogether, these studies promote a better understanding of the specificity of DSB repair in transcriptionally active genes, and notably identification of proteins involved in "Transcription-coupled DSB repair"
2
Liu, Yaqun. "Study of transcription-replication conflict and its role in genomic instability and cancer development." Electronic Thesis or Diss., Université Paris sciences et lettres, 2022. http://www.theses.fr/2022UPSLS083.
Full textAbstract:
Les machineries de réplication et de transcription peuvent provoquer des conflits entre transcription et réplication (TRC), qui se produisent de manière frontale ou co-directionnelle. La collision frontale est considérée comme étant la plus délétère et peut conduire à de l’instabilité génomique au travers des R-loops qui se composent un hybride ADN-ARN et un brin d'ADN déplacé. En analysant les données multi-omiques, nous avons révélé avec succès que la pause transitoire de la fourche de réplication aux 3' des gènes enrichis en R-loops avec collision frontale affecte la stabilité génomique d'une manière dépendante de Topoisomérase1 (Nat.Communs. 2020) puis j'ai développé le premier outil bio-informatique pour analyser de données de réplication (OKseqHMM, disponible sur GitHub, Liu et al. BioRxiv. 2022). Finalement, il a été montré récemment que dans les cellules cancéreuses du sein, les R-loops colocalisent fortement avec une augmentation des cassures de l'ADN, de manière dépendante de la réplication. Nous visons à étudier le TRC dans des cellules cancéreuses et des échantillons de patients cancéreux pour déterminer comment le stress réplicatif induit de l'instabilité génomique dans le développent de cancer, ce qui pourront contribuer à l’établissement de nouvelles stratégies thérapeutiques contre le cancerReplication and transcription machinery can cause transcription-replication conflicts (TRCs), which occur either frontally or co-directionally. The head-on collision is considered to be the most deleterious and can lead to genomic instability through R-loops that consist of a DNA-RNA hybrid and a displaced DNA strand. By analyzing multi-omics data, we successfully revealed that transient replication forks pause at the 3' of genes enriched in R-loops with more head-on collisions affects genomic stability in a Topoisomerase1-dependent manner (Nat. Commons . 2020) then I developed the first bioinformatics tool to analyze replication data (OKseqHMM, available on GitHub, Liu et al. BioRxiv. 2022). Finally, it has recently been shown that in breast cancer cells, R-loops strongly colocalize with an increase in DNA breaks, in a replication-dependent manner. We aim to study TRC in cancer cells and samples from cancer patients to determine how replicative stress induces genomic instability in cancer development, which may contribute to the establishment of new therapeutic strategies against cancer
3
D'ALESSANDRO, GIUSEPPINA. "THE ROLE OF RNA AND DNA:RNA HYBRIDS AT DNA DOUBLE-STRAND BREAKS." Doctoral thesis, Università degli Studi di Milano, 2018. http://hdl.handle.net/2434/562552.
Full textAbstract:
The stability of our genome is constantly challenged by several genotoxic threats. DNA
double-strand breaks (DSBs) are the most dangerous DNA lesions that, if not repaired, can
lead to cancer initiation and progression and/or ageing. These detrimental consequences can
only be avoided if cells promptly recognize the lesions and signal their presence, thus
promoting either efficient repair and transient cell cycle arrest or cell death and cellular
senescence. This is the role of the DNA damage response (DDR) proteins and the newly
identified damage-induced non coding RNAs. We recently discovered that RNA polymerase
II is recruited to DSBs and synthetizes damage-induced non-coding RNAs (dilncRNAs).
DROSHA- and DICER-mediated processing of dilncRNAs generates small RNA species,
named DNA damage response RNA (DDRNAs) (Francia, 2012), that localize to DSBs via
pairing with dilncRNAs and promote DDR signaling (Michelini et al., in press). Similar
small non-coding RNA species discovered in plants are involved in DNA repair by
homologous recombination (HR) (Wei, 2012, Gao, 2014, Wang, 2016). In line with these
results, I report that transcriptional inhibition impairs recruitment of the HR proteins
BRCA1, BRCA2, and RAD51 to DSBs, while partially promoting DNA end resection.
Moreover, I show DNA:RNA hybrids accumulation at DSBs in mammalian cells by both
DRIP analyses and imaging techniques. Damage-induced DNA:RNA hybrids form upon the
hybridization of RNA species, likely dilncRNAs, to the resected DSBs DNA ends generated
during the S/G2 cell cycle phase. I also report that purified recombinant BRCA1 binds
DNA:RNA hybrids in vitro; moreover, DNA:RNA hybrids in vivo contribute to BRCA1
recruitment to DSBs. Consistent with the need to tightly regulate DNA:RNA hybrid levels,
I demonstrate that RNase H2, the major RNase H activity in mammalian nuclei, is recruited
to DSBs through direct interaction with RAD51. In summary, I report for the first time that
DNA:RNA hybrids accumulate at DSBs in mammalian cells in a cell-cycle- and DNA end
resection-depended way. At DSBs, BRCA1 directly recognizes DNA:RNA hybrids and
likely controls their turn-over by mediating the recruitment of RNase H2 via RAD51.
4
Xiong, Yong. "X-Ray crystallographic studies on DNA, RNA hybrids and duplexes containing single bulges /." The Ohio State University, 2000. http://rave.ohiolink.edu/etdc/view?acc_num=osu1488194825668504.
Full text
5
Novoa, Carolina. "RecQ-like helicase SGS1 counteracts DNA : RNA hybrid induced genome instability." Thesis, University of British Columbia, 2017. http://hdl.handle.net/2429/60964.
Full textAbstract:
Dividing cells are constantly under threat from both endogenous and exogenous DNA damaging stresses that can lead to mutations and structural variations in DNA. One contributor to genome instability is three-stranded DNA:RNA hybrid structures called R-loops. Though R-loops are known to induce DNA damage and DNA replication stress, it is unclear whether they are recognized and processed by an established DNA repair pathway prior to inducing DNA breaks. Canonically, DNA repair proteins work downstream of R-loop-induced DNA damage to stimulate repair and suppress genome instability. Recently, the possibility that some DNA repair pathways actively destabilize R-loops, thus preventing unscheduled DNA damage has emerged. Here we identify the helicase SGS1 as a suppressor of R-loop stability. Our data reveals that SGS1 depleted cells accumulate R-loops. In addition, we define a role for transcription in genome instability of cells lacking SGS1, which is consistent with an R-loop based mechanism. Hyper-recombination in SGS1 mutants is dependent on transcript length, transcription rate, and active DNA replication. Also, rDNA instability in sgs1Δ can be suppressed by ectopic expression of RNaseH1, a protein that degrades DNA:RNA hybrids. Interestingly, R-loops are known to form at rDNA loci. We favour a model in which SGS1 contributes to the stabilization of stalled replication forks associated with transcription complexes, and unresolved DNA:RNA hybrids. Finally, we showed that knockdown of the human Sgs1 orthologue BLM in HCT116 cells also led to the accumulation of more R-loops than control HCT116 cells. In summary, our data supports the idea that some DNA repair proteins involved in replication fork stabilization might also prevent and process R-loops.Science, Faculty of
Graduate
6
Yang, Diya. "Genome-wide Analysis of F1 Hybrids to Determine the Initiation of Epigenetic Silencing in Maize." Miami University / OhioLINK, 2021. http://rave.ohiolink.edu/etdc/view?acc_num=miami1610098527086245.
Full text
7
Ly, Danith. "Mechanism of electron transfer in double-stranded DNA and PNA-DNA hybrids, and the development of a fluorescence probe for DNA and RNA detection." Diss., Georgia Institute of Technology, 1998. http://hdl.handle.net/1853/30485.
Full text
8
Rigby, Rachel Elizabeth. "Ribonuclease H2, RNA:DNA hybrids and innate immunity." Thesis, University of Edinburgh, 2011. http://hdl.handle.net/1842/6509.
Full textAbstract:
The activation of the innate immune system is the first line of host defence against infection. Nucleic acids can potently stimulate this response and trigger a series of signalling cascades leading to cytokine production and the establishment of an inflammatory state. Mutations in genes encoding nucleases have been identified in patients with autoimmune diseases, including Aicardi-Goutières syndrome (AGS). This rare childhood inflammatory disorder is characterised by the presence of high levels of the antiviral cytokine interferon-α in the cerebrospinal fluid and blood, which is thought to be produced as a consequence of the activation of the innate immunity by unprocessed self-nucleic acids. This thesis therefore aimed to define the role of one of the AGS nucleases, the Ribonuclease H2 (RNase H2) complex, in innate immunity, and to establish if nucleic acid substrates of this enzyme were able to induce type I interferon production in vitro. The AGS nucleases may function as components of the innate immune response to nucleic acids. Consistent with this hypothesis, RNase H2 was constitutively expressed in immune cells, however, its expression was not upregulated in response to type I interferons. RNase H2-deficient cells responded normally to a range of nucleic acid PAMPs, which implied that a role for RNase H2 as a negative regulator of the immune response was unlikely, in contrast to the reported cellular functions of two other AGS proteins, TREX1 and SAMHD1. Therefore, no clear evidence was found for the direct involvement of RNase H2 in the innate immune response to nucleic acids. An alternative model for the pathogenesis of disease hypothesises that decreased RNase H2 activity within the cell results in an accumulation of RNA:DNA hybrids. To investigate the immunostimulatory potential of such substrates, RNA:DNA hybrids with different physiochemical properties were designed and synthesised. Methods to purify the hybrids from other contaminating nucleic acid species were established and their capacity as activators of the innate immune response tested using a range of in vitro cellular systems. A GU-rich 60 bp RNA:DNA hybrid was shown to be an effective activator of a pro-inflammatory cytokine response exclusively in Flt3-L bone marrow cultures. This response was completely dependent on signalling involving MyD88 and/or Trif, however the specific receptor involved remains to be determined. Reduced cellular RNase H2 activity did not affect the ability of Flt3-L cultures to mount a cytokine response against the RNA:DNA hybrid. These in vitro studies suggested that RNA:DNA hybrids may be a novel nucleic acid PAMP. Taken together, the data in this thesis suggest that the cellular function of RNase H2 is in the suppression of substrate formation rather than as a component of the immune response pathways. Future studies to identify endogenous immunostimulatory RNA:DNA hybrids and the signalling pathways activated by them should provide a detailed understanding of the molecular mechanisms involved in the pathogenesis of AGS and related autoimmune diseases.
9
Islam, Mohammad Kaisarul. "Novel ligands targeting the DNA/RNA hybrid and telomeric quadruplex as potential anticancer agents." Thesis, King's College London (University of London), 2016. https://kclpure.kcl.ac.uk/portal/en/theses/novel-ligands-targeting-the-dnarna-hybrid-and-telomeric-quadruplex-as-potential-anticancer-agents(ce8f3d0e-317d-4c2e-b64a-e13e283f7b95).html.
Full textAbstract:
Telomeres are repetitive sequences of DNA at the ends of chromosomes that become progressively shorter during cell division, acting as a form of “biological clock” causing cell death once they have reached a certain length. Almost 90% of cancer cells overexpress the enzyme telomerase which can lengthen telomeres and confer immortality to the tumour cells. Thus, telomerase has become an important target for drug discovery in the oncology area, and there is also interest from researchers investigating the aging process. During the catalytic cycle of telomerase, a unique DNA/RNA hybrid duplex (DRH) forms that is typically between 6-11 base pairs long and is key to extending the telomere. There is interest in discovering small drug-like molecules that can recognize and bind to this hybrid duplex to inhibit selectively telomerase, either by stabilizing the structure and thereby preventing telomerase dissociation (a key step in the catalytic cycle) or by sufficiently distorting the hybrid duplex to cause the misalignment of key catalytic groups. This project began by using oligonucleotides representing DNA/RNA hybrid duplex (DRH), telomeric G-quadruplex and control duplex DNA sequences to screen against the National Cancer Institute compound libraries (i.e., Diversity Set II, Mechanistic Set and Natural Product Set) using a high throughput Fluorescent Resonance Energy Transfer (FRET)-based DNA thermal denaturation assay to determine binding affinity and specificity. Thirteen novel chemical scaffold families were identified in the assay, compounds which showed a >5 °C selective stabilization of the DNA/RNA hybrid duplex at a 1 μM ligand concentration. Chemical modifications were then made to these scaffolds to generate focused libraries of analogues to improve selectivity, potency and drug-likeness, and to provide Structure-Activity Relationship (SAR) information. A total of 49 novel molecules were synthesized and then screened against an expanded range of four different nucleic acid constructs including telomeric and DNA/RNA hybrid duplex sequences. A number of compounds showed selective DNA/RNA hybrid stabilization potential with some compounds also showing notable telomeric G-quadruplex stabilization without significant affinity for promoter G-quadruplexes (i.e., c-Kit1, c-Kit-2 and c-Myc) and control duplex DNA sequences. The compounds from library-1 provided DNA/RNA hybrid duplex stabilization in the 0.5-7.2 C range and telomeric G-quadruplex stabilization in the 0.2-6.5 C range at a 1 μM ligand concentration. Molecular modelling and molecular dynamics studies confirmed that the methylene spacer between the benzimidazole and phenylene moieties of molecules within library-1 is perfectly shaped to fit within the DRH sequence. In addition, it was confirmed that minor-groove binding and simultaneous intercalation between the nucleobases of a DNA/RNA hybrid duplex requires a linker of specific length (i.e., an eight methylene spacer as in compound 3.3). Selected compounds were then studied further using a variety of biological techniques to confirm selective telomerase inhibition and cell-based assays to utilize their potential as antitumour agents.
10
Beckedorff, Felipe César Ferrarezi. "Recrutamento do complexo repressivo polycomb 2 pelo RNA não codificador longo antissenso ANRASSF1 modula a expressão do gene RASSF1A e a proliferação celular." Universidade de São Paulo, 2012. http://www.teses.usp.br/teses/disponiveis/46/46131/tde-23042013-083641/.
Full textAbstract:
O gene supressor tumoral RASSF1A tem sido associado com redução da proliferação celular em diversos tumores. Sua expressão é regulada por eventos epigenéticos que envolvem o complexo repressivo polycomb (PRC2), no entanto os mecanismos moleculares da modulação do recrutamento deste modificador epigenético para este locus ainda são desconhecidos. Neste trabalho identificamos e caracterizamos ANRASSF1, um RNA não codificador longo (lncRNA) intrônico unspliced, que é transcrito na fita oposta do gene RASSF1A, em várias linhagem celulares e tecidos, e se liga a PRC2. ANRASSF1 é transcrito pela RNAPII, possui cap-5´ e cauda poli-A, além de localizar-se no núcleo e possuir uma meia-vida em média quatro vezes menor comparada com outros lncRNAs ligados à PRC2. A super-expressão ectópica de ANRASSF1 reduziu os níveis de RASSF1A e aumentou a taxa de proliferação em células HeLa, enquanto seu silenciamento provocou efeito oposto. Essas mudanças nos níveis de ANRASSF1 não afetaram a abundância da isoforma RASSF1C em nenhuma das condições. A super-expressão de ANRASSF1 provocou um grande aumento tanto da ocupação de PRC2 como da marca de histona repressiva H3K27me3 especificamente na região promotora RASSF1A. Nenhum efeito da super-expressão de ANRASSF1 foi detectado na ocupação de PRC2 e na histona H3K27me3 nas regiões promotoras de RASSF1C e de outros quatro genes vizinhos, incluindo dois genes supressores tumorais bem caracterizados. Além disso, foi demonstrado que ANRASSF1 forma um híbrido de RNA/DNA e recruta SUZ12, um componente do PRC2, para o promotor de RASSF1A. Notavelmente, foi detectado pelo ensaio de RNase-ChIP que a degradação de ANRASSF1 diminui a ocupação de PRC2 neste promotor. Esses resultados demonstram um novo mecanismo de repressão epigenética do supressor tumoral RASSF1A, envolvendo um lncRNA unspliced antissenso, onde ANRASSF1 reprime seletivamente a expressão da isoforma de RASSF1 que sobrepõe o transcrito antissenso de modo local e específico. Considerando uma perspectiva mais ampla, nossos resultados sugerem que outros lncRNAs intrônicos unspliced não caracterizados no genoma humano podem contribuir para uma modulação epigenética local e específica de cada região em que os lncRNAs são transcritos.Tumor-suppressor RASSF1A gene down-regulation has been implicated in increasing cell proliferation in several tumors. Its expression is regulated by epigenetic events involving polycomb repressive complex 2 (PRC2), however the molecular mechanisms modulating recruitment of this epigenetic modifier to the locus remain largely unknown. Here, we identify and characterize ANRASSF1, an endogenous unspliced long noncoding RNA (lncRNA) that is transcribed from the opposite strand of RASSF1 gene in several cell lines and tissues, and binds to PRC2. ANRASSF1 is transcribed by RNA Polymerase II, 5\'-capped, polyadenylated, displays nuclear localization, and has on average a four-fold shorter half-life compared to other lncRNAs that bind PRC2. ANRASSF1 ectopic overexpression decreases RASSF1A abundance and increases the proliferation rate of HeLa cells, whereas its silencing causes opposite effects. These changes in NRASSF1 levels do not affect RASSF1C isoform abundance. ANRASSF1 overexpression causes a marked increase both in PRC2 occupancy and in histone H3K27me3 repressive mark specifically at the RASSF1A promoter region. No effect of ANRASSF1 overexpression is detected on PRC2 occupancy and on histone H3K27me3 at the promoter regions of RASSF1C and of four other neighbor genes, including two well-characterized tumor suppressor genes. Additionally, we demonstrate that ANRASSF1 forms an RNA/DNA hybrid, and recruits SUZ12, a PRC2 component, to the RASSF1A promoter. Notably, depletion of ANRASSF1 disrupts SUZ12 occupancy on RASSF1A promoter as measured by RNAse-ChIP assay. Together, these results show a new mechanism of epigenetic repression of RASSF1A tumor suppressor gene involving an antisense unspliced lncRNA, in which ANRASSF1 selectively represses expression of the RASSF1 isoform overlapping the antisense transcript in a location-specific manner. In a broader perspective, our findings suggest that other non-characterized unspliced intronic lncRNAs transcribed in the human genome may contribute to a location-specific epigenetic modulation of genes.
Book chapters on the topic "RNA: DNA hybrides"
1
Martins, Angelica N., Weina Ke, Vaishnavi Jawahar, Morriah Striplin, Caryn Striplin, Eric O. Freed, and Kirill A. Afonin. "Intracellular Reassociation of RNA–DNA Hybrids that Activates RNAi in HIV-Infected Cells." In RNA Nanostructures, 269–83. New York, NY: Springer New York, 2017. http://dx.doi.org/10.1007/978-1-4939-7138-1_18.
Full text
2
Rolband, Lewis A., Weina Ke, and Kirill A. Afonin. "Aptamer Conjugated RNA/DNA Hybrid Nanostructures Designed for Efficient Regulation of Blood Coagulation." In RNA Nanostructures, 277–86. New York, NY: Springer US, 2023. http://dx.doi.org/10.1007/978-1-0716-3417-2_19.
Full text
3
García-Rubio, María, Sonia I. Barroso, and Andrés Aguilera. "Detection of DNA-RNA Hybrids In Vivo." In Methods in Molecular Biology, 347–61. New York, NY: Springer New York, 2017. http://dx.doi.org/10.1007/978-1-4939-7306-4_24.
Full text
4
Wagner, Carolin B., and Brian Luke. "DNA–RNA Hybrids at Telomeres in Budding Yeast." In R-Loops, 145–57. New York, NY: Springer US, 2022. http://dx.doi.org/10.1007/978-1-0716-2477-7_10.
Full text
5
Wheelhouse, Richard T., and Jonathan B. Chaires. "Drug Binding to DNA⋅RNA Hybrid Structures." In Methods in Molecular Biology, 55–70. Totowa, NJ: Humana Press, 2009. http://dx.doi.org/10.1007/978-1-60327-418-0_4.
Full text
6
Lane, D. J., and M. L. Collins. "Current Methods for Detection of DNA/Ribosomal RNA Hybrids." In Rapid Methods and Automation in Microbiology and Immunology, 54–75. Berlin, Heidelberg: Springer Berlin Heidelberg, 1991. http://dx.doi.org/10.1007/978-3-642-76603-9_8.
Full text
7
Abakir, Abdulkadir, Fahad Alenezi, and Alexey Ruzov. "Detecting and Mapping N6-Methyladenosine on RNA/DNA Hybrids." In R-Loops, 329–44. New York, NY: Springer US, 2022. http://dx.doi.org/10.1007/978-1-0716-2477-7_22.
Full text
8
Majeed, Saadat, Muhammad Umer Farooq, Sayed Tayyab Raza Naqvi, Batool Fatima, Muhammad Najam-ul-Haq, Sabahat Majeed, Fahad Ali, and Naeem Akhtar Khan. "MOF-based Electrochemical Sensors for DNA/RNA/ATP." In Metal-Organic Frameworks-Based Hybrid Materials for Environmental Sensing and Monitoring, 237–47. New York: CRC Press, 2022. http://dx.doi.org/10.1201/9781003188148-25.
Full text
9
Starczak, Marta, Abdulkadir Abakir, Alexey Ruzov, and Daniel Gackowski. "Detection and Quantification of RNA Modifications on RNA–DNA Hybrids Using SID-UPLC-MS/MS." In R-Loops, 127–43. New York, NY: Springer US, 2022. http://dx.doi.org/10.1007/978-1-0716-2477-7_9.
Full text
10
Silva, Sónia, Cristina Guillén-Mendoza, and Andrés Aguilera. "RNase H1 Hybrid-Binding Domain-Based Tools for Cellular Biology Studies of DNA–RNA Hybrids in Mammalian Cells." In R-Loops, 115–25. New York, NY: Springer US, 2022. http://dx.doi.org/10.1007/978-1-0716-2477-7_8.
Full textConference papers on the topic "RNA: DNA hybrides"
1
Rajput, B., D. Alaimo, A. M. Asselbergs, and E. Reich. "CONSTRUCTION AND EXPRESSION OF HYBRID PLASMINOGEN ACTIVATOR GENES." In XIth International Congress on Thrombosis and Haemostasis. Schattauer GmbH, 1987. http://dx.doi.org/10.1055/s-0038-1644412.
Full textAbstract:
Hybrid plasminogen activator (PA) genes containing fragments of cDNA encoding the non-catalytic part of tissue-PA and the .catalytic domain of urokinase and vice versa were constructed and expressed in Chinese Hamster ovary (CHO) cells. The hybrid nature of the products in stably transformed cells was analyzed at the level of DNA and RNA using probes derived from different regions of the urokinase and tissue-PA cDNAs and at the protein level by means of polyclonal antibodies raised against tissue-PA and urokinase. The hybrid genes made hybrid RNAs and proteins of the expected sizes. The proteins were enzymatically active as determined by zymography and chromogenic enzyme assays and this activity was blocked by the appropriate antibodies. The effect on hybrid PAs of cyanogen bromide cleaved fibrinogen fragments, poly-D-lysine and heparin which are known to affect the activity of tissue-PA and urokinase differently was also studied
2
Maciag, Anna E., Joseph E. Saavedra, Ryan J. Holland, Youseung Kim, Vandana Kumari, Christina E. Luthers, Waheed S. Sehareen, Xinhua Ji, Lucy M. Anderson, and Larry K. Keefer. "Abstract 3334: GSTP1-activated nitric oxide-releasing/PARP inhibitor hybrid prodrugs induce cancer cell death through ROS/RNS, DNA damage, ER stress, and apoptosis." In Proceedings: AACR 104th Annual Meeting 2013; Apr 6-10, 2013; Washington, DC. American Association for Cancer Research, 2013. http://dx.doi.org/10.1158/1538-7445.am2013-3334.
Full textReports on the topic "RNA: DNA hybrides"
1
Dugan, L. Elucidation of the Mechanism of Gene Silencing using Small Interferin RNA: DNA Hybrid Molecules. Office of Scientific and Technical Information (OSTI), February 2006. http://dx.doi.org/10.2172/900164.
Full text
2
Bar-Joseph, Moshe, William O. Dawson, and Munir Mawassi. Role of Defective RNAs in Citrus Tristeza Virus Diseases. United States Department of Agriculture, September 2000. http://dx.doi.org/10.32747/2000.7575279.bard.
Full textAbstract:
This program focused on citrus tristeza virus (CTV), the largest and one of the most complex RNA-plant-viruses. The economic importance of this virus to the US and Israeli citrus industries, its uniqueness among RNA viruses and the possibility to tame the virus and eventually turn it into a useful tool for the protection and genetic improvement of citrus trees justify these continued efforts. Although the overall goal of this project was to study the role(s) of CTV associated defective (d)-RNAs in CTV-induced diseases, considerable research efforts had to be devoted to the engineering of the helper virus which provides the machinery to allow dRNA replication. Considerable progress was made through three main lines of complementary studies. For the first time, the generation of an engineered CTV genetic system that is capable of infecting citrus plants with in vitro modified virus was achieved. Considering that this RNA virus consists of a 20 kb genome, much larger than any other previously developed similar genetic system, completing this goal was an extremely difficult task that was accomplished by the effective collaboration and complementarity of both partners. Other full-length genomic CTV isolates were sequenced and populations examined, resulting in a new level of understanding of population complexities and dynamics in the US and Israel. In addition, this project has now considerably advanced our understanding and ability to manipulate dRNAs, a new class of genetic elements of closteroviruses, which were first found in the Israeli VT isolate and later shown to be omnipresent in CTV populations. We have characterized additional natural dRNAs and have shown that production of subgenomic mRNAs can be involved in the generation of dRNAs. We have molecularly cloned natural dRNAs and directly inoculated citrus plants with 35S-cDNA constructs and have shown that specific dRNAs are correlated with specific disease symptoms. Systems to examine dRNA replication in protoplasts were developed and the requirements for dRNA replication were defined. Several artificial dRNAs that replicate efficiently with a helper virus were created from infectious full-genomic cDNAs. Elements that allow the specific replication of dRNAs by heterologous helper viruses also were defined. The T36-derived dRNAs were replicated efficiently by a range of different wild CTV isolates and hybrid dRNAs with heterologous termini are efficiently replicated with T36 as helper. In addition we found: 1) All CTV genes except of the p6 gene product from the conserved signature block of the Closteroviridae are obligate for assembly, infectivity, and serial protoplast passage; 2) The p20 protein is a major component of the amorphous inclusion bodies of infected cells; and 3) Novel 5'-Co-terminal RNAs in CTV infected cells were characterized. These results have considerably advanced our basic understanding of the molecular biology of CTV and CTV-dRNAs and form the platform for the future manipulation of this complicated virus. As a result of these developments, the way is now open to turn constructs of this viral plant pathogen into new tools for protecting citrus against severe CTV terms and development of virus-based expression vectors for other citrus improvement needs. In conclusion, this research program has accomplished two main interconnected missions, the collection of basic information on the molecular and biological characteristics of the virus and its associated dRNAs toward development of management strategies against severe diseases caused by the virus and building of novel research tools to improve citrus varieties. Reaching these goals will allow us to advance this project to a new phase of turning the virus from a pathogen to an ally.
3
Sink, Ken, Shamay Izhar, and Abraham Nachmias. Asymmetric Somatic Hybridization: Developing a Gene Transfer System for Solanaceous Vegetable Crops. United States Department of Agriculture, February 1996. http://dx.doi.org/10.32747/1996.7613010.bard.
Full textAbstract:
Highly asymmetric somatic hybrid plants were obtained by PEG/DMSO fusion of gamma irradiated (100, 250, 7500 and 1000 Gy) protoplasts of a (KmR-) interspecific hybrid Lycopersicon esculentum x L. pennellii (EP) with protoplasts of eggplant (E). Somatic hybrid calli were selected based on kanamycin resistance and verified by PCR of the NptII gene, RAPD's and Southern's using potato rDNA pTHG2 probes. Flow cytometry indicated all hybrid calli that did not regenerate shoots were 5-9n. Three asymmetric plants regenerated only from callus close to 4n and such calli oly occurred when EP received 100 Gy. The asymmetric plants had eggplant morphology and regenerated from one hybrid callus with 6.29 average size tomato chromosomes. Limited amounts of EP DNA were found in the three somatic hybrid plants H18-1 to -3 by dot-blot hybridization with probe pTHG2, to be equivalent to 6.23, 5.41, and 5.95 % EP, respectively. RFLP analysis of Lycopersicon esculentum and L. pennellii specific chromosomes revealed that only fragments of 8 to 10 out of the 24 EP chromosomes are present in the asymmetric plants. Transgenic plants 2-3, 2-4 and 10-3 were found resistant to verticillium; suggesting successful transfer of the Ve complex from S. torvum to eggplant.
4
Ostersetzer-Biran, Oren, and Jeffrey Mower. Novel strategies to induce male sterility and restore fertility in Brassicaceae crops. United States Department of Agriculture, January 2016. http://dx.doi.org/10.32747/2016.7604267.bard.
Full textAbstract:
Abstract Mitochondria are the site of respiration and numerous other metabolic processes required for plant growth and development. Increased demands for metabolic energy are observed during different stages in the plants life cycle, but are particularly ample during germination and reproductive organ development. These activities are dependent upon the tight regulation of the expression and accumulation of various organellar proteins. Plant mitochondria contain their own genomes (mtDNA), which encode for rRNAs, tRNAs and some mitochondrial proteins. Although all mitochondria have probably evolved from a common alpha-proteobacterial ancestor, notable genomic reorganizations have occurred in the mtDNAs of different eukaryotic lineages. Plant mtDNAs are notably larger and more variable in size (ranging from 70~11,000 kbp in size) than the mrDNAs in higher animals (16~19 kbp). Another unique feature of plant mitochondria includes the presence of both circular and linear DNA fragments, which undergo intra- and intermolecular recombination. DNA-seq data indicate that such recombination events result with diverged mitochondrial genome configurations, even within a single plant species. One common plant phenotype that emerges as a consequence of altered mtDNA configuration is cytoplasmic male sterility CMS (i.e. reduced production of functional pollen). The maternally-inherited male sterility phenotype is highly valuable agriculturally. CMS forces the production of F1 hybrids, particularly in predominantly self-pollinating crops, resulting in enhanced crop growth and productivity through heterosis (i.e. hybrid vigor or outbreeding enhancement). CMS lines have been implemented in some cereal and vegetables, but most crops still lack a CMS system. This work focuses on the analysis of the molecular basis of CMS. We also aim to induce nuclear or organellar induced male-sterility in plants, and to develop a novel approach for fertility restoration. Our work focuses on Brassicaceae, a large family of flowering plants that includes Arabidopsis thaliana, a key model organism in plant sciences, as well as many crops of major economic importance (e.g., broccoli, cauliflower, cabbage, and various seeds for oil production). In spite of the genomic rearrangements in the mtDNAs of plants, the number of genes and the coding sequences are conserved among different mtDNAs in angiosperms (i.e. ~60 genes encoding different tRNAs, rRNAs, ribosomal proteins and subunits of the respiratory system). Yet, in addition to the known genes, plant mtDNAs also harbor numerous ORFs, most of which are not conserved among species and are currently of unknown function. Remarkably, and relevant to our study, CMS in plants is primarily associated with the expression of novel chimericORFs, which likely derive from recombination events within the mtDNAs. Whereas the CMS loci are localized to the mtDNAs, the factors that restore fertility (Rfs) are identified as nuclear-encoded RNA-binding proteins. Interestingly, nearly all of the Rf’s are identified as pentatricopeptide repeat (PPR) proteins, a large family of modular RNA-binding proteins that mediate several aspects of gene expression primarily in plant organelles. In this project we proposed to develop a system to test the ability of mtORFs in plants, which are closely related to known CMS factors. We will induce male fertility in various species of Brassicaceae, and test whether a down-relation in the expression of the recombinantCMS-genes restores fertility, using synthetically designed PPR proteins.
5
Levin, Ilan, John W. Scott, Moshe Lapidot, and Moshe Reuveni. Fine mapping, functional analysis and pyramiding of genes controlling begomovirus resistance in tomato. United States Department of Agriculture, November 2014. http://dx.doi.org/10.32747/2014.7594406.bard.
Full textAbstract:
Abstract. Tomato yellow leaf curl virus (TYLCV), a monopartitebegomovirus, is one of the most devastating viruses of cultivated tomatoes and poses increasing threat to tomato production worldwide. Because all accessions of the cultivated tomato are susceptible to these viruses, wild tomato species have become a valuable resource of resistance genes. QTL controlling resistance to TYLCV and other begomoviruses (Ty loci) were introgressed from several wild tomato species and mapped to the tomato genome. Additionally, a non-isogenic F₁diallel study demonstrated that several of these resistance sources may interact with each other, and in some cases generate hybrid plants displaying lower symptoms and higher fruit yield compared to their parental lines, while their respective resistance genes are not necessarily allelic. This suggests that pyramiding genes originating from different resistance sources can be effective in obtaining lines and cultivars which are highly resistant to begomoviruses. Molecular tools needed to test this hypothesis have been developed by our labs and can thus significantly improve our understanding of the mechanisms of begomovirus resistance and how to efficiently exploit them to develop wider and more durable resistance. Five non-allelic Ty loci with relatively major effects have been mapped to the tomato genome using molecular DNA markers, thereby establishing tools for efficient marker assisted selection, pyramiding of multiple genes, and map based gene cloning: Ty-1, Ty-2, Ty-3, Ty-4, and ty-5. This research focused on Ty-3 and Ty-4 due to their broad range of resistance to different begomoviruses, including ToMoV, and on ty-5 due to its exceptionally high level of resistance to TYLCV and other begomoviruses. Our aims were: (1) clone Ty-3, and fine map Ty-4 and Ty-5 genes, (2)introgress each gene into two backgroundsand develop semi isogenic lines harboring all possible combinations of the three genes while minimizing linkage-drag, (3) test the resulting lines, and F₁ hybrids made with them, for symptom severity and yield components, and (4) identify and functionally characterize candidate genes that map to chromosomal segments which harbor the resistance loci. During the course of this research we have: (1) found that the allelic Ty-1 and Ty-3 represent two alternative alleles of the gene coding DFDGD-RDRP; (2) found that ty-5is highly likely encoded by the messenger RNA surveillance factor PELOTA (validation is at progress with positive results); (3) continued the map-based cloning of Ty-4; (4) generated all possible gene combinations among Ty-1, Ty-3 and ty-5, including their F₁ counterparts, and tested them for TYLCV and ToMoV resistance; (5) found that the symptomless line TY172, carrying ty-5, also carries a novel allele of Ty-1 (termed Ty-1ⱽ). The main scientific and agricultural implications of this research are as follows: (1) We have developed recombination free DNA markers that will substantially facilitate the introgression of Ty-1, Ty-3 and ty-5 as well as their combinations; (2) We have identified the genes controlling TYLCV resistance at the Ty-1/Ty-3 and ty-5 loci, thus enabling an in-depth analyses of the mechanisms that facilitate begomovirus resistance; (3) Pyramiding of Ty resistance loci is highly effective in providing significantly higher TYLCV resistance.
6
Grumet, Rebecca, and Benjamin Raccah. Identification of Potyviral Domains Controlling Systemic Infection, Host Range and Aphid Transmission. United States Department of Agriculture, July 2000. http://dx.doi.org/10.32747/2000.7695842.bard.
Full textAbstract:
Potyviruses form one of the largest and most economically important groups of plant viruses. Individual potyviruses and their isolates vary in symptom expression, host range, and ability to overcome host resistance genes. Understanding factors influencing these biological characteristics is of agricultural importance for epidemiology and deployment of resistance strategies. Cucurbit crops are subject to severe losses by several potyviruses including the highly aggressive and variable zucchini yellow mosaic virus (ZYMV). In this project we sought to investigate protein domains in ZYMV that influence systemic infection and host range. Particular emphasis was on coat protein (CP), because of known functions in both cell to cell and long distance movement, and helper component-protease (HC-Pro), which has been implicated to play a role in symptom development and long distance movement. These two genes are also essential for aphid mediated transmission, and domains that influence disease development may also influence transmissibility. The objectives of the approved BARD project were to test roles of specific domains in the CP and HC-Pro by making sequence alterations or switches between different isolates and viruses, and testing for infectivity, host range, and aphid transmissibility. These objectives were largely achieved as described below. Finally, we also initiated new research to identify host factors interacting with potyviral proteins and demonstrated interaction between the ZYMV RNA dependent RNA polymerase and host poly-(A)-binding protein (Wang et al., in press). The focus of the CP studies (MSU) was to investigate the role of the highly variable amino terminus (NT) in host range determination and systemic infection. Hybrid ZYMV infectious clones were produced by substituting the CP-NT of ZYMV with either the CP-NT from watermelon mosaic virus (overlapping, but broader host range) or tobacco etch virus (TEV) (non- overlapping host range) (Grumet et al., 2000; Ullah ct al., in prep). Although both hybrid viruses initially established systemic infection, indicating that even the non-cucurbit adapted TEV CP-NT could facilitate long distance transport in cucurbits, after approximately 4-6, the plants inoculated with the TEV-CPNT hybrid exhibited a distinct recovery of reduced symptoms, virus titer, and virus specific protection against secondary infection. These results suggest that the plant recognizes the presence of the TEV CP-NT, which has not been adapted to infection of cucurbits, and initiates defense responses. The CP-NT also appears to play a role in naturally occurring resistance conferred by the zym locus in the cucumber line 'Dina-1'. Patterns of virus accumulation indicated that expression of resistance is developmentally controlled and is due to a block in virus movement. Switches between the core and NT domains of ZYMV-NAA (does not cause veinal chlorosis on 'Dina-1'), and ZYMV-Ct (causes veinal chlorosis), indicated that the resistance response likely involves interaction with the CP-NT (Ullah and Grumet, submitted). At the Volcani Center the main thrust was to identify domains in the HC-Pro that affect symptom expression or aphid transmissibility. From the data reported in the first and second year report and in the attached publications (Peng et al. 1998; Kadouri et al. 1998; Raccah et al. 2000: it was shown that: 1. The mutation from PTK to PAK resulted in milder symptoms of the virus on squash, 2. Two mutations, PAK and ATK, resulted in total loss of helper activity, 3. It was established for the first time that the PTK domain is involved in binding of the HC-Pro to the potyvirus particle, and 4. Some of these experiments required greater amount of HC-Pro, therefore a simpler and more efficient purification method was developed based on Ni2+ resin.
7
Dawson, William O., and Moshe Bar-Joseph. Creating an Ally from an Adversary: Genetic Manipulation of Citrus Tristeza. United States Department of Agriculture, January 2004. http://dx.doi.org/10.32747/2004.7586540.bard.
Full textAbstract:
Citrus is one of the major agricultural crops common to Israel and the United States, important in terms of nutrition, foreign exchange, and employment. The economy of both citrus industries have been chronically plagued by diseases caused by Citrus tristeza virus (CTV). The short term solution until virus-resistant plants can be used is the use of mild strain cross-protection. We are custom designing "ideal" protecting viruses to immunize trees against severe isolates of CTV by purposely inoculating existing endangered trees and new plantings to be propagated as infected (protected) citrus budwood. We crossed the substantial technological hurdles necessary to accomplish this task which included developing an infectious cDNA clone which allows in vitro manipulation of the virus and methods to then infect citrus plants. We created a series of hybrids between decline-inducing and mild CTV strains, tested them in protoplasts, and are amplifying them to inoculate citrus trees for evaluation and mapping of disease determinants. We also extended this developed technology to begin engineering transient expression vectors based on CTV as tools for genetic improvement of tree crops, in this case citrus. Because of the long periods between genetic transformation and the ultimate assay of mature tree characteristics, there is a great need for an effective system that allows the expression or suppression of target genes in fruiting plants. Virus-based vectors will greatly expedite progress in citrus genetic improvement. We characterized several components of the virus that provides necessary information for designing virus-based vectors. We characterized the requirements of the 3 ’-nontranslated replication promoter and two 3 ’-ORF subgenomic (sg) mRNA controller elements. We discovered a novel type of 5’-terminal sgRNAs and characterized the cis-acting control element that also functions as a strong promoter of a 3 ’-sgRNA. We showed that the p23 gene controls negative-stranded RNA synthesis and expression of 3 ’ genes. We identified which genes are required for infection of plants, which are host range determinants, and which are not needed for plant infection. We continued the characterization of native dRNA populations and showed the presence of five different classes including class III dRNAs that consists of infectious and self-replicating molecules and class V dRNAs that contain all of the 3 ’ ORFs, along with class IV dRNAs that retain non-contiguous internal sequences. We have constructed and tested in protoplasts a series of expression vectors that will be described in this proposal.