Academic literature on the topic 'Eukaryotic gene regulation'
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Journal articles on the topic "Eukaryotic gene regulation"
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Chin, Jason W. "Eukaryotic gene regulation." Chemistry & Biology 7, no. 1 (January 2000): R26. http://dx.doi.org/10.1016/s1074-5521(00)00071-5.
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Lindahl, G. "Gene Regulation: A Eukaryotic Perspective." International Journal of Biochemistry & Cell Biology 35, no. 1 (January 2003): 111–12. http://dx.doi.org/10.1016/s1357-2725(02)00174-7.
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Marsden, P. "Gene Regulation. A Eukaryotic Perspective." Biochemical Education 19, no. 1 (January 1991): 44–45. http://dx.doi.org/10.1016/0307-4412(91)90163-3.
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Goodbourn, Stephen. "Gene regulation: A eukaryotic perspective." Trends in Genetics 7, no. 10 (October 1991): 340. http://dx.doi.org/10.1016/0168-9525(91)90426-q.
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Mellor, Jane. "Gene regulation: A eukaryotic perspective." Trends in Biochemical Sciences 16 (January 1991): 482–83. http://dx.doi.org/10.1016/0968-0004(91)90186-y.
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Bonifer, Constanze. "Developmental regulation of eukaryotic gene loci." Trends in Genetics 16, no. 7 (July 2000): 310–15. http://dx.doi.org/10.1016/s0168-9525(00)02029-1.
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Nakayama, Koh, and Naoyuki Kataoka. "Regulation of Gene Expression under Hypoxic Conditions." International Journal of Molecular Sciences 20, no. 13 (July 3, 2019): 3278. http://dx.doi.org/10.3390/ijms20133278.
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Eukaryotes are often subjected to different kinds of stress. In order to adjust to such circumstances, eukaryotes activate stress–response pathways and regulate gene expression. Eukaryotic gene expression consists of many different steps, including transcription, RNA processing, RNA transport, and translation. In this review article, we focus on both transcriptional and post-transcriptional regulations of gene expression under hypoxic conditions. In the first part of the review, transcriptional regulations mediated by various transcription factors including Hypoxia-Inducible Factors (HIFs) are described. In the second part, we present RNA splicing regulations under hypoxic conditions, which are mediated by splicing factors and their kinases. This work summarizes and discusses the emerging studies of those two gene expression machineries under hypoxic conditions.
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Chen, Lin. "Combinatorial gene regulation by eukaryotic transcription factors." Current Opinion in Structural Biology 9, no. 1 (February 1999): 48–55. http://dx.doi.org/10.1016/s0959-440x(99)80007-4.
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Gagneux, P. "Gene Regulation: A Eukaryotic Perspective, 4th Edition." Journal of Heredity 94, no. 6 (November 1, 2003): 528–29. http://dx.doi.org/10.1093/jhered/esg102.
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Emery-Corbin, Samantha J., Joshua J. Hamey, Brendan R. E. Ansell, Balu Balan, Swapnil Tichkule, Andreas J. Stroehlein, Crystal Cooper, et al. "Eukaryote-Conserved Methylarginine Is Absent in Diplomonads and Functionally Compensated in Giardia." Molecular Biology and Evolution 37, no. 12 (July 23, 2020): 3525–49. http://dx.doi.org/10.1093/molbev/msaa186.
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Abstract Methylation is a common posttranslational modification of arginine and lysine in eukaryotic proteins. Methylproteomes are best characterized for higher eukaryotes, where they are functionally expanded and evolved complex regulation. However, this is not the case for protist species evolved from the earliest eukaryotic lineages. Here, we integrated bioinformatic, proteomic, and drug-screening data sets to comprehensively explore the methylproteome of Giardia duodenalis—a deeply branching parasitic protist. We demonstrate that Giardia and related diplomonads lack arginine-methyltransferases and have remodeled conserved RGG/RG motifs targeted by these enzymes. We also provide experimental evidence for methylarginine absence in proteomes of Giardia but readily detect methyllysine. We bioinformatically infer 11 lysine-methyltransferases in Giardia, including highly diverged Su(var)3-9, Enhancer-of-zeste and Trithorax proteins with reduced domain architectures, and novel annotations demonstrating conserved methyllysine regulation of eukaryotic elongation factor 1 alpha. Using mass spectrometry, we identify more than 200 methyllysine sites in Giardia, including in species-specific gene families involved in cytoskeletal regulation, enriched in coiled-coil features. Finally, we use known methylation inhibitors to show that methylation plays key roles in replication and cyst formation in this parasite. This study highlights reduced methylation enzymes, sites, and functions early in eukaryote evolution, including absent methylarginine networks in the Diplomonadida. These results challenge the view that arginine methylation is eukaryote conserved and demonstrate that functional compensation of methylarginine was possible preceding expansion and diversification of these key networks in higher eukaryotes.
Dissertations / Theses on the topic "Eukaryotic gene regulation"
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Kielbasa, Szymon M. "Bioinformatics of eukaryotic gene regulation." [S.l.] : [s.n.], 2006. http://deposit.ddb.de/cgi-bin/dokserv?idn=982693192.
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Kiełbasa, Szymon M. "Bioinformatics of eukaryotic gene regulation." Doctoral thesis, Humboldt-Universität zu Berlin, Mathematisch-Naturwissenschaftliche Fakultät I, 2006. http://dx.doi.org/10.18452/15562.
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Die Aufklärung der Mechanismen zur Kontrolle der Genexpression ist eines der wichtigsten Probleme der modernen Molekularbiologie. Detaillierte experimentelle Untersuchungen sind enorm aufwändig aufgrund der komplexen und kombinatorischen Wechselbeziehungen der beteiligten Moleküle. Infolgedessen sind bioinformatische Methoden unverzichtbar. Diese Dissertation stellt drei Methoden vor, die die Vorhersage der regulatorischen Elementen der Gentranskription verbessern. Der erste Ansatz findet Bindungsstellen, die von den Transkriptionsfaktoren erkannt werden. Dieser sucht statistisch überrepräsentierte kurze Motive in einer Menge von Promotersequenzen und wird erfolgreich auf das Genom der Bäckerhefe angewandt. Die Analyse der Genregulation in höheren Eukaryoten benötigt jedoch fortgeschrittenere Techniken. In verschiedenen Datenbanken liegen Hunderte von Profilen vor, die von den Transkriptionsfaktoren erkannt werden. Die Ähnlichkeit zwischen ihnen resultiert in mehrfachen Vorhersagen einer einzigen Bindestelle, was im nachhinein korrigiert werden muss. Es wird eine Methode vorgestellt, die eine Möglichkeit zur Reduktion der Anzahl von Profilen bietet, indem sie die Ähnlichkeiten zwischen ihnen identifiziert. Die komplexe Natur der Wechselbeziehung zwischen den Transkriptionsfaktoren macht jedoch die Vorhersage von Bindestellen schwierig. Auch mit einer Verringerung der zu suchenden Profile sind die Resultate der Vorhersagen noch immer stark fehlerbehafted. Die Zuhilfenahme der unabhängigen Informationsressourcen reduziert die Häufigkeit der Falschprognosen. Die dritte beschriebene Methode schlägt einen neuen Ansatz vor, die die Gen-Anotation mit der Regulierung von multiplen Transkriptionsfaktoren und den von ihnen erkannten Bindestellen assoziiert. Der Nutzen dieser Methode wird anhand von verschiedenen wohlbekannten Sätzen von Transkriptionsfaktoren demonstriert.Understanding the mechanisms which control gene expression is one of the fundamental problems of molecular biology. Detailed experimental studies of regulation are laborious due to the complex and combinatorial nature of interactions among involved molecules. Therefore, computational techniques are used to suggest candidate mechanisms for further investigation. This thesis presents three methods improving the predictions of regulation of gene transcription. The first approach finds binding sites recognized by a transcription factor based on statistical over-representation of short motifs in a set of promoter sequences. A succesful application of this method to several gene families of yeast is shown. More advanced techniques are needed for the analysis of gene regulation in higher eukaryotes. Hundreds of profiles recognized by transcription factors are provided by libraries. Dependencies between them result in multiple predictions of the same binding sites which need later to be filtered out. The second method presented here offers a way to reduce the number of profiles by identifying similarities between them. Still, the complex nature of interaction between transcription factors makes reliable predictions of binding sites difficult. Exploiting independent sources of information reduces the false predictions rate. The third method proposes a novel approach associating gene annotations with regulation of multiple transcription factors and binding sites recognized by them. The utility of the method is demonstrated on several well-known sets of transcription factors. RNA interference provides a way of efficient down-regulation of gene expression. Difficulties in predicting efficient siRNA sequences motivated the development of a library containing siRNA sequences and related experimental details described in the literature. This library, presented in the last chapter, is publicly available at http://www.human-sirna-database.net
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Webb, Sarah. "Structural analysis of eukaryotic gene regulation." Thesis, The University of Sydney, 2014. http://hdl.handle.net/2123/13487.
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This thesis presents data aimed at deepening our understanding of the mechanisms underlying eukaryotic gene regulation. A comprehensive understanding of these mechanisms should ultimately both allow insight into disease processes that arise from defects in gene regulatory circuits and might enable gene expression to be manipulated for application in health, agriculture and industry. The mechanisms that regulate gene expression include chromatin remodelling, post-translational modification and DNA methylation. These mechanisms are able to work together to ensure that the structure of chromatin is accessible to the machinery used for DNA based processes, such as transcription, and also to shield chromatin containing genes that remain unexpressed. These mechanisms are investigated using blood, or erythropoeisis, as the model system, with GATA1 and the NuRD complex being the elements of interest. The erythroid transcriptional factor GATA1 is subject to acetylation in two lysine rich regions, with acetylation in the second of these (at two specific lysine residues) being associated with increased chromatin occupancy as a result of a direct interaction with the bromodomain protein Brd3. As part of an effort to examine the structural and functional consequences of this interaction, it is necessary to produce site-specifically acetylated GATA1. The NuRD complex is a large multi-subunit complex involved in modulating chromatin structure to regulate gene expression. It has two activities, chromatin remodelling and histone deacetylation. Although many papers have been published on the complex, very little is available on the architecture. In order to gain a greater insight into the workings of the NuRD complex, negative stain transmission electron microscopy is used to determine the structure of a near-complete complex.
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Dickinson, P. "Fibronectin gene expression in higher eukaryotic cells." Thesis, University of Manchester, 1987. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.378322.
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Spies, Noah (Noah Walter Benjamin). "Cross-regulation and interaction between eukaryotic gene regulatory processes." Thesis, Massachusetts Institute of Technology, 2012. http://hdl.handle.net/1721.1/72637.
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Thesis (Ph. D.)--Massachusetts Institute of Technology, Dept. of Biology, 2012.This electronic version was submitted by the student author. The certified thesis is available in the Institute Archives and Special Collections.
Cataloged from student submitted PDF version of thesis.
Includes bibliographical references.
Regulation of genes is fundamental to all living processes and can be exerted at many sequential steps. We studied several eukaryotic gene regulatory mechanisms with an emphasis on understanding the interplay between regulatory processes on a genome-wide scale. Gene splicing involves the joining of exonic RNA stretches from within a precursor messenger RNA (mRNA). Splicing typically occurs co-transcriptionally as the pre-mRNA is being produced from the DNA. We explored the relationship between the chromatin state of the gene-encoding DNA and the splicing machinery. We found a marked enrichment for nucleosomes at exonic DNA in human T cells, as compared to surrounding introns, an effect mostly explained by the biased nucleotide content of exons. The use of nucleosome positioning information improved splicing simulation models, suggesting nucleosome positioning may help determine cellular splicing patterns. Additionally, we found several histone marks enriched or depleted at exons compared to the background nucleosome levels, indicative of a histone code for splicing. These results connect the chromatin regulation and mRNA splicing processes in a genome-wide fashion. Another pre-mRNA processing step is cleavage and polyadenylation, which determines the 30 end of the mature mRNA. We found that 3P-Seq was able to quantify the levels of 30 end isoforms, in addition to the method's previous use for annotating mRNA 30 ends. Using 3P-Seq and a transcriptional shutoff experiment in mouse fibroblasts, we investigated the e?effect of nuclear alternative 30 end formation on mRNA stability, typically regulated in the cytoplasm. In genes with multiple, tandem 30 untranslated regions (30 UTRs) produced by alternative cleavage and polyadenylation, we found the shorter UTRs were significantly more stable in general than the longer isoforms. This di?difference was in part explained by the loss of cis-regulatory motifs, such as microRNA targets and PUF-binding sites, between the proximal and distal isoforms. Finally, we characterized the small interfering RNAs (siRNAs) produced from heterochromatic, silenced genomic regions in fission yeast. We observed a considerable bias for siRNAs with a 5' U, and used this bias to infer patterns of siRNA biogenesis. Furthermore, comparisons with between wild-type and the Cid14 non-canonical poly(A) polymerase mutant demonstrated that the exosome, the nuclear surveillance and processing complex, is required for RNA homeostasis. In the absence of a fully functional exosome complex, siRNAs are produced to normal exosome targets, including ribosomal and transfer RNAs, indicating these processes may compete for substrates and underscoring the interconnectedness of gene regulatory systems.
by Noah Spies.
Ph.D.
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Sen, Rwik. "REGULATION OF EUKARYOTIC TRANSCRIPTIONAL ELONGATION AND ASSOCIATED DNA REPAIR." OpenSIUC, 2016. https://opensiuc.lib.siu.edu/dissertations/1205.
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Transcriptional elongation is a crucial step in eukaryotic gene regulation whose mis-regulation leads to cellular pathologies. This makes it quite imperative to aim for a better understanding of the processes regulating transcriptional elongation. An important process promoting the association of RNA Polymerase II (RNAPII) with the coding region of the active gene and hence transcriptional elongation is the monoubiquitination of histone H2B at lysine 123. A complex of an E2 conjugase, Rad6p, and an E3 ligase, Bre1p, is essential for this process. Consistent with the role of histone H2B monoubiquitination in promoting the association of RNAPII with the active gene, this process was found to be impaired in the absence of Rad6p or point mutation of lysine 123 to arginine (H2B-K123R). Intriguingly, the association of RNAPII with the coding region of the active gene was not impaired in the absence of Bre1p, even though Bre1p is essential for histone H2B monoubiquitination. However, deletion of Bre1p’s RING domain that is essential for histone H2B monoubiquitination led to an impaired RNAPII association with the active gene. This observation indicates a role of the non-RING domain of Bre1p in repressing the association of RNAPII with the active gene, resulting in no net decrease in RNAPII occupancy in the absence of Bre1p. Taken together, my results implicated both the stimulatory and repressive roles of the histone H2B ubiquitin ligase Bre1p in regulation of RNAPII association with the coding regions of active genes and hence transcriptional elongation. Interestingly, my work also revealed that for efficient transcriptional elongation by histone H2B monoubiquitination, its optimum level needs to be maintained by a proper balance between Rad6p-Bre1p-mediated ubiquitination and de-ubiquitination (DUB) by the DUB module of SAGA. It was found that Sus1p, a subunit of the DUB module, promotes transcriptional elongation, DNA repair and replication via regulation of histone H2B DUB. In addition to Rad6p- Bre1p and the DUB module, global level of histone H2B monoubiquitination is also critically regulated by Cdk9, a kinase essential for phosphorylation of the serine 2 residue in the C-terminal domain (CTD) of RNAPII, which promotes transcriptional elongation. Apart from serine phosphorylation, proline residues at RNAPII-CTD undergo isomerization by proline isomerases, which also regulate transcription. One of the proline isomerases, Rrd1p, has been previously implicated in transcription in response to rapamycin treatment. Based on this fact and Rrd1p’s known interaction with RNAPII-CTD, we predicted that Rrd1p might regulate transcription independently of rapamycin treatment. In agreement with this hypothesis, our work revealed Rrd1p’s role in facilitating transcription of both rapamycin responsive and non-responsive genes in the absence of rapamycin treatment. Consistently, the absence of Rrd1p led to an impaired nucleosomal disassembly at the active gene, which correlates with the role of Rrd1p in promoting transcription. This is because maintenance of proper nucleosomal dynamics is essential for efficient transcription. It is known that transcriptional elongation is facilitated by the regulation of nucleosomal dynamics via the histone chaperone, FACT. Efficient chromatin reassembly in the wake of elongating RNAPII contributing to the fidelity of transcription is promoted by FACT. Being evolutionarily conserved among eukaryotes, FACT is also known to regulate DNA replication and repair, apart from transcription. Intriguingly, FACT has been found to be upregulated in cancers while its downregulation leads to tumor cell death. However, the mechanism which fine-tunes FACT for normal cellular functions remained unknown. My studies revealed a novel mechanism of regulation of FACT by the ubiquitin-proteasome system in yeast. San1p, an E3 ligase involved in nuclear protein quality control, was found to associate with the active gene and regulate transcriptional elongation through its E3 ligase activity- mediated turnover of Spt16p component of FACT. This regulation was found to maintain optimum level of Spt16p/FACT to engage with the active gene for proper transcriptional elongation, DNA repair and replication. In spite of playing such crucial roles in gene regulation, it was not known how FACT is targeted to the active gene. We discovered that a direct physical interaction between FACT and Cet1p, the mRNA capping enzyme, targets FACT to the active gene independently of Cet1p’s mRNA capping activity. Such targeting of FACT to the active gene leads to the release of promoter proximally paused-RNAPII into transcriptional elongation. However, the progress of RNAPII along the active gene during transcriptional elongation is frequently impeded by various kinds of damages along the underlying template DNA. Even though some of these lesions are co-transcriptionally repaired, it was not known whether the repair of extremely toxic DNA double-strand breaks (DSBs) was coupled to transcription. My results showed that DSBs at the transcriptionally active state of a gene are repaired faster than at the inactive state but such repair was not mediated by a co-transcriptional recruitment of DSB repair factors. This observation is in contrast to other DNA repair pathways such as nucleotide excision repair (NER) where repair factors are co-transcriptionally recruited to the lesion containing DNA. In this regard, we found that an NER factor, Rad14p, co-transcriptionally associates with the active gene in the absence of DNA damage to promote transcription, which unraveled a new role of Rad14p in transcription in addition its established role in NER. In summary, my results provide significant novel insights into the regulation of transcriptional elongation and associated processes leading to better understanding of eukaryotic gene expression.
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Helder, Stephanie. "Investigations into RNA-binding proteins involved in eukaryotic gene regulation." Thesis, The University of Sydney, 2017. http://hdl.handle.net/2123/18597.
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The flood of RNA-related research in recent decades has revealed RNA to be a structurally and functionally diverse class of molecule, one that generates an intricate network of regulation that has been pivotal to the evolution of complex lifeforms. In order to elucidate how RNA achieves biological function through the formation of ribonucleoprotein (RNP) complexes, characterisation of RNA recognition by RNA-binding proteins (RBPs) is an essential step. The rules governing the interaction of RNA and RBPs have proved difficult to define, and in many instances, it is not understood how specificity is achieved. Knowledge of these rules is crucial to our understanding of RNA-related functions and their role in disease, and requires further in-depth characterisation of a wide variety of RNP complexes. The research in this Thesis details the RNA-binding behaviour of two reported RBPs. Firstly, the RNA-binding behaviour of the Drosophila transcription factor bicoid is investigated. For many years it has been believed that the bicoid homeodomain binds the 3′-UTR of the caudal mRNA transcript, yet no binding site or specificity determinants have been reported. The work here attempts to characterise this interaction. Further, other domains in the protein are examined with a view to understanding how biological specificity might be achieved. Secondly, characterisation of the RNA-binding behaviour of the heterodimeric pair of transcription elongation factors, Spt4 and Spt5, is reported. This heterodimer is known to be an important player in transcription and yet remarkably little is known about its function. In the present work, the AA-repeat RNA-binding properties of these proteins are investigated, and complex binding behaviour is reported. Overall, it is shown that the elucidation of RNA-binding activity by proteins is often not straightforward, requiring the application of multiple and increasingly sophisticated techniques if we are to grasp the underlying biology.
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To, Tsz-Leung. "Transcriptional bursting in eukaryotic gene regulation : molecular basis and functional consequences." Thesis, Massachusetts Institute of Technology, 2010. http://hdl.handle.net/1721.1/62062.
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Thesis (Ph. D.)--Massachusetts Institute of Technology, Dept. of Chemical Engineering, 2010.Cataloged from PDF version of thesis.
Includes bibliographical references.
Transcription of mRNA appears to occur in random, intermittent bursts in a large variety of organisms. The statistics of mRNA expression can be described by two parameters: the frequency at which bursts occur (burst frequency) and the average number of mRNA produced within each burst (burst size). The mean steady-state abundance of mRNA is the product of the burst size and burst frequency. Although the experimental evidence for bursty gene transcription is abundant, little is known about its origins and consequences. We utilize single-molecule mRNA imaging and simple stochastic kinetic models to probe and understand both the mechanistic details and functional responses of transcriptional bursting in budding yeast. At the molecular level, we show that gene-specific activators can control both burst size and burst frequency by differentially utilizing kinetically distinct promoter elements. We also recognize the importance of activator residence time and nucleosome positioning on bursting. This investigation exemplifies how we can exploit spontaneous fluctuations in gene expression to uncover the molecular mechanisms and kinetic pathways of transcriptional regulation. At the network level, we demonstrate the important phenotypic consequences of transcriptional bursting by showing how noise itself can generate a bimodal, all-or-none gene expression profile that switches spontaneously between the low and high expression states in a transcriptional positive-feedback loop. Such bimodality is a hallmark in decision-making circuitry within metabolic, developmental, and synthetic gene regulatory networks. Importantly, we prove that the bimodal responses observed in our system are not due to deterministic bistability, which is an often-stated necessary condition for allor- none responses in positive-feedback loops. By clarifying a common misconception, this investigation provides unique biological insights into the molecular components, pathways and mechanisms controlling a measured phenotype.
by Tsz-Leung To.
Ph.D.
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Ferdoush, Jannatul. "Regulation of nuclear phase of eukaryotic gene expression by ubiquitin-proteasome system." OpenSIUC, 2019. https://opensiuc.lib.siu.edu/dissertations/1751.
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Eukaryotic gene expression is a highly synchronized cellular process whose nuclear phase is comprised of transcription, and mRNA processing and export. Transcription can be further comprised of transcription initiation, and elongation. Regulation of transcription initiation, transcription elongation, and mRNA processing and export are crucial for normal cellular function, since misregulation of these processes are associated with various diseases including cancer. Many factors or proteins are associated with these cellular processes which are modulated by different regulatory processes to maintain normal cellular function. Ubiquitin-proteasome system (UPS) is one of the recently studied regulatory processes. Over the years, ubiquitin and 26S proteasome have emerged as important regulatory factors in coordination of transcription and coupled mRNA export. However, the mechanisms as to how the ubiquitin and 26S proteasome regulate transcription and coupled mRNA export have not been clearly elucidated. Therefore, my dissertation has focused on understanding the role of UPS in these important cellular processes: transcription initiation, transcription elongation and mRNA export. The results have shown the non-proteolytic role of 19S RP of 26S proteasome in regulation of transcriptional initiation of SAGA and TFIID-dependent PHO84 gene. It was found that 19S RP facilitates both SAGA- and NuA4-TFIID-dependent transcriptional initiations of PHO84 via increased recruitment of the coactivators SAGA and NuA4 HAT, which promote TFIID-independent and -dependent PIC formation in the presence and absence of an essential nutrient, Pi, in the growth media for transcriptional initiation, respectively. Next, our studies have uncovered the role of UPS in regulation of transcriptional elongation. It was found that E3 ubiquitin ligase, San1, mediated UPS regulation of transcription elongation factor, FACT is required for stimulating nucleosomal reassembly at the coding sequence of active genes for proper transcription elongation. We also found the interaction of FACT with another important transcription elongation factor, Paf1C via NTD (N-ter domain) of Cet1p (mRNA capping enzyme) to regulate transcription elongation.Subsequently, our results revealed a novel regulation of Paf1 component of Paf1C by UPS to regulate its abundance for proper cellular function. Transcription of genes could be blocked by DNA damage which can be repaired by transcription-coupled DNA repair (TCR) pathways. SUMOylation, another PTM (Post-translational modifications) like ubiquitination, is implicated in regulation of many DNA repair pathways including TCR, but it is not clearly understood how SUMOylation and associated enzymes are involved in regulation of such pathways. Here, we revealed the distinct role of SUMO ligases Siz1 and Siz2 in response to several DNA damaging agents such as UV, MMS (methyl methanesulfonate), HU (Hydroxyurea) and H2O2 (Hydrogen peroxide). Finally, we have extended our research works to understand the regulatory mechanisms of mRNA export by UPS. We found the interaction of TREX (Transcription/Export) component Sub2 with Mdm30 (F-box protein) for ubiquitination and proteasomal degradation of Sub2 in a transcription-dependent manner to regulate mRNA export. We also found the role CBC (Cap binding complex) in regulation of nuclear mRNA export. Collectively, the results of this study postulate a better understanding of regulation of transcription initiation, transcription elongation, and mRNA export by UPS.
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Zheng, Qun. "Analysis of the Caenorhabditis elegans rpc-1 gene." Diss., Columbia, Mo. : University of Missouri-Columbia, 2005. http://hdl.handle.net/10355/4129.
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Thesis (Ph.D.)--University of Missouri-Columbia, 2005.The entire dissertation/thesis text is included in the research.pdf file; the official abstract appears in the short.pdf file (which also appears in the research.pdf); a non-technical general description, or public abstract, appears in the public.pdf file. Title from title screen of research.pdf file viewed on (January 25, 2007) Vita. Includes bibliographical references.
Books on the topic "Eukaryotic gene regulation"
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Stephen, Goodbourn, ed. Eukaryotic gene transcription. Oxford: IRL Press at Oxford University Press, 1996.
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Latchman, David S. Gene regulation: A eukaryotic perspective. London: Unwin Hyman, 1990.
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Gene regulation: A eukaryotic perspective. 2nd ed. London: Chapman & Hall, 1995.
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Gene regulation: A eukaryotic perspective. 4th ed. Cheltenham: Nelson Thornes, 2002.
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Gene regulation: A eukaryotic perspective. 5th ed. New York: Taylor & Francis, 2006.
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Wingender, Edgar. Gene regulation in eukaryotes. Weinheim: VCH, 1993.
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R, Kinghorn James, ed. Gene structure in eukaryotic microbes. Oxford: Published for the Society for General Microbiology by IRL Press, 1987.
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Lawrence, Privalsky Martin, ed. Transcriptional corepressors: Mediators of eukaryotic gene repression. Berlin: Springer, 2001.
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Wajapeyee, Narendra, and Romi Gupta, eds. Eukaryotic Transcriptional and Post-Transcriptional Gene Expression Regulation. New York, NY: Springer New York, 2017. http://dx.doi.org/10.1007/978-1-4939-6518-2.
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A, Broda P. M., Oliver S. G. 1949-, and Sims P, eds. The eukaryotic genome: Organisation and regulation. Cambridge: Cambridge University Press, 1993.
Find full textBook chapters on the topic "Eukaryotic gene regulation"
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Gupta, Naba K., Mir F. Ahmad, Debopam Chakrabarti, and Nargis Nasrin. "Roles of Eukaryotic Initiation Factor 2 and Eukaryotic Initiation Factor 2 Ancillary Protein Factors in Eukaryotic Protein Synthesis Initiation." In Translational Regulation of Gene Expression, 287–334. Boston, MA: Springer US, 1987. http://dx.doi.org/10.1007/978-1-4684-5365-2_14.
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Gehrke, Lee. "Differential Translation of Eukaryotic Messenger RNAs." In Translational Regulation of Gene Expression, 367–78. Boston, MA: Springer US, 1987. http://dx.doi.org/10.1007/978-1-4684-5365-2_16.
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Durairaj, Geetha, Shivani Malik, and Sukesh R. Bhaumik. "Eukaryotic Gene Expression by RNA Polymerase II." In Gene Regulation, Epigenetics and Hormone Signaling, 1–28. Weinheim, Germany: Wiley-VCH Verlag GmbH & Co. KGaA, 2017. http://dx.doi.org/10.1002/9783527697274.ch1.
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Jacobson, K. Bruce. "Translational and Nontranslational Mechanisms of Regulation by Eukaryotic Suppressor Mutants." In Translational Regulation of Gene Expression, 379–96. Boston, MA: Springer US, 1987. http://dx.doi.org/10.1007/978-1-4684-5365-2_17.
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Kaspar, Roger L., David R. Morris, and Michael W. White. "Control of Ribosomal Protein Synthesis in Eukaryotic Cells." In Translational Regulation of Gene Expression 2, 335–48. Boston, MA: Springer US, 1993. http://dx.doi.org/10.1007/978-1-4615-2894-4_16.
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Laz, Thomas, John Clements, and Fred Sherman. "The Role of Messenger RNA Sequences and Structures in Eukaryotic Translation." In Translational Regulation of Gene Expression, 413–29. Boston, MA: Springer US, 1987. http://dx.doi.org/10.1007/978-1-4684-5365-2_19.
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Edery, Isaac, Jerry Pelletier, and Nahum Sonenberg. "Role of Eukaryotic Messenger RNA Cap-Binding Protein in Regulation of Translation." In Translational Regulation of Gene Expression, 335–66. Boston, MA: Springer US, 1987. http://dx.doi.org/10.1007/978-1-4684-5365-2_15.
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Walthers, Don, Alvin Go, and Linda J. Kenney. "Regulation of Porin Gene Expression by the Two-Component Regulatory System EnvZ/OmpR." In Bacterial and Eukaryotic Porins, 1–24. Weinheim, FRG: Wiley-VCH Verlag GmbH & Co. KGaA, 2005. http://dx.doi.org/10.1002/3527603875.ch1.
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Peltz, Stuart W., and Allan Jacobson. "Regulation of Eukaryotic Gene Expression at the Level of mRNA Stability: Emergence of General Principles." In Post-transcriptional Control of Gene Expression, 1–7. Berlin, Heidelberg: Springer Berlin Heidelberg, 1996. http://dx.doi.org/10.1007/978-3-642-60929-9_1.
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Varani, Gabriele, Peter Bayer, Paul Cole, Andres Ramos, and Luca Varani. "RNA Structure and RNA-Protein Recognition During Regulation of Eukaryotic Gene Expression." In RNA Biochemistry and Biotechnology, 195–216. Dordrecht: Springer Netherlands, 1999. http://dx.doi.org/10.1007/978-94-011-4485-8_15.
Full textConference papers on the topic "Eukaryotic gene regulation"
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Pannekok, H., A. J. Van Zonneveid, C. J. M. de vries, M. E. MacDonald, H. Veerman, and F. Blasi. "FUNCTIONAL PROPERTIES OF DELETION-MUTANTS OF TISSUE-TYPE PLASMINOGEN ACTIVATOR." In XIth International Congress on Thrombosis and Haemostasis. Schattauer GmbH, 1987. http://dx.doi.org/10.1055/s-0038-1643724.
Full textAbstract:
Over the past twenty-five years, genetic methods have generated a wealth of information on the regulation and the structure-function relationship of bacterial genes.These methods are based on the introduction of random mutations in a gene to alter its function. Subsequently, genetic techniques cure applied to localize the mutation, while the nature of the impairedfunction could be determined using biochemical methods. Classic examples of this approach is now considered to be the elucidation of the structure and function of genes, constituting the Escherichia coli lactose (lac) and tryptophan (trp) operons,and the detailed establishment of the structure and function of the repressor (lacl) of the lac operon. Recombinant DNA techniques and the development of appropriate expression systems have provided the means both to study structure and functionof eukaryotic (glyco-) proteins and to create defined mutations with a predestinedposition. The rationale for the construction of mutant genes should preferentiallyrely on detailed knowledge of the three-dimensional structure of the gene product.Elegant examples are the application of in vitro mutagenesis techniques to substitute amino-acid residues near the catalytic centre of subtilisin, a serine proteasefrom Bacillus species and to substituteanamino acid in the reactive site (i.e. Pi residue; methionine) of α-antitrypsin, a serine protease inhibitor. Such substitutions have resulted into mutant proteins which are less susceptible to oxidation and, in some cases, into mutant proteins with a higher specific activity than the wild-type protein.If no data are available on the ternary structure of a protein, other strategies have to be developed to construct intelligent mutants to study the relation between the structure and the function of a eukaryotic protein. At least for a number of gene families, the gene structure is thought to be created by "exon shuffling", an evolutionary recombinational process to insert an exon or a set of exons which specify an additional structural and/or functional domain into a pre-existing gene. Both the structure of the tissue-type plasminogen activator protein(t-PA) and the t-PA gene suggest that this gene has evolved as a result of exon shuffling. As put forward by Gilbert (Science 228 (1985) 823), the "acid test"to prove the validity of the exon shuffling theory is either to delete, insert or to substitute exon(s) (i.e. in the corresponding cDNA) and toassay the properties of the mutant proteins to demonstrate that an exon or a set of adjacent exons encode (s) an autonomousfunction. Indeed, by the construction of specific deletions in full-length t-PA cDNA and expression of mutant proteins intissue-culture cells, we have shown by this approach that exon 2 of thet-PA gene encodes the function required forsecretion, exon 4 encodes the "finger" domain involved in fibrin binding(presumably on undegraded fibrin) and the set of exons 8 and 9 specifies kringle 2, containing a lysine-binding sit(LBS) which interacts with carboxy-terminal lysines, generated in fibrin after plasmic digestion. Exons 10 through 14 encode the carboxy-ter-minal light chain of t-PA and harbor the catalytic centre of the molecule and represents the predominant "target site" for the fast-acting endothelial plasminogen activator inhibitor (PAI-1).As a follow-up of this genetic approach to construct deletion mutants of t-PA, we also created substitution mutants of t-PA. Different mutants were constructed to substitute cDNA encoding thelight chain of t-PA by cDNA encoding the B-chain of urokinase (u-PA), in order to demonstrate that autonomous structural and functional domains of eitherone of the separate molecules are able toexert their intrinsic properties in a different context (C.J.M. de Vries et al., this volume). The possibilities and the limitations of this approach to study the structure and the function of t-PA and of other components of the fibrinolytic process will be outlined.
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"Bacteriophages as vectors of gene transfer from prokaryotes to eukaryotes." In Bioinformatics of Genome Regulation and Structure/Systems Biology (BGRS/SB-2022) :. Institute of Cytology and Genetics, the Siberian Branch of the Russian Academy of Sciences, 2022. http://dx.doi.org/10.18699/sbb-2022-074.
Full textReports on the topic "Eukaryotic gene regulation"
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Chamovitz, Daniel, and Albrecht Von Arnim. Translational regulation and light signal transduction in plants: the link between eIF3 and the COP9 signalosome. United States Department of Agriculture, November 2006. http://dx.doi.org/10.32747/2006.7696515.bard.
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The COP9 signalosome (CSN) is an eight-subunit protein complex that is highly conserved among eukaryotes. Genetic analysis of the signalosome in the plant model species Arabidopsis thaliana has shown that the signalosome is a repressor of light dependent seedling development as mutant Arabidopsis seedlings that lack this complex develop in complete darkness as if exposed to light. These mutant plants die following the seedling stage, even when exposed to light, indicating that the COP9 signalosome also has a central role in the regulation of normal photomorphogenic development. The biochemical mode of action of the signalosome and its position in eukaryotic cell signaling pathways is a matter of controversy and ongoing investigation, and recent results place the CSN at the juncture of kinase signaling pathways and ubiquitin-mediated protein degradation. We have shown that one of the many CSN functions may relate to the regulation of translation through the interaction of the CSN with its related complex, eukaryotic initiation factor (eIF3). While we have established a physical connection between eIF3 subunits and CSN subunits, the physiological and developmental significance of this interaction is still unknown. In an effort to understand the biochemical activity of the signalosome, and its role in regulating translation, we originally proposed to dissect the contribution of "h" subunit of eIF3 (eIF3h) along the following specific aims: (i) Isolation and phenotypic characterization of an Arabidopsis loss-of-function allele for eIF3h from insertional mutagenesis libraries; (ii) Creation of designed gain and loss of function alleles for eIF3h on the basis of its nucleocytoplasmic distribution and its yeast-two-hybrid interactions with other eIF3 and signalosome partner proteins; (iii) Determining the contribution of eIF3h and its interaction with the signalosome by expressing specific mutants of eIF3h in the eIF3h- loss-of function background. During the course of the research, these goals were modified to include examining the genetic interaction between csn and eif3h mutations. More importantly, we extended our effort toward the genetic analysis of mutations in the eIF3e subunit, which also interacts with the CSN. Through the course of this research program we have made several critical scientific discoveries, all concerned with the apparent diametrically opposed roles of eIF3h and eIF3e. We showed that: 1) While eIF3e is essential for growth and development, eIF3h is not essential for growth or basal translation; 2) While eIF3e has a negative role in translational regulation, eIF3h is positively required for efficient translation of transcripts with complex 5' UTR sequences; 3) Over-accumulation of eIF3e and loss-of-function of eIF3h both lead to cop phenotypes in dark-grown seedlings. These results were published in one publication (Kim et al., Plant Cell 2004) and in a second manuscript currently in revision for Embo J. Are results have led to a paradigm shift in translation research – eIF3 is now viewed in all systems as a dynamic entity that contains regulatory subuits that affect translational efficiency. In the long-term agronomic outlook, the proposed research has implications that may be far reaching. Many important plant processes, including developmental and physiological responses to light, abiotic stress, photosynthate, and hormones operate in part by modulating protein translation [23, 24, 40, 75]. Translational regulation is slowly coming of age as a mechanism for regulating foreign gene expression in plants, beginning with translational enhancers [84, 85] and more recently, coordinating the expression of multiple transgenes using internal ribosome entry sites. Our contribution to understanding the molecular mode of action of a protein complex as fundamental as eIF3 is likely to lead to advances that will be applicable in the foreseeable future.
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Coplin, David, Isaac Barash, and Shulamit Manulis. Role of Proteins Secreted by the Hrp-Pathways of Erwinia stewartii and E. herbicola pv. gypsophilae in Eliciting Water-Soaking Symptoms and Initiating Galls. United States Department of Agriculture, June 2001. http://dx.doi.org/10.32747/2001.7580675.bard.
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Many bacterial pathogens of plants can inject pathogenicity proteins into host cells using a specialized type III secretion system encoded by hrpgenes. This system deliver effector proteins, into plant cells that function in both susceptible and resistant interactions. We have found that the virulence of Erwinia stewartii(Es; syn. Pantoea stewartii) and Erwinia herbicola pv. gypsophilae (Ehg, syn. Pantoea agglomerans), which cause Stewart's wilt of corn and galls on Gypsophila, respectively, depends on hrpgenes. The major objectives of this project were: To increase expression of hrpgenes in order to identify secreted proteins; to identify genes for proteins secreted by the type-III systems and determine if they are required for pathogenicity; and to determine if the secreted proteins can function within eukaryotic cells. We found that transcription of the hrp and effector genes in Es and Ehg is controlled by at least four genes that constitute a regulatory cascade. Environmental and/or physiological signaling appears to be mediated by the HrpX/HrpY two component system, with HrpX functioning as a sensor-kinase and HrpY as a response regulator. HrpYupregulateshrpS, which encodes a transcriptional enhancer. HrpS then activates hrpL, which encodes an alternate sigma factor that recognizes "hrp boxes". All of the regulatory genes are essential for pathogenicity, except HrpX, which appears only to be required for induction of the HR in tobacco by Es. In elucidating this regulatory pathway in both species, we made a number of significant new discoveries. HrpX is unusual for a sensor-kinase because it is cytoplasmic and contains PAS domains, which may sense the redox state of the bacterium. In Es, a novel methyl-accepting protein may function upstream of hrpY and repress hrp gene expression in planta. The esaIR quorum sensing system in Es represses hrp gene expression in Es in response to cell-density. We have discovered six new type III effector proteins in these species, one of which (DspE in Ehg and WtsE in Es) is common to both pathogens. In addition, Es wtsG, which is a homolog of an avrPpiB from P. syringae pv. pisi, and an Ehg ORF, which is a homolog of P. syringae pv. phaseolicola AvrPphD, were both demonstrated to encode virulence proteins. Two plasmidborne, Ehg Hop proteins, HsvG and PthG, are required for infection of gypsophilia, but interestingly, PthG also acts as an Avr elicitor in beets. Using a calmodulin-dependent adenylate cyclase (cyaA) reporter gene, we were successful in demonstrating that an HsvG-CyaA fusion protein can be transferred into human HeLa cells by the type-III system of enteropathogenic E. coli. This is a highly significant accomplishment because it is the first direct demonstration that an effector protein from a plant pathogenic bacterium is capable of being translocated into a eukaryotic cell by a type-III secretion system. Ehg is considered a limiting factor in Gypsophila production in Israel and Stewart’s Wilt is a serious disease in the Eastern and North Central USA, especially on sweet corn in epidemic years. We believe that our basic research on the characterization of type III virulence effectors should enable future identification of their receptors in plant cells. This may lead to novel approaches for genetically engineering resistant plants by modifying their receptors or inactivating effectors and thus blocking the induction of the susceptible response. Alternatively, hrp gene regulation might also provide a target for plant produced compounds that interfere with recognition of the host by the pathogen. Such strategies would be broadly applicable to a wide range of serious bacterial diseases on many crops throughout the USA and Israel.
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Schuster, Gadi, and David Stern. Integration of phosphorus and chloroplast mRNA metabolism through regulated ribonucleases. United States Department of Agriculture, August 2008. http://dx.doi.org/10.32747/2008.7695859.bard.
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New potential for engineering chloroplasts to express novel traits has stimulated research into relevant techniques and genetic processes, including plastid transformation and gene regulation. This proposal continued our long time BARD-funded collaboration research into mechanisms that influence chloroplast RNA accumulation, and thus gene expression. Previous work on cpRNA catabolism has elucidated a pathway initiated by endonucleolytic cleavage, followed by polyadenylation and exonucleolytic degradation. A major player in this process is the nucleus-encoded exoribonuclease/polymerasepolynucleotidephoshorylase (PNPase). Biochemical characterization of PNPase has revealed a modular structure that controls its RNA synthesis and degradation activities, which in turn are responsive to the phosphate (P) concentration. However, the in vivo roles and regulation of these opposing activities are poorly understood. The objectives of this project were to define how PNPase is controlled by P and nucleotides, using in vitro assays; To make use of both null and site-directed mutations in the PNPgene to study why PNPase appears to be required for photosynthesis; and to analyze plants defective in P sensing for effects on chloroplast gene expression, to address one aspect of how adaptation is integrated throughout the organism. Our new data show that P deprivation reduces cpRNA decay rates in vivo in a PNPasedependent manner, suggesting that PNPase is part of an organismal P limitation response chain that includes the chloroplast. As an essential component of macromolecules, P availability often limits plant growth, and particularly impacts photosynthesis. Although plants have evolved sophisticated scavenging mechanisms these have yet to be exploited, hence P is the most important fertilizer input for crop plants. cpRNA metabolism was found to be regulated by P concentrations through a global sensing pathway in which PNPase is a central player. In addition several additional discoveries were revealed during the course of this research program. The human mitochondria PNPase was explored and a possible role in maintaining mitochondria homeostasis was outlined. As polyadenylation was found to be a common mechanism that is present in almost all organisms, the few examples of organisms that metabolize RNA with no polyadenylation were analyzed and described. Our experiment shaded new insights into how nutrient stress signals affect yield by influencing photosynthesis and other chloroplast processes, suggesting strategies for improving agriculturally-important plants or plants with novel introduced traits. Our studies illuminated the poorly understood linkage of chloroplast gene expression to environmental influences other than light quality and quantity. Finely, our finding significantly advanced the knowledge about polyadenylation of RNA, the evolution of this process and its function in different organisms including bacteria, archaea, chloroplasts, mitochondria and the eukaryotic cell. These new insights into chloroplast gene regulation will ultimately support plant improvement for agriculture
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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.
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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.
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Schuster, Gadi, and David Stern. Integrated Studies of Chloroplast Ribonucleases. United States Department of Agriculture, September 2011. http://dx.doi.org/10.32747/2011.7697125.bard.
Full textAbstract:
Gene regulation at the RNA level encompasses multiple mechanisms in prokaryotes and eukaryotes, including splicing, editing, endo- and exonucleolytic cleavage, and various phenomena related to small or interfering RNAs. Ribonucleases are key players in nearly all of these post-transcriptional mechanisms, as the catalytic agents. This proposal continued BARD-funded research into ribonuclease activities in the chloroplast, where RNase mutation or deficiency can cause metabolic defects and is often associated with plant chlorosis, embryo or seedling lethality, and/or failure to tolerate nutrient stress. The first objective of this proposal was to examined a series of point mutations in the PNPase enzyme of Arabidopsis both in vivo and in vitro. This goal is related to structure-function analysis of an enzyme whose importance in many cellular processes in prokaryotes and eukaryotes has only begun to be uncovered. PNPase substrates are mostly generated by endonucleolytic cleavages for which the catalytic enzymes remain poorly described. The second objective of the proposal was to examine two candidate enzymes, RNase E and RNase J. RNase E is well-described in bacteria but its function in plants was still unknown. We hypothesized it catalyzes endonucleolytic cleavages in both RNA maturation and decay. RNase J was recently discovered in bacteria but like RNase E, its function in plants had yet to be explored. The results of this work are described in the scientific manuscripts attached to this report. We have completed the first objective of characterizing in detail TILLING mutants of PNPase Arabidopsis plants and in parallel introducing the same amino acids changes in the protein and characterize the properties of the modified proteins in vitro. This study defined the roles for both RNase PH core domains in polyadenylation, RNA 3’-end maturation and intron degradation. The results are described in the collaborative scientific manuscript (Germain et al 2011). The second part of the project aimed at the characterization of the two endoribonucleases, RNase E and RNase J, also in this case, in vivo and in vitro. Our results described the limited role of RNase E as compared to the pronounced one of RNase J in the elimination of antisense transcripts in the chloroplast (Schein et al 2008; Sharwood et al 2011). In addition, we characterized polyadenylation in the chloroplast of the green alga Chlamydomonas reinhardtii, and in Arabidopsis (Zimmer et al 2009). Our long term collaboration enabling in vivo and in vitro analysis, capturing the expertise of the two collaborating laboratories, has resulted in a biologically significant correlation of biochemical and in planta results for conserved and indispensable ribonucleases. These new insights into chloroplast gene regulation will ultimately support plant improvement for agriculture.
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Tzfira, Tzvi, Michael Elbaum, and Sharon Wolf. DNA transfer by Agrobacterium: a cooperative interaction of ssDNA, virulence proteins, and plant host factors. United States Department of Agriculture, December 2005. http://dx.doi.org/10.32747/2005.7695881.bard.
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Agrobacteriumtumefaciensmediates genetic transformation of plants. The possibility of exchanging the natural genes for other DNA has led to Agrobacterium’s emergence as the primary vector for genetic modification of plants. The similarity among eukaryotic mechanisms of nuclear import also suggests use of its active elements as media for non-viral genetic therapy in animals. These considerations motivate the present study of the process that carries DNA of bacterial origin into the host nucleus. The infective pathway of Agrobacterium involves excision of a single-stranded DNA molecule (T-strand) from the bacterial tumor-inducing plasmid. This transferred DNA (T-DNA) travels to the host cell cytoplasm along with two virulence proteins, VirD2 and VirE2, through a specific bacteriumplant channel(s). Little is known about the precise structure and composition of the resulting complex within the host cell and even less is known about the mechanism of its nuclear import and integration into the host cell genome. In the present proposal we combined the expertise of the US and Israeli labs and revealed many of the biophysical and biological properties of the genetic transformation process, thus enhancing our understanding of the processes leading to nuclear import and integration of the Agrobacterium T-DNA. Specifically, we sought to: I. Elucidate the interaction of the T-strand with its chaperones. II. Analyzing the three-dimensional structure of the T-complex and its chaperones in vitro. III. Analyze kinetics of T-complex formation and T-complex nuclear import. During the past three years we accomplished our goals and made the following major discoveries: (1) Resolved the VirE2-ssDNA three-dimensional structure. (2) Characterized VirE2-ssDNA assembly and aggregation, along with regulation by VirE1. (3) Studied VirE2-ssDNA nuclear import by electron tomography. (4) Showed that T-DNA integrates via double-stranded (ds) intermediates. (5) Identified that Arabidopsis Ku80 interacts with dsT-DNA intermediates and is essential for T-DNA integration. (6) Found a role of targeted proteolysis in T-DNA uncoating. Our research provide significant physical, molecular, and structural insights into the Tcomplex structure and composition, the effect of host receptors on its nuclear import, the mechanism of T-DNA nuclear import, proteolysis and integration in host cells. Understanding the mechanical and molecular basis for T-DNA nuclear import and integration is an essential key for the development of new strategies for genetic transformation of recalcitrant plant species. Thus, the knowledge gained in this study can potentially be applied to enhance the transformation process by interfering with key steps of the transformation process (i.e. nuclear import, proteolysis and integration). Finally, in addition to the study of Agrobacterium-host interaction, our research also revealed some fundamental insights into basic cellular mechanisms of nuclear import, targeted proteolysis, protein-DNA interactions and DNA repair.