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Journal articles on the topic "TRNA Function"
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Mangroo, Dev, Xin-Qi Wu, and Uttam L. Rajbhandary. "Escherichia coliinitiator tRNA: structure–function relationships and interactions with the translational machinery." Biochemistry and Cell Biology 73, no. 11-12 (December 1, 1995): 1023–31. http://dx.doi.org/10.1139/o95-109.
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We showed previously that the sequence and (or) structural elements important for specifying the many distinctive properties of Escherichia coli initiator tRNA are clustered in the acceptor stem and in the anticodon stem and loop. This paper briefly describes this and reviews the results of some recently published studies on the mutant initiator tRNAs generated during this work. First, we have studied the effect of overproduction of methionyl-tRNA transformylase (MTF) and initiation factors IF2 and IF3 on activity of mutant initiator tRNAs mat are defective at specific steps in the initiation pathway. Overproduction of MTF rescued specifically the activity of mutant tRNAs defective in formylation but not mutants defective in binding to the P site. Overproduction of IF2 increased me activity of all mutant tRNAs having the CUA anticodon but not of mutant tRNA having me GAC anticodon. Overproduction of IF3 had no effect on the activity of any of me mutant tRNAs tested. Second, for functional studies of mutant initiator tRNA in vivo, we used a CAU→CUA anticodon sequence mutant mat can initiate protein synthesis from UAG instead of AUG. In contrast with me wild-type initiator tRNA, the mutant initiator tRNA has a 2-methylthio-N6-isopentenyl adenosine (ms2i6A) base modification next to the anticodon. Interestingly, this base modification is now important for activity of the mutant tRNA in initiation. In a miaA strain of E. coli deficient in biosynthesis of ms2i6A, the mutant initiator tRNA is much less active in initiation. The defect is specifically in binding to the ribosomal P site.Key words: initiator tRNA, initiation Factors, formylation, P site binding, base modification.
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Schaffer, Ashleigh E., Otis Pinkard, and Jeffery M. Coller. "tRNA Metabolism and Neurodevelopmental Disorders." Annual Review of Genomics and Human Genetics 20, no. 1 (August 31, 2019): 359–87. http://dx.doi.org/10.1146/annurev-genom-083118-015334.
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tRNAs are short noncoding RNAs required for protein translation. The human genome includes more than 600 putative tRNA genes, many of which are considered redundant. tRNA transcripts are subject to tightly controlled, multistep maturation processes that lead to the removal of flanking sequences and the addition of nontemplated nucleotides. Furthermore, tRNAs are highly structured and posttranscriptionally modified. Together, these unique features have impeded the adoption of modern genomics and transcriptomics technologies for tRNA studies. Nevertheless, it has become apparent from human neurogenetic research that many tRNA biogenesis proteins cause brain abnormalities and other neurological disorders when mutated. The cerebral cortex, cerebellum, and peripheral nervous system show defects, impairment, and degeneration upon tRNA misregulation, suggesting that they are particularly sensitive to changes in tRNA expression or function. An integrated approach to identify tRNA species and contextually characterize tRNA function will be imperative to drive future tool development and novel therapeutic design for tRNA-associated disorders.
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Francis, M. A., and U. L. Rajbhandary. "Expression and function of a human initiator tRNA gene in the yeast Saccharomyces cerevisiae." Molecular and Cellular Biology 10, no. 9 (September 1990): 4486–94. http://dx.doi.org/10.1128/mcb.10.9.4486-4494.1990.
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We showed previously that the human initiator tRNA gene, in the context of its own 5'- and 3'-flanking sequences, was not expressed in Saccharomyces cerevisiae. Here we show that switching its 5'-flanking sequence with that of a yeast arginine tRNA gene allows its functional expression in yeast cells. The human initiator tRNA coding sequence was either cloned downstream of the yeast arginine tRNA gene, with various lengths of intergenic spacer separating them, or linked directly to the 5'-flanking sequence of the yeast arginine tRNA coding sequence. The human initiator tRNA made in yeast cells can be aminoacylated with methionine, and it was clearly separated from the yeast initiator and elongator methionine tRNAs by RPC-5 column chromatography. It was also functional in yeast cells. Expression of the human initiator tRNA in transformants of a slow-growing mutant yeast strain, in which three of the four endogenous initiator tRNA genes had been inactivated by gene disruption, resulted in enhancement of the growth rate. The degree of growth rate enhancement correlated with the steady-state levels of human tRNA in the transformants. Besides providing a possible assay for in vivo function of mutant human initiator tRNAs, this work represents the only example of the functional expression of a vertebrate RNA polymerase III-transcribed gene in yeast cells.
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Francis, M. A., and U. L. Rajbhandary. "Expression and function of a human initiator tRNA gene in the yeast Saccharomyces cerevisiae." Molecular and Cellular Biology 10, no. 9 (September 1990): 4486–94. http://dx.doi.org/10.1128/mcb.10.9.4486.
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We showed previously that the human initiator tRNA gene, in the context of its own 5'- and 3'-flanking sequences, was not expressed in Saccharomyces cerevisiae. Here we show that switching its 5'-flanking sequence with that of a yeast arginine tRNA gene allows its functional expression in yeast cells. The human initiator tRNA coding sequence was either cloned downstream of the yeast arginine tRNA gene, with various lengths of intergenic spacer separating them, or linked directly to the 5'-flanking sequence of the yeast arginine tRNA coding sequence. The human initiator tRNA made in yeast cells can be aminoacylated with methionine, and it was clearly separated from the yeast initiator and elongator methionine tRNAs by RPC-5 column chromatography. It was also functional in yeast cells. Expression of the human initiator tRNA in transformants of a slow-growing mutant yeast strain, in which three of the four endogenous initiator tRNA genes had been inactivated by gene disruption, resulted in enhancement of the growth rate. The degree of growth rate enhancement correlated with the steady-state levels of human tRNA in the transformants. Besides providing a possible assay for in vivo function of mutant human initiator tRNAs, this work represents the only example of the functional expression of a vertebrate RNA polymerase III-transcribed gene in yeast cells.
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von Pawel-Rammingen, U., S. Aström, and A. S. Byström. "Mutational analysis of conserved positions potentially important for initiator tRNA function in Saccharomyces cerevisiae." Molecular and Cellular Biology 12, no. 4 (April 1992): 1432–42. http://dx.doi.org/10.1128/mcb.12.4.1432-1442.1992.
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The conserved positions of the eukaryotic cytoplasmic initiator tRNA have been suggested to be important for the initiation of protein synthesis. However, the role of these positions is not known. We describe in this report a functional analysis of the yeast initiator methionine tRNA (tRNA(iMet)), using a novel in vivo assay system which is not dependent on suppressor tRNAs. Strains of Saccharomyces cerevisiae with null alleles of the four initiator methionine tRNA (IMT) genes were constructed. Consequently, growth of these strains was dependent on tRNA(iMet) encoded from a plasmid-derived gene. We used these strains to investigate the significance of the conserved nucleosides of yeast tRNA(iMet) in vivo. Nucleotide substitutions corresponding to the nucleosides of the yeast elongator methionine tRNA (tRNA(MMet)) have been made at all conserved positions to identify the positions that are important for tRNA(iMet) to function in the initiation process. Surprisingly, nucleoside changes in base pairs 3-70, 12-23, 31-39, and 29-41, as well as expanding loop I by inserting an A at position 17 (A17) had no effect on the tester strain. Nucleotide substitutions in positions 54 and 60 to cytidines and guanosines (C54, G54, C60, and G60) did not prevent cell growth. In contrast, the double mutation U/rT54C60 blocked cell growth, and changing the A-U base pair 1-72 to a G-C base pair was deleterious to the cell, although these tRNAs were synthesized and accepted methionine in vitro. From our data, we suggest that an A-U base pair in position 1-72 is important for tRNA(iMet) function, that the hypothetical requirement for adenosines at positions 54 and 60 is invalid, and that a U/rT at position 54 is an antideterminant distinguishing an elongator from an initiator tRNA in the initiation of translation.
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von Pawel-Rammingen, U., S. Aström, and A. S. Byström. "Mutational analysis of conserved positions potentially important for initiator tRNA function in Saccharomyces cerevisiae." Molecular and Cellular Biology 12, no. 4 (April 1992): 1432–42. http://dx.doi.org/10.1128/mcb.12.4.1432.
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The conserved positions of the eukaryotic cytoplasmic initiator tRNA have been suggested to be important for the initiation of protein synthesis. However, the role of these positions is not known. We describe in this report a functional analysis of the yeast initiator methionine tRNA (tRNA(iMet)), using a novel in vivo assay system which is not dependent on suppressor tRNAs. Strains of Saccharomyces cerevisiae with null alleles of the four initiator methionine tRNA (IMT) genes were constructed. Consequently, growth of these strains was dependent on tRNA(iMet) encoded from a plasmid-derived gene. We used these strains to investigate the significance of the conserved nucleosides of yeast tRNA(iMet) in vivo. Nucleotide substitutions corresponding to the nucleosides of the yeast elongator methionine tRNA (tRNA(MMet)) have been made at all conserved positions to identify the positions that are important for tRNA(iMet) to function in the initiation process. Surprisingly, nucleoside changes in base pairs 3-70, 12-23, 31-39, and 29-41, as well as expanding loop I by inserting an A at position 17 (A17) had no effect on the tester strain. Nucleotide substitutions in positions 54 and 60 to cytidines and guanosines (C54, G54, C60, and G60) did not prevent cell growth. In contrast, the double mutation U/rT54C60 blocked cell growth, and changing the A-U base pair 1-72 to a G-C base pair was deleterious to the cell, although these tRNAs were synthesized and accepted methionine in vitro. From our data, we suggest that an A-U base pair in position 1-72 is important for tRNA(iMet) function, that the hypothetical requirement for adenosines at positions 54 and 60 is invalid, and that a U/rT at position 54 is an antideterminant distinguishing an elongator from an initiator tRNA in the initiation of translation.
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Strobel, M. C., and J. Abelson. "Effect of intron mutations on processing and function of Saccharomyces cerevisiae SUP53 tRNA in vitro and in vivo." Molecular and Cellular Biology 6, no. 7 (July 1986): 2663–73. http://dx.doi.org/10.1128/mcb.6.7.2663-2673.1986.
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The Saccharomyces cerevisiae leucine-inserting amber suppressor tRNA gene SUP53 (a tRNALeu3 allele) was used to investigate the relationship between precursor tRNA structure and mature tRNA function. This gene encodes a pre-tRNA which contains a 32-base intron. The mature tRNASUP53 contains a 5-methylcytosine modification of the anticodon wobble base. Mutations were made in the SUP53 intron. These mutant genes were transcribed in an S. cerevisiae nuclear extract preparation. In this extract, primary tRNA gene transcripts are end-processed and base modified after addition of cofactors. The base modifications made in vitro were examined, and the mutant pre-tRNAs were analyzed for their ability to serve as substrates for partially purified S. cerevisiae tRNA endonuclease and ligase. Finally, the suppressor function of these mutant tRNA genes was assayed after their integration into the S. cerevisiae genome. Mutant analysis showed that the totally intact precursor tRNA, rather than any specific sequence or structure of the intron, was necessary for efficient nonsense suppression by tRNASUP53. Less efficient suppressor activity correlated with the absence of the 5-methylcytosine modification. Most of the intron-altered precursor tRNAs were successfully spliced in vitro, indicating that modifications are not critical for recognition by the tRNA endonuclease and ligase.
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Strobel, M. C., and J. Abelson. "Effect of intron mutations on processing and function of Saccharomyces cerevisiae SUP53 tRNA in vitro and in vivo." Molecular and Cellular Biology 6, no. 7 (July 1986): 2663–73. http://dx.doi.org/10.1128/mcb.6.7.2663.
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The Saccharomyces cerevisiae leucine-inserting amber suppressor tRNA gene SUP53 (a tRNALeu3 allele) was used to investigate the relationship between precursor tRNA structure and mature tRNA function. This gene encodes a pre-tRNA which contains a 32-base intron. The mature tRNASUP53 contains a 5-methylcytosine modification of the anticodon wobble base. Mutations were made in the SUP53 intron. These mutant genes were transcribed in an S. cerevisiae nuclear extract preparation. In this extract, primary tRNA gene transcripts are end-processed and base modified after addition of cofactors. The base modifications made in vitro were examined, and the mutant pre-tRNAs were analyzed for their ability to serve as substrates for partially purified S. cerevisiae tRNA endonuclease and ligase. Finally, the suppressor function of these mutant tRNA genes was assayed after their integration into the S. cerevisiae genome. Mutant analysis showed that the totally intact precursor tRNA, rather than any specific sequence or structure of the intron, was necessary for efficient nonsense suppression by tRNASUP53. Less efficient suppressor activity correlated with the absence of the 5-methylcytosine modification. Most of the intron-altered precursor tRNAs were successfully spliced in vitro, indicating that modifications are not critical for recognition by the tRNA endonuclease and ligase.
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Oberbauer, Vera, and Matthias Schaefer. "tRNA-Derived Small RNAs: Biogenesis, Modification, Function and Potential Impact on Human Disease Development." Genes 9, no. 12 (December 5, 2018): 607. http://dx.doi.org/10.3390/genes9120607.
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Transfer RNAs (tRNAs) are abundant small non-coding RNAs that are crucially important for decoding genetic information. Besides fulfilling canonical roles as adaptor molecules during protein synthesis, tRNAs are also the source of a heterogeneous class of small RNAs, tRNA-derived small RNAs (tsRNAs). Occurrence and the relatively high abundance of tsRNAs has been noted in many high-throughput sequencing data sets, leading to largely correlative assumptions about their potential as biologically active entities. tRNAs are also the most modified RNAs in any cell type. Mutations in tRNA biogenesis factors including tRNA modification enzymes correlate with a variety of human disease syndromes. However, whether it is the lack of tRNAs or the activity of functionally relevant tsRNAs that are causative for human disease development remains to be elucidated. Here, we review the current knowledge in regard to tsRNAs biogenesis, including the impact of RNA modifications on tRNA stability and discuss the existing experimental evidence in support for the seemingly large functional spectrum being proposed for tsRNAs. We also argue that improved methodology allowing exact quantification and specific manipulation of tsRNAs will be necessary before developing these small RNAs into diagnostic biomarkers and when aiming to harness them for therapeutic purposes.
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Tucker, Jessica M., Aaron M. Schaller, Ian Willis, and Britt A. Glaunsinger. "Alteration of the Premature tRNA Landscape by Gammaherpesvirus Infection." mBio 11, no. 6 (December 15, 2020): e02664-20. http://dx.doi.org/10.1128/mbio.02664-20.
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ABSTRACTTransfer RNAs (tRNAs) are transcribed by RNA polymerase III (RNAPIII) and play a central role in decoding our genome, yet their expression and noncanonical function remain understudied. Many DNA tumor viruses enhance the activity of RNAPIII, yet whether infection alters tRNA expression is largely unknown. Here, we present the first genome-wide analysis of how viral infection alters the tRNAome. Using a tRNA-specific sequencing method (DM-tRNA-seq), we find that the murine gammaherpesvirus MHV68 induces global changes in premature tRNA (pre-tRNA) expression, with 14% of tRNA genes upregulated more than 3-fold, indicating that differential tRNA gene induction is a characteristic of DNA virus infection. Elevated pre-tRNA expression corresponds to increased RNAPIII occupancy for the subset of tRNA genes tested; additionally, posttranscriptional mechanisms contribute to the accumulation of pre-tRNA species. We find increased abundance of tRNA fragments derived from pre-tRNAs upregulated by viral infection, suggesting that noncanonical tRNA cleavage is also affected. Furthermore, pre-tRNA accumulation, but not RNAPIII recruitment, requires gammaherpesvirus-induced degradation of host mRNAs by the virally encoded mRNA endonuclease muSOX. We hypothesize that depletion of pre-tRNA maturation or turnover machinery contributes to robust accumulation of full-length pre-tRNAs in infected cells. Collectively, these findings reveal pervasive changes to tRNA expression during DNA virus infection and highlight the potential of using viruses to explore tRNA biology.IMPORTANCE Viral infection can dramatically change the gene expression landscape of the host cell, yet little is known regarding changes in noncoding gene transcription by RNA polymerase III (RNAPIII). Among these are transfer RNAs (tRNAs), which are fundamental in protein translation, yet whose gene regulatory features remain largely undefined in mammalian cells. Here, we perform the first genome-wide analysis of tRNA expression changes during viral infection. We show that premature tRNAs accumulate during infection with the model gammaherpesvirus MHV68 as a consequence of increased transcription, but that transcripts do not undergo canonical maturation into mature tRNAs. These findings underscore how tRNA expression is a highly regulated process, especially during conditions of elevated RNAPIII activity.
Dissertations / Theses on the topic "TRNA Function"
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Boonyalai, Nonlawat. "Lysyl-tRNA synthetase : structure-function studies." Thesis, Imperial College London, 2006. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.429379.
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Kelly, Paul Michael. "Characterizing the Function of Alanyl-tRNA Synthetase Activity in Microbial Translation." The Ohio State University, 2020. http://rave.ohiolink.edu/etdc/view?acc_num=osu1586259003806337.
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Dhungel, Nripesh. "tRNA Splicing Endonuclease: Novel and Essential Function Beyond tRNA Splicing and Subunit interactions." The Ohio State University, 2012. http://rave.ohiolink.edu/etdc/view?acc_num=osu1337968400.
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Swinehart, William E. Jr. "A Biochemical Investigation of Saccharomyces cerevisiae Trm10 and Implications of 1-methylguanosine for tRNA Structure and Function." The Ohio State University, 2015. http://rave.ohiolink.edu/etdc/view?acc_num=osu1429867956.
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Rogers, Theresa Elizabeth. "Elucidating the Function of a Pseudo-tRNA in Bacillus cereus." The Ohio State University, 2010. http://rave.ohiolink.edu/etdc/view?acc_num=osu1291213318.
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Faruggio, Dawn C. (Dawn Catherine) 1965. "Stucture-function relationships of human initiator tRNA mutants and attempted regulated expresion of tRNA genes in yeast." Thesis, Massachusetts Institute of Technology, 1995. http://hdl.handle.net/1721.1/32665.
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Picchioni, Daria. "Biological function of SLIMP, a mitochondrial seryl-tRNA synthetase paralog." Doctoral thesis, Universitat de Barcelona, 2014. http://hdl.handle.net/10803/283974.
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Our research group focuses on protein translation and more specifically on the mechanism of transfer RNA (tRNA) aminoacylation by a family of essential and universal enzymes called aminoacyl-tRNA synthetases (aaRSs). aaRS are essential and universal components of the genetic code and their long evolutionary history explains the growing number of functions being discovered for aaRS and for aaRS homologues, beyond their canonical role in gene translation.
During the process of constructing a model for human disorders caused by mitochondrial tRNA aminoacylation deficiencies in Drosophila melanogaster in the laboratory has been identified a previously uncharacterized paralog of a seryl-tRNA synthetase (SerRS) named SLIMP.
Given the conservation of SLIMP in all available insect genomes, a characterization of the phenotype upon SLIMP depletion was conducted in Drosophila melanogaster. We analyzed the specific effect of SLIMP knockdown in muscles and glia, and we observed a dramatic reduction of the lifespan in both mutants. Interestingly, SLIMP knockdown in the glia causes neurodegeneration visualized as vacuolization in the brain of mutant flies.
The sequence identity that SLIMP shares with SerRS allowed to predict that this protein has an identical fold of a seryl-tRNA synthetase. It was previously shown that SLIMP does not posses tRNA aminoacylation activity, but it retains the property to bind mitochondrial tRNASer isoacceptors as a possible reflection of the evolutionary origin of the protein. We characterized the RNA-and DNA-binding property of SLIMP through in vitro and in vivo methodologies. We found that SLIMP binds in vitro all RNA sequences that are encoded in the mitochondrial genome. We suggest that the composition of these sequences (AT enrichment) and the respective folding energy (low .G) are the main determinants for SLIMP binding to RNA. We performed ribonucleoprotein immunoprecipitation assay (RIP) in cells to study SLIMP-RNA interaction in vivo and we showed that SLIMP interact with almost all mitochondrial transcripts. Pull-down experiments combined to mass spectrometry analysis revealed at least two SLIMP protein interactors: SRS2 and LON protease. We demonstrated that SLIMP and SRS2 are interdependent, as the knockdown or the overexpression of one protein reduced or increase respectively the level of the other. We suggest that SLIMP and SRS2 might form a functional complex that is maintained at a given SLIMP:SRS2 ratio. LON was the other identified SLIMP interactor. Our data shows that LON and SRS2 do not interact, but SRS2 might be a substrate of LON proteolytic activity. We also found OPA1 as a potential interactor of LON. OPA1 is a dynamin-like GTPase that is responsible for inner membrane fusion. Our results indicated that the overexpression of LON protease results in an increase of the smaller OPA1 isoforms that correlates with mitochondrial stress. Given the proposed role of SLIMP in RNA binding, we aimed to determine whether SLIMP has an effect on mitochondrial transcription. The results obtained by Northern blot screening of some mitochondrial mRNAs revealed that knockdown of SLIMP significantly reduced the steady-state levels of COX2 and COX3 mRNA, and 12S and 16S rRNAs whereas tRNA levels were found constant. Our results suggest that the defect in transcription upon SLIMP depletion, might be limited to mature mRNAs and may reveal a specific function for SLIMP in the binding and stabilization of processed mitochondrial mRNAs rather than a role in the control of their transcriptional rate or processing. We showed that knockdown of SLIMP affects also cellular growth and cell cycle progression, in fact, upon SLIMP depletion, we observed a dramatic increase of cells in G2/M phase. This result suggests that the mitochondrial role of SLIMP, or a consequence of its function, may be acting as a crosstalk between mitochondria and nuclear transcription factors that regulate cell proliferation. Collectively, the work described in this thesis has contributed to the characterization of a mitochondrial seryl-tRNA synthetase paralog that has acquired an essential function in insects despite a relatively modest divergence from a canonical SerRS structure.El nostre grup de recerca es centra en la traducció de proteïnes i més específicament en el mecanisme d’aminoacilació dels àcids ribonucleics (ARNs) de transferència (ARNt) per una família d’enzims essencials i universals anomenats aminoacil-ARNt sintetases (aaRSs). Al laboratori s’han analitzat el paper de les aaRSs en la traducció proteica, les seves funcions no canòniques, la seva evolució, així com la seva implicació en malalties humanes. Les aaRSs són components universals i essencials del codi genètic. La seva llarga historia evolutiva explica el creixent número de funcions que s’estan descobrint, tant per a elles com per a proteïnes paràlogues, més enllà del seu paper canònic en traducció genètica. Al laboratori, durant el procés d’obtenció d’un model a Drosophila melanogaster per a l’estudi de malalties humanes degudes a deficiències en l’aminoacilació d’ARNt, es va identificar un nou gen, paràlog de la seril-ARNt sintetasa (SeRS) mitocondrial, anomenat SLIMP. La proteïna SLIMP representa un nou tipus de proteïna similar a aaRS que ha adquirit una funció essencial a insectes, tot i la relativament baixa divergència respecta a una estructura d’SeRS canònica. Tot i amb això, són necessaris estudis addicionals per a identificar el paper biològic de SLIMP. Per aconseguir aquesta fita, s’ha portat a terme el projecte descrit en aquest manuscrit, el qual consisteix en anàlisis addicionals del fenotip resultant de la depleció de SLIMP in vivo, seguits d’estudis detallats de les interaccions moleculars amb àcids nucleics i proteïnes, per acabar amb un estudi dels efectes de SLIMP en la fisiologia cel•lular. En conjunt, els nostres resultats demostren que SLIMP s’uneix específicament a ARNs mitocondrials in vivo i in vitro. SLIMP interacciona amb SerRS2 i les dues són interdependents a nivell d’estabilitat proteica. La depleció de SLIMP o de SerRS2 redueix els nivells basals d’alguns ARNm mitocondrials, però la transcripció d’ARNt és manté inalterada. Es proposa un rol en la regulació post-transcripcional o en l’estabilitat dels ARNm madurs. Hem observat també que la depleció de SLIMP indueix l’aturada del cicle cel•lular en la transició G2/M. Aquests resultats suggereixen que SLIMP, o una conseqüència de la seva funció, podria tenir un paper d’enllaç entre els mitocondris i els factors de transcripció nuclears que regulen la proliferació cel•lular.
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Cacan, Ercan. "Evolutionary synthetic biology: structure/function relationships within the protein translation system." Thesis, Georgia Institute of Technology, 2011. http://hdl.handle.net/1853/45838.
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Production of mutant biological molecules for understanding biological principles or as therapeutic agents has gained considerable interest recently. Synthetic genes are today being widely used for production of such molecules due to the substantial decrease in the costs associated with gene synthesis technology. Along one such line, we have engineered tRNA genes in order to dissect the effects of G:U base-pairs on the accuracy of the protein translation machinery. Our results provide greater detail into the thermodynamic interactions between tRNA molecules and an Elongation Factor protein (termed EF-Tu in bacteria and eEF1A in eukaryotes) and how these interactions influence the delivery of aminoacylated tRNAs to the ribosome. We anticipate that our studies not only shed light on the basic mechanisms of molecular machines but may also help us to develop therapeutic or novel proteins that contain unnatural amino acids. Further, the manipulation of the translation machinery holds promise for the development of new methods to understand the origins of life.
Along another line, we have used the power of synthetic biology to experimentally validate an evolutionary model. We exploited the functional diversity contained within the EF-Tu/eEF1A gene family to experimentally validate the model of evolution termed ‘heterotachy’. Heterotachy refers to a switch in a site’s mutational rate class. For instance, a site in a protein sequence may be invariant across all bacterial homologs while that same site may be highly variable across eukaryotic homologs. Such patterns imply that the selective constraints acting on this site differs between bacteria and eukaryotes. Despite intense efforts and large interest in understanding these patterns, no studies have experimentally validated these concepts until now. In the present study, we analyzed EF-Tu/eEF1A gene family members between bacteria and eukaryotes to identify heterotachous patterns (also called Type-I functional divergence). We applied statistical tests to identify sites possibly responsible for biomolecular functional divergence between EF-Tu and eEF1A. We then synthesized protein variants in the laboratory to validate our computational predictions. The results demonstrate for the first time that the identification of heterotachous sites can be specifically implicated in functional divergence among homologous proteins.
In total, this work supports an evolutionary synthetic biology paradigm that in one direction uses synthetic molecules to better understand the mechanisms and constraints governing biomolecular behavior while in another direction uses principles of molecular sequence evolution to generate novel biomolecules that have utility for industry and/or biomedicine.
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Mirando, Adam Christopher. "Characterization Of A Non-Canonical Function For Threonyl-Trna Synthetase In Angiogenesis." ScholarWorks @ UVM, 2015. http://scholarworks.uvm.edu/graddis/523.
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In addition to its canonical role in aminoacylation, threonyl-tRNA synthetase (TARS) possesses pro-angiogenic activity that is susceptible to the TARS-specific antibiotic borrelidin. However, the therapeutic benefit of borrelidin is offset by its strong toxicity to living cells. The removal of a single methylene group from the parent borrelidin generates BC194, a modified compound with significantly reduced toxicity but comparable anti-angiogenic potential. Biochemical analyses revealed that the difference in toxicities was due to borrelidin's stimulation of amino acid starvation at ten-fold lower concentrations than BC194. However, both compounds were found to inhibit in vitro and in vivo models of angiogenesis at sub-toxic concentrations, suggesting a similar mechanism that is distinct from the toxic responses. Crystal structures of TARS in complex with each compound indicated that the decreased contacts in the BC194 structure may render it more susceptible to competition with the canonical substrates and permit sufficient aminoacylation activity over a wider concentration of inhibitor. Conversely, both borrelidin and BC194 induce identical conformational changes in TARS, providing a rationale for their comparable effects on angiogenesis. The mechanisms of TARS and borrelidin-based compounds on angiogenesis were subsequently tested using zebrafish and cell-based models. These data revealed ectopic branching, non-functional vessels, and increased cell-cell contracts following BC194-treatment or knockdown of TARS expression, suggesting a role for the enzyme in the maturation and guidance of nascent vasculature. Using various TARS constructs this function was found to be dependent on two interactions or activities associated with the TARS enzyme that are distinct from its canonical aminoacylation activity. Furthermore, observations that TARS may influence VEGF expression and purinergic signaling suggest the possibility for a receptor-mediated response. Taken together, the results presented here demonstrate a clear role for TARS in angiogenesis, independent of its primary function in translation. Although the exact molecular mechanisms through which TARS and borrelidin regulate this activity remain to be determined, these data provide a foundation for future investigations of TARS's function in vascular biology and its use as a target for angiogenesis-based therapy.
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Kubicek, Charles E. 1981. "Identifying targets and function of the ubiquitin related modifier Urm1 in Saccharomyces cerevisiae." Thesis, University of Oregon, 2009. http://hdl.handle.net/1794/10310.
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xi, 81 p. : ill. A print copy of this thesis is available through the UO Libraries. Search the library catalog for the location and call number.Post-translational modification of proteins is an important cellular method of controlling various aspects of protein activity, including protein-protein interactions, half- life, and transport. An important class of post-translational modifications involves the ubiquitin family of proteins. In these modifications, a small protein, such as ubiquitin, is conjugated to a target protein through an isopeptide bond. Conjugation by a ubiquitin family member acts as a signal to regulate the activity, function, or stability of the target protein. Urm1, a ubiquitin-like protein conserved throughout all eukaryotes, was initially identified in S. cerevisiae. Loss of Urm1 leads to the disruption of a variety of cellular processes, including oxidative stress response, filamentous growth, and temperature sensitivity. This body of work comprises efforts to identify novel targets of Urm1, the mechanism by which Urm1 is attached to target proteins, and the physiological consequences of such conjugation. To gain understanding of the function and mechanism of Urm1 conjugation, the only known conjugate of Urm1, the peroxiredoxin reductase Ahp1, was examined in an effort to identify the site of modification on Ahp1 and to evaluate the physiological consequences of urmylation of Ahp1. I then completed a series of screens--a synthetic lethal screen, a two-hybrid screen, and a protein over-expression screen--to identify novel Urm1 conjugates and cellular functions dependent on Urm1. Of particular interest were genes identified in the synthetic lethal screen, namely PTC1, which encodes a protein phosphatase, and a set of genes encoding the Elongator complex, which functions in transcriptional elongation and tRNA modification. During this time period, other groups showed that thiolation of tRNAs depends on Urm1. Thus, Urm1 does not function only in protein conjugation, but also as a sulfur carrier in the thiolation of tRNA. Interestingly, I identified Elp2, a component of the Elongator complex, as a new Urm1-conjugate. Because Elp2 is also required for tRNA modification, perhaps Urm1 plays more than one role in tRNA modification. Loss of tRNA modification may disrupt many cellular functions and could explain the variety of urm1 mutant phenotypes. I have determined that all known Urm1 dependent processes are also associated with tRNA modification.
Committee in charge: Karen Guillemin, Chairperson, Biology; George Sprague, Advisor, Biology; Alice Barkan, Member, Biology; Kenneth Prehoda, Member, Chemistry; Tom Stevens, Outside Member, Chemistry
Books on the topic "TRNA Function"
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Dieter, Söll, and RajBhandary Uttam, eds. tRNA: Structure, biosynthesis, and function. Washington, D.C: ASM Press, 1995.
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Knoll, Franz, and Thomas Vogel. Design for Robustness. Zurich, Switzerland: International Association for Bridge and Structural Engineering (IABSE), 2009. http://dx.doi.org/10.2749/sed011.
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<p>Robustness is the ability to survive unforeseen circum-stances without undue damage or loss of function. It has become a requirement expressed in modern building codes, mostly without much advice as to how it can be achieved. Engineering has developed some approaches based on tra-ditional practice as well as recent insight. However, know-ledge about robustness remains scattered and ambiguous, making it difficult to apply to many specific cases.<p> The authors' attempt to collect and review elements, methods and strategies toward structural robustness, using a holistic, almost philosophical approach. This leads to a set of consid-erations to guide selection and implementation of measures in specific cases, followed by a collection of applications and examples from the authors practice.<p>The world, engineering and construction are imperfect and not entirely predictable. Robustness provides a measure of structural safety beyond traditional codified design rules.
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Soll, Dieter. Trna: Structure, Biosynthesis, and Function. ASM Press, 1995.
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Soll, Dieter, and Uttam L. RajBhandary. TRNA: Structure, Biosynthesis, and Function. Wiley & Sons, Limited, John, 2014.
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Divan, Aysha, and Janice A. Royds. 3. RNA. Oxford University Press, 2016. http://dx.doi.org/10.1093/actrade/9780198723882.003.0003.
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The first RNA molecules to be discovered were those involved in protein synthesis, mRNA, transfer RNA (tRNA), and ribosomal RNA (rRNA). In recent years, a vast number of additional RNA molecules have been identified. ‘RNA’ explains that these are non-coding RNAs that are not involved in protein synthesis, but influence many normal cellular and disease processes by regulating gene expression. RNA interference (RNAi) as one of the main ways in which gene expression is regulated is described with applications to therapy. Classes of RNA, including long non-coding RNAs and catalytic RNAs, are explained along with RNA techniques used to study RNA molecule and gene function.
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Shen, Wenyan *. Studies of the structure and function of "Bacillus subtilis" tRNAs in "Escherichia coli". 1991.
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Schreiner, Helen Carol. Replication control in broad-host-range plasmid RK2: Kil and kor functions affect expression of the essential replication gene trfA. 1988.
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Sedel, Frédéric, and Carla E. M. Hollak. Disorders of Thiamine Metabolism. Oxford University Press, 2016. http://dx.doi.org/10.1093/med/9780199972135.003.0028.
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Thiamine is a water-soluble vitamin acting in the mitochondria as a cofactor for energy metabolism and, in the cytoplasm, in the pentose phosphate biosynthetic pathway. Its transport through the plasma membrane requires two transporters with overlapping functions: THTR1 encoded by SLC19A2, and THTR2 encoded by SLC19A3. Thiamine is transformed into its active form, thiamine pyrophosphate (TPP) by a kinase encoded by the TPK1 gene. Then it may enter the mitochondria through a TPP transporter encoded by SLC25A19. Mutations in SLC19A2 cause thiamine-responsive megaloblastic anemia (TRMA). Mutations in SLC19A3 cause biotin/thiamine–responsive basal ganglia disease. Mutations in SLC25A19 may cause early microcephaly with death in infancy (also called Amish microcephaly) or a later-onset bilateral striatal necrosis with progressive peripheral neuropathy. Recently, mutations in the TPK1 gene have been associated with recurrent encephalopathy with mild lactic acidosis.
Book chapters on the topic "TRNA Function"
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Martin, Nancy C. "Organellar tRNAs: Biosynthesis and Function." In tRNA, 127–40. Washington, DC, USA: ASM Press, 2014. http://dx.doi.org/10.1128/9781555818333.ch9.
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Björk, Glenn R. "Biosynthesis and Function of Modified Nucleosides." In tRNA, 165–205. Washington, DC, USA: ASM Press, 2014. http://dx.doi.org/10.1128/9781555818333.ch11.
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Meinnel, Thierry, Yves Mechulam, and Sylvain Blanquet. "Aminoacyl-tRNA Synthetases: Occurrence, Structure, and Function." In tRNA, 251–92. Washington, DC, USA: ASM Press, 2014. http://dx.doi.org/10.1128/9781555818333.ch14.
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Baron, Christian, and August Böck. "The Selenocysteine-Inserting tRNA Species: Structure and Function." In tRNA, 529–44. Washington, DC, USA: ASM Press, 2014. http://dx.doi.org/10.1128/9781555818333.ch26.
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Sherman, Joyce M., M. John Rogers, and Dieter Söll. "Recognition in the Glutamine tRNA System: from Structure to Function." In tRNA, 395–409. Washington, DC, USA: ASM Press, 2014. http://dx.doi.org/10.1128/9781555818333.ch19.
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Schuster, Jens, and Mario Mörl. "Mitochondrial tRNA editing." In Mitochondrial Function and Biogenesis, 81–96. Berlin, Heidelberg: Springer Berlin Heidelberg, 2004. http://dx.doi.org/10.1007/b95713.
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Maréchal-Drouard, L., P. Guillemaut, H. Pfitzingzer, and J. H. Weil. "Chloroplast tRNAs and tRNA genes: structure and function." In The Translational Apparatus of Photosynthetic Organelles, 45–57. Berlin, Heidelberg: Springer Berlin Heidelberg, 1991. http://dx.doi.org/10.1007/978-3-642-75145-5_4.
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Suzuki, Tsutomu. "Biosynthesis and function of tRNA wobble modifications." In Fine-Tuning of RNA Functions by Modification and Editing, 23–69. Berlin, Heidelberg: Springer Berlin Heidelberg, 2005. http://dx.doi.org/10.1007/b106361.
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Alfonzo, Juan D. "Editing of tRNA for Structure and Function." In Nucleic Acids and Molecular Biology, 33–50. Berlin, Heidelberg: Springer Berlin Heidelberg, 2008. http://dx.doi.org/10.1007/978-3-540-73787-2_2.
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Shigi, Naoki. "Sulfur Modifications in tRNA: Function and Implications for Human Disease." In Modified Nucleic Acids in Biology and Medicine, 55–71. Cham: Springer International Publishing, 2016. http://dx.doi.org/10.1007/978-3-319-34175-0_3.
Full textConference papers on the topic "TRNA Function"
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Williams, Tamara, Adam Mirando, Christopher Francklyn, and Karen M. Lounsbury. "Abstract 3897: A novel function for threonyl-tRNA synthetase as a stimulator of angiogenesis and endothelial cell migration." 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.am2013-3897.
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Shigematsu, M., K. Pawar, T. Kawamura, D. A. Deshpande, and Y. Kirino. "Expression and Functional Role of tRNA-Derived Non-Coding RNAs in Asthma." In American Thoracic Society 2019 International Conference, May 17-22, 2019 - Dallas, TX. American Thoracic Society, 2019. http://dx.doi.org/10.1164/ajrccm-conference.2019.199.1_meetingabstracts.a6114.
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Honda, Shozo, Megumi Shigematsu, Phillipe Loher, Juan P. Palazzo, Issei Imoto, Isidore Rigoutsos, and Yohei Kirino. "Abstract 2665: SHOT-RNAs: A novel class of tRNA-derived functional RNAs expressed in hormone-dependent cancers." In Proceedings: AACR 107th Annual Meeting 2016; April 16-20, 2016; New Orleans, LA. American Association for Cancer Research, 2016. http://dx.doi.org/10.1158/1538-7445.am2016-2665.
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Yi Lee, Ching, Ching I Chen, and Meng-Cong Zheng. "User Experience of Taiwan Railway Ticket Vending Machine." In 13th International Conference on Applied Human Factors and Ergonomics (AHFE 2022). AHFE International, 2022. http://dx.doi.org/10.54941/ahfe1001699.
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Since its inception, the Taiwan Railway Ticket Vending Machine (TRA-TVM) has been used only by a few people because of its complex functions. Meanwhile, the Taiwan Railway ticket offices always have long queues. In light of this phenomenon, the Taiwan Railways Administration has released a new system in 2021. This study aims to determine whether the new system can meet users' needs to achieve an entirely self-service ticketing process.This study evaluated the usability, user psychology, and behavior of the TRA-TVM. Participants with different levels of self-efficacy (SE) were selected before the test. The sample consisted of 36 participants (16 males and 20 females) who didn't use the TRA-TVM before. In addition, the whole test was executed online. Conducted the evaluation as follows: (1) Filled out the questionnaires and finished the function sort of the TRA-TVM before the tasks to understand the participants' thoughts and past experiences. (2) Participants were asked to operate three tasks, one of which was related to the hardware interaction of the system. (3) At the end of each task, participants had a semi-structured interview to clarify how they operated during the task. (4) When finishing all the tasks, participants need to complete the QUIS and SUS questionnaires to indicate their evaluation of what they felt about this system.The results show a high error rate in the initial operation stage for the subjects when selecting the ticketing method and when customizing the station or the time of arrival and departure. The participants' performance showed that users made more errors in purchasing tickets than in operating other system functions and found that there is no absolute relationship between the level of SE and operational performance. Only in the "Ticket Refund(Q6)" function found a significant difference between the participants' SE and the number of errors. Those with high self-efficacy (HSE) had a significantly higher error rate for the refund function than those with low self-efficacy (LSE). In the semi-structured interview, HSE mentioned that they were unnoticed the feedback on the system and therefore were not sure how to operate it. Overall, the most often mentioned problem was "Confused," like the order of the ticketing process and the logic of the operation. For example, some participants felt that the function of "Seats Left(Q2)" shouldn't be included as a ticketing method when purchasing a ticket. Or the current operational logic made it impossible for participants to understand that they couldn't use "Express Ticket Purchase" or "Non-Reserved Trains" to purchase an unreserved ticket with a bicycle. In addition, the users' initial mental model may cause the user to make poor decisions and may even intensify or reduce the users' feelings during the process. Future optimization of the system should be prioritized at two stages, namely the selection of the ticketing method and the customizing station or time of arrival and departure. These findings could be used to improve the user experience in the future, allowing users to have a better experience during the ticketing process.
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Andreev, D. E., M. Terenin, S. E. Dmitriev, M. Niepmann, and I. N. Shatsky. "MOONLIGHTING FUNCTIONS OF GLYCYL-TRNA SYNTHETASE IN MAMMALIAN CELLS: THE ROLE IN THE TRANSLATION INITIATION ON THE IRES ELEMENTS OF ENTEROVIRUS RNAS." In Viruses: Discovering Big in Small. TORUS PRESS, 2019. http://dx.doi.org/10.30826/viruses-2019-13.
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Castilla, Andre Coutinho, Priscilla Koch Wagner, Jessicados Santos de Oliveira, Maria Fernanda Barbosa Wanderley, and Anthony Moreno Eigier. "Neuralmed trIA: Automated Screening of Urgent Findings on Head CT Scans." In XIII Congresso Paulista de Neurologia. Zeppelini Editorial e Comunicação, 2021. http://dx.doi.org/10.5327/1516-3180.342.
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Background:NeuralMed is a startup specialized in the development of artificial intelligence applications in medicine. Objectives: We developed a screening system for head computed tomography scans (HCT) that prioritizes patients with urgent pathologies. This study describes the development of the algorithm for detection and localization of intracranial hemorrhage. Design and setting: This is an observational study on HCT performed at hospitals in State of São Paulo. Methodology: The algorithm was built from 8432 HCT. We used unenhanced axial images after post-processing and normalization. The set is split into 80% for training, 10% for validation and 10% for testing. We used a MobileNet network pre-trained with ImageNet weights combined with a long short-term memory with categorical cross-entropy as loss function. The model’s outputs are hemorrhage, no findings, and other pathologies. A gradient class activation map was applied to identify and localize the hemorrhages. Results: Internal validation showed an area under the ROC curve (AUROC) of 96%, sensitivity of 87% and positive predictive value (PPV) of 96%. External validation was performed on 125 exams collected in a period after the training group obtaining an AUROC of 86%, sensitivity of 78% and PPV of 81%. Conclusion: After detecting an identifying the bleedings it is possible to order the patient queue prioritizing those most likely to have abnormalities and life-threatening situations. The algorithm also indicates the lesion location by showing the regions of the images that most activated the neural network.
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Kubisz, P., S. Brahimi, and S. Cronberg. "INFLUENCE OF L0MUSTIN ON SOME PLATELET FUNCTIONS IN VITRO." In XIth International Congress on Thrombosis and Haemostasis. Schattauer GmbH, 1987. http://dx.doi.org/10.1055/s-0038-1643443.
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Lomustin (CCNU) is a cytostatic drug that is used in the trea-tement of some forms of solid tumors and hemoblastosis. The most important form of toxicity associated with its use consists of thrombocytopenia often associated with hemorrhagic syndrome. Lomustin may exert its action on the formation of platelets, but it may also interfere with platelets themselves.In the present study its influence on the platelet function was investigated using a wide battery of tests.Platelet-rich plasma was incubated with lomustin (1,5 to 15 jjg/ml and 30 minutes at 37°C) in vitro.This inhibited strongly aggregation by ADP (2 × 10-6 M), adrenaline (2×10−6 M) and collagen (15 ¼g/ml). Platelet factor 4 release was almost completly inhibited independently of the inducer used. 14C-serotonin release was decreased by approximately 70% when incubation proceeded in the presence of 15 ¼g/ml of lomustin, and completly at higher concentration. Platelet factor 3 availability was also significantly impaired. Reptilase clot retraction was diminished, regardless of inducer used. In the presence of N-ethyl maleimide (1 mmol) significantLy lower malondialdehyde was produced in platelets incubated with lomustin, than in that from normal controls. After addition of arachidonic acid (1 mmol), the platelet synthesized Less thromboxane B2. Lomustin did not interfere with coagulation factors and did not induce overt fibrinolysis.Its is concluded that lomustin acts as an inhibitor of platelet functions and can induce an acquired thrombocytopathy.Thus impairement of platelet functions might play a part in hemorrhagic complications accompanying in some cases lomustin therapy.
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Xiao, Y., E. R. Hsieh, Steve S. Chung, T. R. Chen, S. A. Huang, T. J. Chen, and Osbert Cheng. "Novel Concept of the Transistor Variation Directed Toward the Circuit Implementation of Physical Unclonable Function (PUF) and True-random-number Generator (TRNG)." In 2019 IEEE International Electron Devices Meeting (IEDM). IEEE, 2019. http://dx.doi.org/10.1109/iedm19573.2019.8993496.
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Bang, K. H., and W. H. Choo. "Flow Boiling in Minichannels of Copper, Brass, and Aluminum Round Tubes." In ASME 2004 2nd International Conference on Microchannels and Minichannels. ASMEDC, 2004. http://dx.doi.org/10.1115/icmm2004-2381.
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The past work on flow boiling heat transfer in minichannels ranging one to three millimeters of hydraulic diameter has indicated that the local heat transfer coefficients are largely independent of mass flux and vapor quality, but mainly a function of wall heat flux. The present work is a revisit of flow boiling in minichannels by conducting experiment using 1.67 mm inner diameter tubes of three different materials; aluminum, brass, and copper, to investigate an effect of the tube inner surface conditions with the focus on an effect on nucleate boiling. Tests were conducted for R-22, a fixed mass flux of 600 kg/m2s, 5∼30 kW/m2 of wall heat flux, 0.0∼0.9 of local vapor quality. The present experimental data confirmed that the flow boiling heat transfer coefficient in a minichannel varies only by heat flux, independent of mass flux and vapor quality. The effect of tube material was found small for the tubes used in the present work. The present data were well predicted by the correlation proposed by Tran et al. (1996).
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Chen, I.-Chang, Shu-Keng Hsu, Teh-Juan Wu, Li-Hsien Yen, Yusin Lee, Dung-Ying Lin, Chuen-Yih Chen, Wei-Hsun Lee, and Guo-Wei Su. "RDSP: A Railway Decision Support Platform for Integrating and Bridging Existed Legacy Systems." In 2013 Joint Rail Conference. American Society of Mechanical Engineers, 2013. http://dx.doi.org/10.1115/jrc2013-2442.
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Railway system operation is a very complicated task and must be supported by the coordination of several systems including engineering, transportation, locomotive maintenance and management, ticket system and passengers service, etc. Ideally, a modern railway enterprise information system should be an integrated, consisted, and site-opened database to support railway system operation. However, multiple isolated applications instead of one integrated enterprise information system are often formed due to historical factors such as independent departments, budgetary constraint and application requirements diversification. Take Taiwan Railways Administration (TRA) as an example, four major departments have their own support systems and databases that store critical planning and operational data. Those systems are isolated and can hardly communicate with each other. As a result, most cross-systems information analyses or data reference for decision making are done manually, which significantly affect the organizational efficiency of TRA. To address the aforementioned issues, a railway decision support platform (RDSP) for integrating the legacy systems (databases) is proposed in this paper. RDSP is designed to be a railway enterprise information system that supports the critical functions of data warehouse and decision support. Furthermore, RDSP is built by integrating the existing legacy systems rather than by building it from scratch. A data bridging system (HDBS) including four modules are design and implemented, input module for connecting the external data sources, output module for exporting integrated report or dumping data by predefined criteria for other systems, configuration module with a web-based user interface for setting up the periodic operations of data input or output tasks, and DB connection module for connecting external databases. Various types of railway system data are designed in RDSP schema and collected, including facilities, timetable, train services records, tickets, centralized traffic control (CTC) system records, and automatic train protection (ATP) system records. RDSP provides a system framework to integrate many isolated island-style databases that currently exist in TRA, and can form a cross-enterprise database that serves as the primary and only data platform. To demonstrate the efficacy of the RDSP, a spatiotemporal ticket-selling analysis report, a train delay cause analysis report, and a timetable planning software (TrainWorld) are designed on top of it. In the future, RDSP will play a major supportive role in infrastructure maintenance, operations, decision support, and planning.
Reports on the topic "TRNA Function"
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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.