Demonstration of translation elongation factor 3 activity from a non-fungal species, Phytophthora infestans

Mateyak, Maria K.; Pupek, Justyna K.; Garino, Alexandra E.; Knapp, McCllelan C.; Colmer, Sarah; Kinzy, Terri Goss; Dunaway, Stephen

doi:10.1371/journal.pone.0190524

Cited by 18 publications

(18 citation statements)

References 30 publications

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“…The conserved nuclear gene EF1α , a G-protein that binds and recruits aa-tRNAs to the A-site of the ribosome, has been valuable as a higher-level phylogenetic marker in insects and it has also been widely used for stable reference gene 44,45 . For example, EF1α was the relatively stable gene for developmental stages and photoperiods in Harmonia axyridis 33 , for cuticle in S .…”

Section: Discussionmentioning

confidence: 99%

Evaluation and Validation of Reference Genes for Quantitative Real-Time PCR in Helopeltis theivora Waterhouse (Hemiptera: Miridae)

Zheng

Meng

Zhu

et al. 2019

Sci Rep

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Helopeltis theivora Waterhouse is a predominant sucking pest in many tropic economic crops, such as tea, cocoa and coffee. Quantitative real-time PCR (qRT-PCR) is one of the most powerful tools to analyze the gene expression level and investigate the mechanism of insect physiology at transcriptional level. Gene expression studies utilizing qRT-PCR have been applied to numerous insects so far. However, no universal reference genes could be used for H. theivora. To obtain accurate and reliable normalized data in H. theivora, twelve candidate reference genes were examined under different tissues, developmental stages and sexes by using geNorm, NormFinder, BestKeeper, Delta Ct and RefFinder algorithms, respectively. The results revealed that the ideal reference genes differed across the treatments, and the consensus rankings generated from stability values provided by these programs suggested a combination of two genes for normalization. To be specific, RPS3A and Actin were the best suitable reference genes for tissues, RPL13A and GAPDH were suitable for developmental stages, EF1α and RPL13A were suitable for sexes, and RPL13A and RPS3A were suitable for all samples. This study represents the first systematic analysis of reference genes for qRT-PCR experiments in H. theivora, and the results can provide a credible normalization for qRT-PCR data, facilitating transcript profiling studies of functional genes in this insect.

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Section: Discussionmentioning

confidence: 99%

Evaluation and Validation of Reference Genes for Quantitative Real-Time PCR in Helopeltis theivora Waterhouse (Hemiptera: Miridae)

Zheng

Meng

Zhu

et al. 2019

Sci Rep

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“…the elongation factor eEF2/EF-G 1 – and some are lineage-specific, such as elongation factor 3, eEF3, a member of the ABCF ATPase family 2,3 . While initial analysis of eEF3 distribution suggested it a fungi-specific translational factor 4 , its distribution is broader, with eEF3-like homologues found in non-fungal species, such as oomycete Phytophthora infestans 5 , choanoflagellates, and various distantly related algae 3 . The protein is essential both for the viability of Saccharomyces cerevisiae 6 and for peptide elongation in a reconstituted yeast translational system 7,8 .…”

Section: Introductionmentioning

confidence: 99%

Ribosome profiling analysis of eEF3-depletedSaccharomyces cerevisiae

Kasari

Margus

Atkinson

et al. 2018

Preprint

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In addition to the standard set of translation factors common in eukaryotic organisms, protein synthesis in the yeast Saccharomyces cerevisiae requires an ABCF ATPase factor eEF3, eukaryotic Elongation Factor 3. eEF3 is an E-site binder that was originally identified as an essential factor involved in the elongation stage of protein synthesis. Recent biochemical experiments suggest an additional function of eEF3 in ribosome recycling. We have characterised the global effects of eEF3 depletion on translation using ribosome profiling. Depletion of eEF3 results in decreased ribosome density at the stop codon, indicating that ribosome recycling does not become rate limiting when eEF3 levels are low. Consistent with a defect in translation elongation, eEF3 depletion causes a moderate redistribution of ribosomes towards the 5’ part of the open reading frames. We observed no E-site codon-or amino acid-specific ribosome stalling upon eEF3 depletion, supporting its role as a general elongation factor. Surprisingly, depletion of eEF3 leads to a relative decrease in P-site proline stalling, which we hypothesise is a secondary effect of generally decreased translation and/or decreased competition for the E-site with eIF5A.

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“…All chloroviruses also encode a homolog of translational elongation factor 3 (EF-3) [ 13 ]. EF-3 plays a role in optimizing the accuracy of mRNA decoding at the ribosomal acceptor site during protein synthesis in fungi [ 35 , 36 ]; EF-3 has been reported recently in algae [ 37 ]. The role this putative enzyme plays in chlorovirus translation is unknown.…”

Section: Resultsmentioning

confidence: 99%

Diversity of tRNA Clusters in the Chloroviruses

Duncan

Dunigan

Etten

2020

Viruses

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Viruses rely on their host’s translation machinery for the synthesis of their own proteins. Problems belie viral translation when the host has a codon usage bias (CUB) that is different from an infecting virus due to differences in the GC content between the host and virus genomes. Here, we examine the hypothesis that chloroviruses adapted to host CUB by acquisition and selection of tRNAs that at least partially favor their own CUB. The genomes of 41 chloroviruses comprising three clades, each infecting a different algal host, have been sequenced, assembled and annotated. All 41 viruses not only encode tRNAs, but their tRNA genes are located in clusters. While differences were observed between clades and even within clades, seven tRNA genes were common to all three clades of chloroviruses, including the tRNAArg gene, which was found in all 41 chloroviruses. By comparing the codon usage of one chlorovirus algal host, in which the genome has been sequenced and annotated (67% GC content), to that of two of its viruses (40% GC content), we found that the viruses were able to at least partially overcome the host’s CUB by encoding tRNAs that recognize AU-rich codons. Evidence presented herein supports the hypothesis that a chlorovirus tRNA cluster was present in the most recent common ancestor (MRCA) prior to divergence into three clades. In addition, the MRCA encoded a putative isoleucine lysidine synthase (TilS) that remains in 39/41 chloroviruses examined herein, suggesting a strong evolutionary pressure to retain the gene. TilS alters the anticodon of tRNAMet that normally recognizes AUG to then recognize AUA, a codon for isoleucine. This is advantageous to the chloroviruses because the AUA codon is 12–13 times more common in the chloroviruses than their host, further helping the chloroviruses to overcome CUB. Among large DNA viruses infecting eukaryotes, the presence of tRNA genes and tRNA clusters appear to be most common in the Phycodnaviridae and, to a lesser extent, in the Mimiviridae.

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Demonstration of translation elongation factor 3 activity from a non-fungal species, Phytophthora infestans

Cited by 18 publications

References 30 publications

Evaluation and Validation of Reference Genes for Quantitative Real-Time PCR in Helopeltis theivora Waterhouse (Hemiptera: Miridae)

Evaluation and Validation of Reference Genes for Quantitative Real-Time PCR in Helopeltis theivora Waterhouse (Hemiptera: Miridae)

Ribosome profiling analysis of eEF3-depletedSaccharomyces cerevisiae

Diversity of tRNA Clusters in the Chloroviruses

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