scholarly journals Arabidopsis thaliana class II poly(A)-binding proteins are required for efficient multiplication of turnip mosaic virus

2008 ◽  
Vol 89 (9) ◽  
pp. 2339-2348 ◽  
Author(s):  
Philippe J. Dufresne ◽  
Eliane Ubalijoro ◽  
Marc G. Fortin ◽  
Jean-François Laliberté
Keyword(s):  
Arabidopsis Thaliana ◽  
Mosaic Virus ◽  
Class Ii ◽  
Turnip Mosaic Virus ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Mrna Levels ◽  
Binding Assays ◽  
Rna Accumulation

The poly(A)-binding protein (PABP) is an important translation initiation factor that binds to the polyadenylated 3′ end of mRNA. We have previously shown that PABP2 interacts with the RNA-dependent RNA polymerase (RdRp) and VPg-Pro of turnip mosaic virus (TuMV) within virus-induced vesicles. At least eight PABP isoforms are produced in Arabidopsis thaliana, three of which (PABP2, PABP4 and PABP8) are highly and broadly expressed and probably constitute the bulk of PABP required for cellular functions. Upon TuMV infection, an increase in protein and mRNA expression from PAB2, PAB4 and PAB8 genes was recorded. In vitro binding assays revealed that RdRp and the viral genome-linked protein (VPg-Pro) interact preferentially with PABP2 but are also capable of interaction with one or both of the other class II PABPs (i.e. PABP4 and PABP8). To assess whether PABP is required for potyvirus replication, A. thaliana single and double pab knockouts were isolated and inoculated with TuMV. All lines showed susceptibility to TuMV. However, when precise monitoring of viral RNA accumulation was performed, it was found to be reduced by 2.2- and 3.5-fold in pab2 pab4 and pab2 pab8 mutants, respectively, when compared with wild-type plants. PABP levels were most significantly reduced in the membrane-associated fraction in both of these mutants. TuMV mRNA levels thus correlated with cellular PABP concentrations in these A. thaliana knockout lines. These data provide further support for a role of PABP in potyvirus replication.

scholarly journals Interaction of VPg-Pro of Turnip mosaic virus with the translation initiation factor 4E and the poly(A)-binding protein in planta

2004 ◽  
Vol 85 (4) ◽  
pp. 1055-1063 ◽  
Author(s):  
Simon Léonard ◽  
Catherine Viel ◽  
Chantal Beauchemin ◽  
Nicole Daigneault ◽  
Marc G. Fortin ◽  
...  
Keyword(s):  
Mosaic Virus ◽  
Binding Protein ◽  
Gradient Centrifugation ◽  
Direct Interaction ◽  
Turnip Mosaic Virus ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Metal Chelation ◽  
In Planta

The viral protein linked to the genome (VPg) of Turnip mosaic virus (TuMV) interacts in vitro with the translation eukaryotic initiation factor (eIF) 4E. In the present study, we investigated the consequence of TuMV infection on eIF4E expression. Two isomers are present in plants, namely eIF4E and eIF(iso)4E. Expression of the latter was detected in both TuMV-infected and mock-inoculated Brassica perviridis plants, but expression of eIF4E was found only in infected plants. Membranes from TuMV-infected or mock-inoculated tissues were separated by sucrose gradient centrifugation and fractions were collected. Immunoblot analyses showed that 6K2-VPg-Pro/VPg-Pro polyproteins were associated with endoplasmic reticulum membranes and were the viral forms likely to interact with eIF(iso)4E and eIF4E. In planta interaction between 6K2-VPg-Pro/VPg-Pro and eIF(iso)4E/eIF4E was confirmed by co-purification by metal chelation chromatography. The poly(A)-binding protein (PABP) was also found to co-purify with VPg-Pro. Direct interaction between VPg-Pro and PABP was shown by an ELISA-based binding assay. These experiments suggest that a multi-protein complex may form around VPg-Pro of TuMV.

scholarly journals Multiple Resistance Phenotypes to Lettuce mosaic virus Among Arabidopsis thaliana Accessions

2003 ◽  
Vol 16 (7) ◽  
pp. 608-616 ◽  
Author(s):  
Frédéric Revers ◽  
Thomas Guiraud ◽  
Marie-Christine Houvenaghel ◽  
Thierry Mauduit ◽  
Olivier Le Gall ◽  
...  
Keyword(s):  
Arabidopsis Thaliana ◽  
Mosaic Virus ◽  
Resistance Gene ◽  
Recombinant Inbred Lines ◽  
Cell Movement ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Lettuce Mosaic Virus ◽  
Genetic Analyses ◽  
Large Variability

With the aim to characterize plant and viral factors involved in the molecular interactions between plants and potyviruses, a Lettuce mosaic virus (LMV)-Arabidopsis thaliana pathosystem was developed. Screening of Arabi-dopsis accessions with LMV isolates indicated the existence of a large variability in the outcome of the interaction, allowing the classification of Arabidopsis accessions into seven susceptibility groups. Using a reverse genetic approach, the genome-linked protein of LMV, a multifunc-tional protein shown to be involved in viral genome amplification and movement of potyviruses, was established as the viral determinant responsible for the ability to overcome the resistance of the Niederzenz accession to LMV-0. Preliminary genetic analyses from F2 and recombinant inbred lines available between susceptible and resistant Arabidopsis accessions revealed the existence of at least three resistance phenotypes to LMV with different genetic bases. One dominant resistance gene, designated LLM1, involved in blocking the replication or cell-to-cell movement of the LMV-0 isolate in the Columbia accession, was mapped to chromosome I and shown to be linked to the marker nga280. At the same time, genetic analyses of segregating F2 populations were consistent with the restriction of the systemic movement of the LMV-AF199 isolate in Columbia being controlled by two dominant genes and with the complete resistance to all tested LMV isolates of the Cape Verde islands (Cvi) accession being conferred by a single recessive resistance gene. Sequencing of the eu-karyotic translation initiation factor 4E genes from the different LMV-resistant Arabidopsis accessions showed that these factors are not directly involved in the characterized resistance phenotypes.

scholarly journals Identification of a Translation Initiation Factor 3 (eIF3) Core Complex, Conserved in Yeast and Mammals, That Interacts with eIF5

1998 ◽  
Vol 18 (8) ◽  
pp. 4935-4946 ◽  
Author(s):  
Lon Phan ◽  
Xiaolong Zhang ◽  
Katsura Asano ◽  
James Anderson ◽  
Hans-Peter Vornlocher ◽  
...  
Keyword(s):  
Translation Initiation ◽  
Gel Filtration ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Start Codon ◽  
Binding Assays ◽  
Core Complex ◽  
Eukaryotic Translation ◽  
Translation Initiation Factor 3

ABSTRACT Only five of the nine subunits of human eukaryotic translation initiation factor 3 (eIF3) have recognizable homologs encoded in theSaccharomyces cerevisiae genome, and only two of these (Prt1p and Tif34p) were identified previously as subunits of yeast eIF3. We purified a polyhistidine-tagged form of Prt1p (His-Prt1p) by Ni2+ affinity and gel filtration chromatography and obtained a complex of ≈600 kDa composed of six polypeptides whose copurification was completely dependent on the polyhistidine tag on His-Prt1p. All five polypeptides associated with His-Prt1p were identified by mass spectrometry, and four were found to be the other putative homologs of human eIF3 subunits encoded in S. cerevisiae: YBR079c/Tif32p, Nip1p, Tif34p, and YDR429c/Tif35p. The fifth Prt1p-associated protein was eIF5, an initiation factor not previously known to interact with eIF3. The purified complex could rescue Met-tRNAi Met binding to 40S ribosomes in defective extracts from a prt1 mutant or extracts from which Nip1p had been depleted, indicating that it possesses a known biochemical activity of eIF3. These findings suggest that Tif32p, Nip1p, Prt1p, Tif34p, and Tif35p comprise an eIF3 core complex, conserved between yeast and mammals, that stably interacts with eIF5. Nip1p bound to eIF5 in yeast two-hybrid and in vitro protein binding assays. Interestingly, Sui1p also interacts with Nip1p, and both eIF5 and Sui1p have been implicated in accurate recognition of the AUG start codon. Thus, eIF5 and Sui1p may be recruited to the 40S ribosomes through physical interactions with the Nip1p subunit of eIF3.

Turnip mosaic virus VPg interacts with Arabidopsis thaliana eIF(iso)4E and inhibits in vitro translation

Biochimie ◽  
2008 ◽  
Vol 90 (10) ◽  
pp. 1427-1434 ◽  
Author(s):  
Hiroshi Miyoshi ◽  
Hayato Okade ◽  
Shinji Muto ◽  
Noriko Suehiro ◽  
Hideki Nakashima ◽  
...  
Keyword(s):  
Arabidopsis Thaliana ◽  
Mosaic Virus ◽  
Turnip Mosaic Virus ◽  
In Vitro Translation

scholarly journals The Absence of Eukaryotic Initiation Factor eIF(iso)4E Affects the Systemic Spread of a Tobacco etch virus Isolate in Arabidopsis thaliana

2013 ◽  
Vol 26 (4) ◽  
pp. 461-470 ◽  
Author(s):  
Carlos A. Contreras-Paredes ◽  
Laura Silva-Rosales ◽  
José-Antonio Daròs ◽  
Naholi D. Alejandri-Ramírez ◽  
Tzvetanka D. Dinkova
Keyword(s):  
Arabidopsis Thaliana ◽  
Mosaic Virus ◽  
Resistance Mechanism ◽  
Tobacco Etch Virus ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Viral Translation ◽  
Systemic Spread ◽  
Viral Coat

Translation initiation factor eIF4E exerts an important role during infection of viral species in the family Potyviridae. Particularly, a eIF(iso)4E family member is required for Arabidopsis thaliana susceptibility to Turnip mosaic virus, Lettuce mosaic virus, and Tobacco etch virus (TEV). In addition, a resistance mechanism named restriction of TEV movement (RTM) in A. thaliana controls the systemic spread of TEV in Col-0 ecotype. Here, we describe that TEV-TAMPS, a Mexican isolate, overcomes the RTM resistance mechanism reported for TEV-7DA infection of the Col-0 ecotype but depends on eIF(iso)4E for its systemic spread. To understand at which level eIF(iso)4E participates in A. thaliana TEV-TAMPS infection, the viral RNA replication and translation were measured. The absence or overexpression of eIF(iso)4E did not affect viral translation, and replication was still observed in the absence of eIF(iso)4E. However, the TEV-TAMPS systemic spread was completely abolished in the null mutant. The viral protein genome-linked (VPg) precursor NIa was found in coimmunoprecipitated complexes with both, eIF(iso)4E and eIF4E. However, the viral coat protein (CP) was only present in the eIF(iso)4E complexes. Since both the VPg and the CP proteins are needed for systemic spread, we propose a role of A. thaliana eIF(iso)4E in the movement of TEV-TAMPS within this host.

scholarly journals The Poly(A) Binding Protein Is Internalized in Virus-Induced Vesicles or Redistributed to the Nucleolus during Turnip Mosaic Virus Infection

2007 ◽  
Vol 81 (20) ◽  
pp. 10905-10913 ◽  
Author(s):  
Chantal Beauchemin ◽  
Jean-François Laliberté
Keyword(s):  
Confocal Microscopy ◽  
Mosaic Virus ◽  
Binding Protein ◽  
Subcellular Fractionation ◽  
Turnip Mosaic Virus ◽  
Translation Initiation Factor ◽  
Host Protein ◽  
Initiation Factor ◽  
Gradient Fractionation ◽  
Mosaic Virus Infection

ABSTRACT Poly(A) binding protein 2 (PABP2) of Arabidopsis thaliana was previously shown to interact with VPg-Pro of turnip mosaic virus (TuMV) and may consequently play an important role during infection. Subcellular fractionation experiments revealed that PABP2 was predominantly a cytoplasmic soluble protein in healthy plants. However, in TuMV-infected plants, a subpopulation of PABP2 was membrane associated or was localized in the nucleus. Confocal microscopy experiments indicated that PABP2 was partially retargeted to the nucleolus in the presence of TuMV VPg-Pro. In addition, the membrane association of PABP2 during TuMV infection resulted from the internalization of the host protein in 6K-VPg-Pro-induced vesicles, as shown by a combination of confocal microscopy and sucrose gradient fractionation experiments. This redistribution of an important translation initiation factor to the nucleolus and to membrane structure likely underlies two important processes of the TuMV replication cycle.

Discovery of N-Phenyl-4-(1H-pyrrol-3-yl)pyrimidin-2-amine Derivatives as Potent Mnk2 Inhibitors: Design, Synthesis, SAR Analysis, and Evaluation of in vitro Anti-leukaemic Activity

2019 ◽  
Vol 15 (6) ◽  
pp. 602-623 ◽  
Author(s):  
Ahmed M. Abdelaziz ◽  
Sarah Diab ◽  
Saiful Islam ◽  
Sunita K.C. Basnet ◽  
Benjamin Noll ◽  
...  
Keyword(s):  
Proliferative Activity ◽  
Myeloid Cell ◽  
Structural Diversity ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Cell Leukaemia ◽  
Aberrant Expression ◽  
Design Synthesis ◽  
Induced Apoptosis

Background:Aberrant expression of eukaryotic translation initiation factor 4E (eIF4E) is common in many types of cancer including acute myeloid leukaemia (AML). Phosphorylation of eIF4E by MAPK-interacting kinases (Mnks) is essential for the eIF4E-mediated oncogenic activity. As such, the pharmacological inhibition of Mnks can be an effective strategy for the treatment of cancer.Methods:A series of N-phenyl-4-(1H-pyrrol-3-yl)pyrimidin-2-amine derivatives was designed and synthesised. The Mnk inhibitory activity of these derivatives as well as their anti-proliferative activity against MV4-11 AML cells was determined.Results:These compounds were identified as potent Mnk2 inhibitors. Most of them demonstrated potent anti-proliferative activity against MV4-11 AML cells. The cellular mechanistic studies of the representative inhibitors revealed that they reduced the level of phosphorylated eIF4E and induced apoptosis by down-regulating the anti-apoptotic protein myeloid cell leukaemia 1 (Mcl-1) and by cleaving poly(ADP-ribose)polymerase (PARP). The lead compound 7k possessed desirable pharmacokinetic properties and oral bioavailability.Conclusion:This work proposes that exploration of the structural diversity in the context of Nphenyl- 4-(1H-pyrrol-3-yl)pyrimidin-2-amine would offer potent and selective Mnk inhibitors.

scholarly journals Comparison of two Turnip mosaic virus P1 proteins in their ability to co-localize with the Arabidopsis thaliana G3BP-2 protein

Virus Genes ◽  
2021 ◽  
Vol 57 (2) ◽  
pp. 233-237
Author(s):  
Hendrik Reuper ◽  
Björn Krenz
Keyword(s):  
Arabidopsis Thaliana ◽  
Mosaic Virus ◽  
Specific Interaction ◽  
Viral Proteins ◽  
Turnip Mosaic Virus ◽  
Stress Conditions ◽  
C Terminus ◽  
Positive Sense ◽  
Key Factor ◽  
Large Host

AbstractTurnip mosaic virus (TuMV), belonging to the genus Potyvirus (family Potyviridae), has a large host range and consists of a single-stranded positive sense RNA genome encoding 12 proteins, including the P1 protease. This protein which is separated from the polyprotein by cis cleavage at its respective C-terminus, has been attributed with different functions during potyviral infection of plants. P1 of Turnip mosaic virus (P1-TuMV) harbors an FGSF-motif and FGSL-motif at its N-terminus. This motif is predicted to be a binding site for the host Ras GTPase-activating protein-binding protein (G3BP), which is a key factor for stress granule (SG) formation in mammalian systems and often targeted by viruses to inhibit SG formation. We therefore hypothesized that P1-TuMV might interact with G3BP to control and regulate plant SGs to optimize cellular conditions for the production of viral proteins. Here, we analyzed the co-localization of the Arabidopsis thaliana G3BP-2 with the P1 of two TuMV isolates, namely UK 1 and DEU 2. Surprisingly, P1-TuMV-DEU 2 co-localized with AtG3BP-2 under abiotic stress conditions, whereas P1-TuMV-UK 1 did not. AtG3BP-2::RFP showed strong SGs formation after stress, while P1-UK 1::eGFP maintained a chloroplastic signal under stress conditions, the signal of P1-DEU 2::eGFP co-localized with that of AtG3BP-2::RFP. This indicates a specific interaction between P1-DEU 2 and the AtG3BP family which is not solely based on the canonical interaction motifs.

scholarly journals Circadian clock control of eIF2α phosphorylation is necessary for rhythmic translation initiation

2020 ◽  
Vol 117 (20) ◽  
pp. 10935-10945 ◽  
Author(s):  
Shanta Karki ◽  
Kathrina Castillo ◽  
Zhaolan Ding ◽  
Olivia Kerr ◽  
Teresa M. Lamb ◽  
...  
Keyword(s):  
Circadian Clock ◽  
Translation Initiation ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Nutrient Metabolism ◽  
Eif2α Phosphorylation ◽  
Eif2α Kinase ◽  
The Impact

The circadian clock in eukaryotes controls transcriptional and posttranscriptional events, including regulation of the levels and phosphorylation state of translation factors. However, the mechanisms underlying clock control of translation initiation, and the impact of this potential regulation on rhythmic protein synthesis, were not known. We show that inhibitory phosphorylation of eIF2α (P-eIF2α), a conserved translation initiation factor, is clock controlled in Neurospora crassa, peaking during the subjective day. Cycling P-eIF2α levels required rhythmic activation of the eIF2α kinase CPC-3 (the homolog of yeast and mammalian GCN2), and rhythmic activation of CPC-3 was abolished under conditions in which the levels of charged tRNAs were altered. Clock-controlled accumulation of P-eIF2α led to reduced translation during the day in vitro and was necessary for the rhythmic synthesis of select proteins in vivo. Finally, loss of rhythmic P-eIF2α levels led to reduced linear growth rates, supporting the idea that partitioning translation to specific times of day provides a growth advantage to the organism. Together, these results reveal a fundamental mechanism by which the clock regulates rhythmic protein production, and provide key insights into how rhythmic translation, cellular energy, stress, and nutrient metabolism are linked through the levels of charged versus uncharged tRNAs.

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