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Parasitology Research

, Volume 118, Issue 10, pp 2843–2855 | Cite as

Functional characterization of the translation initiation factor eIF4E of Echinococcus granulosus

  • Filipe Santos Pereira-Dutra
  • Martin Cancela
  • Bruna Valandro Meneghetti
  • Henrique Bunselmeyer Ferreira
  • Karina Mariante Monteiro
  • Arnaldo ZahaEmail author
Genetics, Evolution, and Phylogeny - Original Paper
  • 121 Downloads

Abstract

The eukaryotic initiation factor 4E (eIF4E) specifically recognizes the 5′ mRNA cap, a rate-limiting step in the translation initiation process. Although the 7-methylguanosine cap (MMGcap) is the most common 5′ cap structure in eukaryotes, the trans-splicing process that occurs in several organism groups, including nematodes and flatworms, leads to the addition of a trimethylguanosine cap (TMGcap) to some RNA transcripts. In some helminths, eIF4E can have a dual capacity to bind both MMGcap and TMGcap. In the present work, we evaluated the distribution of eIF4E protein sequences in platyhelminths and we showed that only one gene coding for eIF4E is present in most parasitic flatworms. Based on this result, we cloned the Echinococcus granulosus cDNA sequence encoding eIF4E in Escherichia coli, expressed the recombinant eIF4E as a fusion protein to GST, and tested its ability to capture mRNAs through the 5′ cap using pull-down assay and qPCR. Our results indicate that the recombinant eIF4E was able to bind preferentially 5′-capped mRNAs compared with rRNAs from total RNA preparations of E. granulosus. By qPCR, we observed an enrichment in MMG-capped mRNA compared with TMG-capped mRNAs among Eg-eIF4E-GST pull-down RNAs. Eg-eIF4E structural model using the Schistosoma mansoni eIF4E as template showed to be well preserved with only a few differences between chemically similar amino acid residues at the binding sites. These data showed that E. granulosus eIF4E can be used as a potential tool to study full-length 5′-capped mRNA, besides being a potential drug target against parasitic flatworms.

Keywords

Capped RNAs mRNA purification Echinococcus granulosus eIF4E 

Notes

Author contributions

Co-wrote and edited the manuscript, retrieved sequences, performed phylogenetic analysis, and performed the experiments: FSPD and MC; performed the structural and docking analyses of proteins: BVM; conceived and designed the experiments: FSPD, MC, KMM, and AZ. Contributed reagents/materials/analysis tools: HBF and AZ. Wrote the paper: FSPD, MC, BVM, and AZ. All authors read and approved the final manuscript.

Funding

This study was financed in part by the Coordenação de Aperfeiçoamento de Pessoal de Nível Superior–Brasil (CAPES) (Finance Code 001 and grant number 1278/2011) and by the Conselho Nacional de Desenvolvimento Científico e Tecnológico (grant number 472316/2013–3). FSPD and BVM were recipients of a CAPES MSc. fellowship and MC was recipient of a CAPES postdoctoral fellowship.

Supplementary material

436_2019_6421_MOESM1_ESM.docx (48 kb)
Supplementary Table 1 (DOCX 47 kb)
436_2019_6421_MOESM2_ESM.docx (43 kb)
Supplementary Table 2 (DOCX 43 kb)
436_2019_6421_MOESM3_ESM.docx (44 kb)
Supplementary Table 3 (DOCX 43 kb)

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Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Laboratório de Biologia Molecular de Cestodeos, Centro de BiotecnologiaUniversidade Federal do Rio Grande do Sul, UFRGSPorto AlegreBrazil
  2. 2.Programa de Pós-Graduação em Biologia Celular e Molecular, Centro de BiotecnologiaUFRGSPorto AlegreBrazil
  3. 3.Departamento de Biologia Molecular e Biotecnologia, Instituto de BiociênciasUFRGSPorto AlegreBrazil

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