title: Binding specificities and potential roles of isoforms of eukaryotic initiation factor 4E in Leishmania.
abstract: The 5' cap structure of trypanosomatid mRNAs, denoted cap 4, is a complex structure that contains unusual modifications on the first four nucleotides. We examined the four eukaryotic initiation factor 4E (eIF4E) homologues found in the Leishmania genome database. These proteins, denoted LeishIF4E-1 to LeishIF4E-4, are located in the cytoplasm. They show only a limited degree of sequence homology with known eIF4E isoforms and among themselves. However, computerized structure prediction suggests that the cap-binding pocket is conserved in each of the homologues, as confirmed by binding assays to m(7)GTP, cap 4, and its intermediates. LeishIF4E-1 and LeishIF4E-4 each bind m(7)GTP and cap 4 comparably well, and only these two proteins could interact with the mammalian eIF4E binding protein 4EBP1, though with different efficiencies. 4EBP1 is a translation repressor that competes with eIF4G for the same residues on eIF4E; thus, LeishIF4E-1 and LeishIF4E-4 are reasonable candidates for serving as translation factors. LeishIF4E-1 is more abundant in amastigotes and also contains a typical 3' untranslated region element that is found in amastigote-specific genes. LeishIF4E-2 bound mainly to cap 4 and comigrated with polysomal fractions on sucrose gradients. Since the consensus eIF4E is usually found in 48S complexes, LeishIF4E-2 could possibly be associated with the stabilization of trypanosomatid polysomes. LeishIF4E-3 bound mainly m(7)GTP, excluding its involvement in the translation of cap 4-protected mRNAs. It comigrates with 80S complexes which are resistant to micrococcal nuclease, but its function is yet unknown. None of the isoforms can functionally complement the Saccharomyces cerevisiae eIF4E, indicating that despite their structural conservation, they are considerably diverged.
authors.affiliation: Department of Life Sciences, Ben Gurion University, P.O. Box 653, Beer Sheva 84105, Israel.
authors.forename: Yael
authors.initials: Y
authors.lastname: Yoffe
authors.forename: Joanna
authors.initials: J
authors.lastname: Zuberek
authors.forename: Asaf
authors.initials: A
authors.lastname: Lerer
authors.forename: Magdalena
authors.initials: M
authors.lastname: Lewdorowicz
authors.forename: Janusz
authors.initials: J
authors.lastname: Stepinski
authors.forename: Michael
authors.initials: M
authors.lastname: Altmann
authors.forename: Edward
authors.initials: E
authors.lastname: Darzynkiewicz
authors.forename: Michal
authors.initials: M
authors.lastname: Shapira
mesh_terms: D020413:3' Untranslated Regions; D000818:Animals; D001665:Binding Sites / Q000235:genetics; D039561:Eukaryotic Initiation Factor-4E / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D005786:Gene Expression Regulation; D017125:Genes, Protozoan; D066298:In Vitro Techniques; D007700:Kinetics; D018320:Leishmania major / Q000235:genetics / Q000378:metabolism*; D007894:Leishmania mexicana / Q000235:genetics / Q000378:metabolism*; D008958:Models, Molecular; D020033:Protein Isoforms / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017434:Protein Structure, Tertiary; D015800:Protozoan Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012314:RNA Cap Analogs / Q000235:genetics / Q000378:metabolism; D012315:RNA Caps / Q000235:genetics / Q000378:metabolism; D016053:RNA, Protozoan / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D013045:Species Specificity
chemical_list: D020413:3' Untranslated Regions; D039561:Eukaryotic Initiation Factor-4E; D020033:Protein Isoforms; D015800:Protozoan Proteins; D012314:RNA Cap Analogs; D012315:RNA Caps; D016053:RNA, Protozoan; D011993:Recombinant Fusion Proteins