eIF4F-like complexes formed by cap-binding homolog TbEIF4E5 with TbEIF4G1 or TbEIF4G2 are implicated in post-transcriptional regulation in Trypanosoma brucei

Abstract

Members of the eIF4E mRNA cap-binding family are involved in translation and the modulation of transcript availability in other systems as part of a three-component complex including eIF4G and eIF4A. The kinetoplastids possess four described eIF4E and five eIF4G homologs. We have identified two new eIF4E family proteins in Trypanosoma brucei, and define distinct complexes associated with the fifth member, TbEIF4E5. The cytosolic TbEIF4E5 protein binds cap 0 in vitro. TbEIF4E5 was found in association with two of the five TbEIF4Gs. TbIF4EG1 bound TbEIF4E5, a 47.5-kDa protein with two RNA-binding domains, and either the regulatory protein 14-3-3 II or a 117.5-kDa protein with guanylyltransferase and methyltransferase domains in a potentially dynamic interaction. The TbEIF4G2/TbEIF4E5 complex was associated with a 17.9-kDa hypothetical protein and both 14-3-3 variants I and II. Knockdown of TbEIF4E5 resulted in the loss of productive cell movement, as evidenced by the inability of the cells to remain in suspension in liquid culture and the loss of social motility on semisolid plating medium, as well as a minor reduction of translation. Cells appeared lethargic, as opposed to compromised in flagellar function per se. The minimal use of transcriptional control in kinetoplastids requires these organisms to implement downstream mechanisms to regulate gene expression, and the TbEIF4E5/TbEIF4G1/117.5-kDa complex in particular may be a key player in that process. We suggest that a pathway involved in cell motility is affected, directly or indirectly, by one of the TbEIF4E5 complexes.

Two related trypanosomatid eIF4G homologues have functional differences compatible with distinct roles during translation initiation

“…This conservation is in striking contrast with the divergent N and C-terminal regions of the other 3 trypanosomatid eIF4G homologues (EIF4G1, EIF4G2 and EIF4G5) which have recently been seen to participate in novel eIF4F-like complexes that do not seem to have primary roles in translation. 40,41 The presence of putative MA3/ HEAT-2 and W2/HEAT-3 domains in at least 2 trypanosomatid eIF4G homologues confirms the likely ancient origin of the eIF4G tripartite structure, despite the fact that both MA3 and W2 are missing from yeast eIF4G and the W2 domain is not found in plant eIF4G homologues. 32 The evidence presented clearly distinguishes EIF4G3 and EIF4G4 functionally.…”

“…More recently EIF4G1, EIF4G2 and EIF4G5 have been shown to form complexes with 2 novel eIF4E homologues, EIF4E5 and EIF4E6, but they have not been linked to the translation initiation process. 40,41 Here, the functional properties and individual roles of EIF4G3 and EIF4G4 have been further investigated in both L. major and T. brucei. First, binding to eIF4Es, eIF4A and PABPs were investigated in vitro.…”

Structural basis for LeishIF4E-1 modulation by an interacting protein in the human parasite Leishmania major

“…Trypanosomatids encode several isoforms of the same initiation factor; there are six highly diverged paralogs of the human eIF4E cap-binding protein (LeishIF4E-1 to LeishIF4E-6). These have a small degree of conservation among themselves (40–60%) and also differ significantly from their counterparts in higher eukaryotes (30–40% sequence identity) (Supplementary Figure S1A) (24,26,27). Of the six paralogs, LeishIF4E-1 is highly expressed in axenic amastigotes and binds the m 7 GTP at 37°C, suggesting its involvement in translation initiation in amastigotes (23).…”

A Glycosylation Mutant of Trypanosoma brucei Links Social Motility Defects In Vitro to Impaired Colonization of Tsetse Flies In Vivo

“…Somewhat surprisingly, none of the procyclins is required for SoMo (12). The three mutants so far found to be defective all are motility mutants (13,16).Rft1 is an endoplasmic reticular protein involved in the conversion of Man 5 GlcNAc 2 -PP-dolichol (M5-DLO) to M9-DLO, the precursor for N-linked glycans (17-19). The protein is essential in yeast; in humans, mutations have been linked to congenital …”

“…Somewhat surprisingly, none of the procyclins is required for SoMo (12). The three mutants so far found to be defective all are motility mutants (13,16).…”