LeishIF4E2 is a cap-binding protein that plays a role in Leishmania cell cycle progression.

Leishmania encode six paralogs of the cap-binding protein eIF4E and five eIF4G candidates, forming unique complexes. Two cap-binding proteins, LeishIF4E1 and LeishIF4E2, do not bind any identified LeishIF4Gs, thus their roles are intriguing. Here, we combine structural prediction, proteomic analysis, and interaction assays to shed light on LeishIF4E2 function. A nonconserved C-terminal extension was identified through structure prediction and sequence alignment. m7 GTP-binding assays involving both recombinant and transgenic LeishIF4E2 with and without the C-terminal extension revealed that this extension functions as a regulatory gate, modulating the cap-binding activity of LeishIF4E2. The interactomes of the two LeishIF4E2 versions were investigated, highlighting the role of the C-terminal extension in binding to SLBP2. SLBP2 is known to interact with a stem-loop structure in the 3' UTRs of histone mRNAs. Consistent with the predicted inhibitory effect of SLBP2 on histone expression in Xenopus laevis, a hemizygous deletion mutant of LeishIF4E2, exhibited an upregulation of several histones. We therefore propose that LeishIF4E2 is involved in histone expression, possibly through its interaction between SLBP2 and LeishIF4E2, thus affecting cell cycle progression. In addition, cell synchronization showed that LeishIF4E2 expression decreased during the S-phase, when histones are known to be synthesized. Previous studies in T. brucei also highlighted an association between TbEIF4E2 and SLBP2, and further reported on an interaction between TbIF4E2 and S-phase-abundant mRNAs. Our results show that overexpression of LeishIF4E2 correlates with upregulation of cell cycle and chromosome maintenance proteins. Along with its effect on histone expression, we propose that LeishIF4E2 is involved in cell cycle progression.

LeishIF3d is a non-canonical cap-binding protein in Leishmania.

Translation of most cellular mRNAs in eukaryotes proceeds through a cap-dependent pathway, whereby the cap-binding complex, eIF4F, anchors the pre-initiation complex at the 5' end of mRNAs driving translation initiation. The genome of Leishmania encodes a large repertoire of cap-binding complexes that fulfill a variety of functions possibly involved in survival along the life cycle. However, most of these complexes function in the promastigote life form that resides in the sand fly vector and decrease their activity in amastigotes, the mammalian life form. Here we examined the possibility that LeishIF3d drives translation in Leishmania using alternative pathways. We describe a non-canonical cap-binding activity of LeishIF3d and examine its potential role in driving translation. LeishIF3d is required for translation, as reducing its expression by a hemizygous deletion reduces the translation activity of the LeishIF3d(+/-) mutant cells. Proteomic analysis of the mutant cells highlights the reduced expression of flagellar and cytoskeletal proteins, as reflected in the morphological changes observed in the mutant cells. Targeted mutations in two predicted alpha helices diminish the cap-binding activity of LeishIF3d. Overall, LeishIF3d could serve as a driving force for alternative translation pathways, although it does not seem to offer an alternative pathway for translation in amastigotes.

Proteomic analysis of heterotic seed germination in maize using F1 hybrid DHM 117 and its parental inbreds.

A number of differentially expressed proteins (DEPs) were identified in comparative two-dimensional gel electrophoresis analysis of dry and 24-h water-imbibed seeds of maize F1 hybrid DHM 117 (BML 6 × BML 7) and its parental inbreds. Of the DEPs, 53.4% (86/161) in dry seeds and 58% (127/219) in water-imbibed seeds exhibited a nonadditive pattern in the F1 hybrid as compared to parental inbreds. A total of 30 DEPs were categorized into different biological processes, most of which were related to metabolism and energy (34%), followed by storage proteins (27%), stress response (23%), transcription and translation (7%), cell cycle (3%), and hormone biosynthesis (3%). The transcript accumulation pattern of 8 selected genes corresponding to DEPs was examined using qRTPCR. Interestingly, LEA protein Rab28 showed higher accumulation in dry seeds at both protein and transcript levels, whereas indole3-acetaldehyde oxidase showed lower accumulation in water-imbibed seeds of the F1 hybrid than the female parent at the protein level. Thus, the DEPs particularly involved in metabolic and energy processes, as well as hormone biosynthesis in the F 1 hybrid, might be responsible for heterotic seed germination in the F1 hybrid. The DEPs identified in this study provide a scope for improving the seed germination trait of agricultural crops.