Structural differences in translation initiation between pathogenic trypanosomatids and their mammalian hosts

Abstract

The 43S pre-initiation complex (PIC) assembly requires establishment of numerous interactions among eukaryotic initiation factors (eIFs), Met-tRNA i Met and the small ribosomal subunit (40S). Owing to several differences in the structure and composition of kinetoplastidian 40S compared to their mammalian counterparts, translation initiation in trypanosomatids is suspected to display substantial variability. Here, we determined the structure of the 43S PIC from Trypanosoma cruzi, the Chagas disease parasite, showing numerous specific features, such as different eIF3 structure and interactions with the large rRNA expansion segments 9 S , 7 S and 6 S , and the association of a kinetoplastid-specific ~245 kDa DDX60-like helicase. We also revealed a previously undetermined binding site of the eIF5 C-terminal domain, and terminal tails of eIF2β, eIF1, eIF1A and eIF3 c and d subunits, uncovering molecular details of their critical activities.One Sentence Summary: The 43S pre-initiation complex structure from Trypanosoma cruzi reveals kinetoplastid-specific features of translation initiation. Main Text:The first critical initiation step in eukaryotes is the assembly of the 43S PIC comprising the 40S, the eIF2•GTP•Met-tRNAi Met ternary complex, and eIFs 1, 1A, 3 and 5 (1, 2). It is followed by the recruitment of the mRNA promoted by the mRNA cap-binding complex comprising eIF4A, 4B and 4F (3, 4), forming the 48S PIC. The 48S PIC then scans the 5' untranslated region (UTR) of mRNA in the 5' to 3' direction till a start codon is encountered, upon which the majority of eIFs sequentially disassemble from the 40S and the resulting 48S initiation complex (48S IC) joins the large ribosomal subunit (60S) to form an elongation-competent 80S ribosome. Kinetoplastids is a group of flagellated unicellular eukaryotic parasites that have a complex life cycle. They spend part of their life cycle in the insect guts before being transmitted to the mammalian host upon biting. Common kinetoplastids include human pathogens such as Trypanosoma cruzi, Trypanosoma brucei and Leishmania spp., etiologic agents of Chagas disease, African sleeping sickness and leishmaniasis, respectively. However, most of the related public health measures are mainly preventative and therapeutic strategies are extremely limited and often highly toxic. Since kinetoplastids have diverged early from other eukaryotes, their mRNA translational machineries developed unique molecular features unseen in other eukaryotic species. For instance, their 40S contains a kinetoplastid-specific ribosomal protein (KSRP) (5) and unusually oversized ribosomal RNA (rRNA) expansion segments (ES S ) (6). Since these unique features may play specific roles in kinetoplastidian mRNA translation, they provide potential specific drug targets.It was proposed that two particularly oversized expansion segments, ES6 S and ES7 S located near the mRNA exit channel on the kinatoplastidian 40S, may contribute to modulating translation initiation in kinetoplastids by interacting with the structural co...