Biology subjects

Hashem, Y.

Publications and source records attributed to Hashem, Y..

3 recordsLinked to original sources

bioRxiv2019

Unique features of mRNA translation initiation in trypanosomatids

Canonical mRNA translation in eukaryotes begins with the formation of the 43S pre-initiation complex (PIC). Its assembly requires the binding of several eukaryotic initiation factors (eIF 1, 1A, 2, 3 and 5), Met-tRNAiMet and the small ribosomal subunit (40S). Compared to their mammalian hosts, trypanosomatids present significant structural differences in their 40S suggesting substantial variability in translation initiation. Here we determined the structure of the 43S PIC from Trypanosoma cruzi, the parasite causing the Chagas disease. Our structure shows numerous specific features, such as the variant eIF3 structure and its unique interactions with the large rRNA ESs 9S, 7S and 6S, and the association of a kinetoplastid-specific ~245 kDa DDX60-like helicase. It also revealed the so-far-elusive 40S-binding site of the eIF5 C-terminal domain and the structures of key terminal tails of several conserved eIFs underlying their activities within the PIC. Our results are corroborated by GST-pulldown assays in both human and T. cruzi and mass-spectrometry data.

molecular biology↗

bioRxiv2019

Structural insights into the Kozak sequence interactions within the mammalian late-stage initiation complexes

In higher eukaryotes, the mRNA sequence in direct vicinity of the start codon, called the Kozak sequence (CRCCaugG, where R is a purine), is known to influence the rate of the initiation process. However, the molecular basis underlying its role remains poorly understood. Here, we present the cryo-electron microscopy (cryo-EM) structures of mammalian late-stage 48S initiation complexes (LS48S IC) in the presence of two different native mRNA sequences, {beta}-globin and histone 4 (H4) at overall resolution of 3[A] and 3.5[A], respectively. Our high-resolution structures unravel key interactions from the mRNA to eukaryotic initiation factors (eIF): 1A, 2, 3, 18S rRNA, and several 40S ribosomal proteins. In addition, we were able to study the structural role of ABCE1 in the formation of native 48S ICs. Our results reveal a comprehensive map of the ribosome/eIFs -mRNA and -tRNA interactions and suggest the impact of mRNA sequence on the structure of the LS48S IC.

molecular biology↗

bioRxiv2019

Cryo-EM structure of the RNA-rich plant mitochondrial ribosome

The vast majority of eukaryotic cells contain mitochondria, essential powerhouses and metabolic hubs1. These organelles have a bacterial origin and were acquired during an early endosymbiosis event2. Mitochondria possess specialized gene expression systems composed of various molecular machines including the mitochondrial ribosomes (mitoribosomes). Mitoribosomes are in charge of translating the few essential mRNAs still encoded by mitochondrial genomes3. While chloroplast ribosomes strongly resemble those of bacteria4,5, mitoribosomes have diverged significantly during evolution and present strikingly different structures across eukaryotic species6-10. In contrast to animals and trypanosomatides, plants mitoribosomes have unusually expanded ribosomal RNAs and conserved the short 5S rRNA, which is usually missing in mitoribosomes11. We have previously characterized the composition of the plant mitoribosome6 revealing a dozen plant-specific proteins, in addition to the common conserved mitoribosomal proteins. In spite of the tremendous recent advances in the field, plant mitoribosomes remained elusive to high-resolution structural investigations, and the plant-specific ribosomal features of unknown structures. Here, we present a cryo-electron microscopy study of the plant 78S mitoribosome from cauliflower at near-atomic resolution. We show that most of the plant-specific ribosomal proteins are pentatricopeptide repeat proteins (PPR) that deeply interact with the plant-specific rRNA expansion segments. These additional rRNA segments and proteins reshape the overall structure of the plant mitochondrial ribosome, and we discuss their involvement in the membrane association and mRNA recruitment prior to translation initiation. Finally, our structure unveils an rRNA-constructive phase of mitoribosome evolution across eukaryotes.

molecular biology↗