Molecular Architecture of the 40S⋅eIF1⋅eIF3 Translation Initiation Complex

Abstract

SummaryEukaryotic translation initiation requires the recruitment of the large, multiprotein eIF3 complex to the 40S ribosomal subunit. We present X-ray structures of all major components of the minimal, six-subunit Saccharomyces cerevisiae eIF3 core. These structures, together with electron microscopy reconstructions, cross-linking coupled to mass spectrometry, and integrative structure modeling, allowed us to position and orient all eIF3 components on the 40S⋅eIF1 complex, revealing an extended, modular arrangement of eIF3 subunits. Yeast eIF3 engages 40S in a clamp-like manner, fully encircling 40S to position key initiation factors on opposite ends of the mRNA channel, providing a platform for the recruitment, assembly, and regulation of the translation initiation machinery. The structures of eIF3 components reported here also have implications for understanding the architecture of the mammalian 43S preinitiation complex and the complex of eIF3, 40S, and the hepatitis C internal ribosomal entry site RNA.

Eukaryote-specific extensions in ribosomal proteins of the small subunit: Structure and function

“…57 Very recently, Erzberger and coworkers presented an X-ray structure of the universally conserved core subunits of eIF3 (at »3.5 A ) and modeled the eIF3 complex position on the yeast 40S ribosomal subunit by employing cryo-EM and cross-linking of the factor (s) followed by mass spectrometry (MS). 58 This reconstruction provided evidence supporting some of the previously reported interactions (between eIF3 and eukaryote-specific 40S regions) observed by biochemical, genetic and structural analyses, but also revealed a number of new/ alternative interactions. 58 While the exact architecture of the eIF3 40S complex remains to be Figure 5.…”

“…58 This reconstruction provided evidence supporting some of the previously reported interactions (between eIF3 and eukaryote-specific 40S regions) observed by biochemical, genetic and structural analyses, but also revealed a number of new/ alternative interactions. 58 While the exact architecture of the eIF3 40S complex remains to be Figure 5. Sequence alignment of uS7 proteins from different organisms showing length variability of eukaryotespecific N-terminal extensions.…”

Integrative structure and functional anatomy of a nuclear pore complex

“…7A–C; shown in blue). Therefore, the dominant cluster essentially satisfies the cross-linking data within its uncertainty (the false detection rate is approximately 5% to 10% 138,139 ). Most of the cross-link violations are small, and can be rationalized by local structural fluctuations, coarse-grained representations of some Nup domains, and/or finite structural sampling, as shown in Extended Data Fig.…”

Eukaryotic aspects of translation initiation brought into focus

“…eIF4 is known to bind both the cap and, through poly-A binding protein, the tail of the translated mRNA, a pattern suspected to result in the proximity of the translational termination and start sites with obvious consequences for the efficiency of recycling of terminating ribosomes [11]; however, its exact structural arrangement with respect to the 40S remains one of the gaping holes in our knowledge. With the advent of higher-resolution cryo-electron microscopy (EM), it has proved possible, however, to obtain a relatively complete intermediate-resolution description of the organization of eIF3 from both yeast [8,9,13] and mammals [14][15][16] (figure 2). eIF3 circumnavigates the entire 40S ribosomal subunit [8,13], recruiting and coordinating essentially every element of the translational apparatus.…”

“…With the advent of higher-resolution cryo-electron microscopy (EM), it has proved possible, however, to obtain a relatively complete intermediate-resolution description of the organization of eIF3 from both yeast [8,9,13] and mammals [14][15][16] (figure 2). eIF3 circumnavigates the entire 40S ribosomal subunit [8,13], recruiting and coordinating essentially every element of the translational apparatus. Two key conserved sub-complexes occupy the vicinity of each end of the mRNA channel on the solvent-exposed surface of the small 40S subunit [8,15], stabilizing mRNA interacting helicases and contacting eIF4, while from both sides of the ribosomal subunit protein extensions of eIF3 interact with and recruit the evolutionarily conserved machinery that operates within the mRNA-binding channel and the decoding site.…”