Abstract
Eukaryotic translation initiation factor 3 (eIF3) is a key regulator of translation initiation, but its in vivo assembly and molecular functions remain unclear. Here we show that eIF3 from Neurospora crassa is structurally and compositionally similar to human eIF3. N. crassa eIF3 forms a stable 12-subunit complex linked genetically and biochemically to the 13th subunit, eIF3j, which in humans modulates mRNA start codon selection. Based on N. crassa genetic analysis, most subunits in eIF3 are essential. Subunits that can be deleted (e, h, k and l) map to the right side of the eIF3 complex, suggesting that they may coordinately regulate eIF3 function. Consistent with this model, subunits eIF3k and eIF3l are incorporated into the eIF3 complex as a pair, and their insertion depends on the presence of subunit eIF3h, a key regulator of vertebrate development. Comparisons to other eIF3 complexes suggest that eIF3 assembles around an eIF3a and eIF3c dimer, which may explain the coordinated regulation of human eIF3 levels. Taken together, these results show that Neurospora crassa eIF3 provides a tractable system for probing the structure and function of human-like eIF3 in the context of living cells.
Highlights
The regulation of protein synthesis in eukaryotes occurs predominantly during translation initiation
Using an eIF3j query from Aspergillus niger, we identified a orthologue for eIF3j
The Neurospora DeIF3kl strain exhibited nearly wildtype linear growth rates and conidiation on sucrose minimal media (Figure 2A and B, Table S2 in File S1). These results suggest that subunits k and l are not required for assembly of the remainder of the Eukaryotic translation initiation factor 3 (eIF3) complex in vivo
Summary
The regulation of protein synthesis in eukaryotes occurs predominantly during translation initiation. Translation initiation in eukaryotes is regulated by a number of eukaryotic initiation factors (eIFs) whose specific roles in this process remain unclear. EIF3 is the largest eIF, consisting of 13 non-identical protein subunits named eIF3a through eIF3m [1]. EIF3 is required for genomic RNA recruitment to the small ribosomal subunit during viral internal ribosome entry site (IRES)-dependent translation [2,4,5,6]. EIF3 subunits have been tied to developmental pathways that may require eIF3 to recruit mRNAs to PICs [8,9,10]. The overall architecture of human eIF3 has recently been described [11], the specific functions of its subunits and its in vivo assembly pathway remain unclear [3,11,12]
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