The Interaction between the Ribosomal Stalk Proteins and Translation Initiation Factor 5B Promotes Translation Initiation

Abstract

Ribosomal stalk proteins recruit translation elongation GTPases to the factor-binding center of the ribosome. Initiation factor 5B (eIF5B in eukaryotes and aIF5B in archaea) is a universally conserved GTPase that promotes the joining of the large and small ribosomal subunits during translation initiation. Here we show that aIF5B binds to the C-terminal tail of the stalk protein. In the cocrystal structure, the interaction occurs between the hydrophobic amino acids of the stalk C-terminal tail and a small hydrophobic pocket on the surface of the GTP-binding domain (domain I) of aIF5B. A substitution mutation altering the hydrophobic pocket of yeast eIF5B resulted in a marked reduction in ribosome-dependent eIF5B GTPase activity In yeast cells, the eIF5B mutation affected growth and impaired expression during amino acid starvation via a defect in start site selection for the first upstream open reading frame in mRNA, as observed with the eIF5B deletion mutant. The deletion of two of the four stalk proteins diminished polyribosome levels (indicating defective translation initiation) and starvation-induced expression, both of which were suppressible by eIF5B overexpression. Thus, the mutual interaction between a/eIF5B and the ribosomal stalk plays an important role in subunit joining during translation initiation .

Ribosomal mutation in helix 32 of 18S rRNA alters fidelity of eukaryotic translation start site selection

“…A,B). If the increase in free 60S ribosome pool causes faster 80S formation then it should have prevented leaky scanning for uORF and exacerbated the Sui − phenotype . No such effects were observed for the C1209U mutant 40S ribosome suggesting that the resultant conformation changes in this ribosome may be delaying steps necessary for 80S formation.…”

Direct visualization of translational GTPase factor-pool formed around the archaeal ribosomal P-stalk by high-speed atomic force microscopy

“…In translation initiation, the ribosomal stalk interacts with the GTPase initiation factor IF2 (aIF5B in archaea and eIF5B in eukaryotes) to promote large and small subunit joining and selectivity for the initiation AUG codon (45,46). The ribosomal stalk also plays a functional role in the recruitment of ribosome recycling factor ABCE1 and its ATP hydrolysis in archaea and eukaryotes (47).…”

“…The ribosomal stalk also plays a functional role in the recruitment of ribosome recycling factor ABCE1 and its ATP hydrolysis in archaea and eukaryotes (47). Point mutations disrupting the binding of the ribosomal stalk to eIF5B and ABCE1 inhibit cell viability, suggesting that the ribosomal stalk maintains these functional interactions with translation initiation and recycling factors in vivo (46,47). Therefore, at least four translation factors appear to compete with each other as the binding partners for the ribosomal stalk.…”

Phosphorylation of the N‐terminal domain of ribosomal P‐stalk protein uL10 governs its association with the ribosome

“…The P-stalk is the central element of the GTPase-associated center (GAC) and is constituted by three proteins: P1, P2, and uL10 (formerly named P0) [16], which together form a pentameric complex uL10 (P1-P2) 2 [17]. The primary role of the P-stalk is interaction with translational GTPases and stimulation of their GTPase activity during each step of translation [18][19][20][21][22][23][24]. The phenomenon of multiplication of P-stalk proteins was functionally linked with the decoding event, thus associating the stalk also with translational fidelity [25].…”

“…The phenomenon of multiplication of P-stalk proteins was functionally linked with the decoding event, thus associating the stalk also with translational fidelity [25]. The stalk proteins were named P-proteins [26], because they are phosphorylated by CK2 protein kinase at their conserved C termini [27][28][29], that is, a protein element that is regarded as a functional region responsible for direct interactions with translational factors or ribosome-inactivating proteins [23,24,30,31]. However, despite the decades of work, the function of this PTM remains undiscovered.…”