Translation initiation complex formation in the crenarchaeon Sulfolobus solfataricus

Abstract

The function of initiation factors in and the sequence of events during translation initiation have been intensively studied in Bacteria and Eukaryotes, whereas in Archaea knowledge on these functions/processes is limited. By employing chemical probing, we show that translation initiation factor aIF1 of the model crenarchaeon Sulfolobus solfataricus binds to the same area on the ribosome as the bacterial and eukaryal orthologs. Fluorescence energy transfer assays (FRET) showed that aIF1, like its eukaryotic and bacterial orthologs, has a fidelity function in translation initiation complex formation, and that both aIF1 and aIF1A exert a synergistic effect in stimulating ribosomal association of the Met-tRNAi Met binding factor a/eIF2. However, as in Eukaryotes their effect on a/eIF2 binding appears to be indirect. Moreover, FRET was used to analyze for the first time the sequence of events toward translation initiation complex formation in an archaeal model system. These studies suggested that a/eIF2-GTP binds first to the ribosome and then recruits Met-tRNAi Met , which appears to comply with the operational mode of bacterial IF2, and deviates from the shuttle function of the eukaryotic counterpart eIF2. Thus, despite the resemblance of eIF2 and a/eIF2, recruitment of initiator tRNA to the ribosome is mechanistically different in Pro-and Eukaryotes.

Translation initiation: variations in the mechanism can be anticipated

“…A large-scale analysis of completed prokaryotic genomes showed that a significant proportion of ORFs is leaderless or led by SD free 5 0 -UTRs [51]. These features of 5 0 -UTR are especially dominant in Archaea [52,53]. In some cases of SD-independent translation, the initiation is mediated by ribosomal protein S1 [47,52].…”

Binding of the 5′-Triphosphate End of mRNA to the γ-Subunit of Translation Initiation Factor 2 of the Crenarchaeon Sulfolobus solfataricus

“…The γ-subunit of the crenarchaeon Sulfolobus solfataricus IF2 (SsoIF2γ) serves, besides its central role in Met-tRNA i binding on the ribosome [11], an additional function: SsoIF2γ binds to the 5′-triphosphate end of mRNA and protects its 5′-part from degradation [12]. Similarly, the eukaryotic IF2 has also been shown to interact with mRNA [13,14].…”

Haloferax volcanii, a Prokaryotic Species that Does Not Use the Shine Dalgarno Mechanism for Translation Initiation at 5′-UTRs

“…An in vitro translation system has been established for Sulfolobus solfataricus , which was used to study different aspects of translation initiation [21], [29]–[32]. Notably, the substitution of two nucleotides of the SD motif resulted in a total inhibition of translation, indicating that the SD mechanism is operating in this crenarchaeal species [29].…”