Factor-Dependent Internal Ribosome Entry Site and -1 Programmed Frameshifting Signal in the Bemisia-Associated...

Area of Science:

• Molecular Biology
• Virology
• RNA Structure and Function

Background:

• Dicistrovirus intergenic (IGR) internal ribosome entry sites (IRESs) typically employ a streamlined, factor-independent translation initiation mechanism.
• These IRESs usually possess two to three overlapping pseudoknots that interact with ribosomal subunits.
• A previous study predicted an atypical IGR IRES and a -1 programmed frameshift (-1 FS) signal in whitefly Bemisia-associated dicistrovirus 2 (BaDV-2).

Purpose of the Study:

• To investigate the functional characteristics of the predicted BaDV-2 -1 FS signal and IGR IRES.
• To elucidate the mechanism of translation initiation employed by BaDV-2.
• To characterize the RNA structural elements involved in BaDV-2 translation regulation.

Main Methods:

• Utilized bicistronic reporters to assess the activity of the BaDV-2 -1 FS signal and IGR IRES in vitro.
• Employed deletion and mutational analyses to map the IRES element and identify key structural features.
• Investigated ribosome binding and sensitivity to translation initiation factor inhibitors (eIF2, eIF4A).

Main Results:

• The BaDV-2 -1 FS signal drives frameshifting in vitro, mediated by a slippery sequence and a downstream stem-loop.
• The BaDV-2 IGR IRES supports translation initiation from an AUG codon but requires host factors, unlike typical dicistrovirus IRESs.
• The BaDV-2 IRES is sensitive to eIF2 and eIF4A inhibitors and appears unstructured or adopts dynamic conformations during initiation.

Conclusions:

• Demonstrated the first -1 FS frameshifting signal in the dicistrovirus family.
• Identified a novel, factor-dependent IRES mechanism within dicistroviruses, challenging previous models.
• Highlighted the diverse strategies viral RNA structures employ for regulating protein synthesis.