Lettuce Big-Vein Associated Virus ORF3 Encodes a Functional 30K Movement Protein

Area of Science:

• Plant virology
• Molecular biology
• Structural biology

Background:

• Movement proteins (MPs) are crucial for plant virus cell-to-cell and systemic movement.
• The 30K superfamily represents the largest group of MPs, characterized by conserved jelly-roll structures despite low sequence similarity.
• Lettuce big-vein associated virus (LBVaV) is linked to severe lettuce big-vein disease (LBVD) and facilitates the spread of other viruses, but its proteins remain uncharacterized.

Purpose of the Study:

• To identify and characterize the movement proteins of Lettuce big-vein associated virus (LBVaV).
• To investigate the role of LBVaV ORF3 in plant virus movement and its potential classification within the 30K superfamily of MPs.

Main Methods:

• Utilized AlphaFold2 for protein structure prediction and FoldSeek for structure-based homology searches to annotate LBVaV open reading frames (ORFs).
• Performed microscopy to determine the subcellular localization of LBVaV ORF3.
• Conducted in planta expression studies to assess the ability of ORF3 to facilitate the movement of movement-impaired viruses.

Main Results:

• All LBVaV ORFs were annotated, with ORF3 identified as belonging to the 30K superfamily of MPs.
• LBVaV ORF3 exhibits low sequence identity (5-11%) to related MPs but shares the characteristic jelly-roll fold.
• Microscopy confirmed ORF3 localization at plasmodesmata, and its expression enhanced cell-to-cell movement of two distinct plant viruses.

Conclusions:

• The AlphaFold2-FoldSeek strategy successfully identified a novel MP (LBVaV ORF3) despite low sequence homology.
• LBVaV ORF3 functions as a movement protein, localizing to plasmodesmata and promoting viral spread.
• This finding contributes to understanding the diversity of MPs and viral movement mechanisms in plants.