|Chavez,, Juan -|
|Ju,, Ho-Jong -|
|Weisbrod,, Chad -|
|Eng,, Jimmy -|
|Bruce,, James -|
Submitted to: Journal of Proteome Research
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: March 6, 2012
Publication Date: May 4, 2012
Citation: Chavez, J., Cilia, M., Ju, H., Weisbrod, C., Eng, J.K., Gray, S.M., Bruce, J.E. 2012. Cross-linking measurements of the potato leafroll virus reveal protein interaction topologies required for virion stability, aphid transmission, and virus-plant interactions. Journal of Proteome Research. 11(5):2968-2981. Interpretive Summary: Insect-transmitted viruses threaten animal and plant health on a global scale. Potato leafroll virus (PLRV) is transmitted by aphids, is part of a larger family of viruses that infect potato, wheat, oats, barley, and rye, and causes serious economic problems for farmers in the United States and world-wide. Currently there is limited host resistance to this group of viruses and no effective control measures for aphid vectors. There is also no structural information on the capsids of these viruses that could facilitate an understanding of how the viruses interact with insect vectors and plant hosts. Here we used a novel chemical cross-linking reagent called Protein Interaction Reporter to study the architecture of the PLRV capsid. The Protein Interaction Reporter links proteins together and these links enable us to visualize the surfaces and surface features of proteins. The Protein Interaction Reporter enables measurements of these protein links using a mass spectrometer. The Protein Interaction Reporter enabled us to discover the regions of the PLRV particles that are involved in stabilizing the cage-like icosahedral structure of the capsid and contributing to aphid-transmission and virus interaction with host plants. The Protein Interaction Reporter is particularly useful to explore virus particle structures that are difficult to study using other methods and to understand which regions of the capsid are involved in biological function.
Technical Abstract: Protein interactions are critical determinants of insect-transmission for viruses in the family Luteoviridae. Two luteovirid structural proteins, the capsid protein (CP) and the readthrough protein (RTP), contain multiple functional domains that regulate virus transmission. There is no structural information available for these economically important viruses. We used Protein Interaction Reporter (PIR) technology, a strategy that uses chemical cross-linking and high resolution mass spectrometry, to discover topological features of the Potato leafroll virus (PLRV) CP and RTP that are required for the diverse biological functions of PLRV virions. Four cross-linked sites were repeatedly detected, one linking CP monomers, two within the RTP, and one linking the RTP and CP. Virus mutants with triple amino acid deletions immediately adjacent to or encompassing the cross-linked sites were defective in virion stability, RTP incorporation into the capsid, and aphid transmission. Plants infected with a new, infectious PLRV mutant lacking 26 amino acids encompassing a cross-linked site in the RTP exhibited a delay in the appearance of systemic infection symptoms. PIR technology provided the first structural insights into luteoviruses which are crucially lacking and that are involved in vector-virus and plant-virus interactions. These are the first cross-linking measurements on any infectious, insect-transmitted virus.