Proteomics of the Autographa californica nucleopolyhedrovirus budded virions

Abstract

Baculoviruses produce two progeny phenotypes during their replication cycles. The occlusion-derived virus (ODV) is responsible for initiating primary infection in the larval midgut, and the budded virus (BV) phenotype is responsible for the secondary infection. The proteomics of several baculovirus ODVs have been revealed, but so far, no extensive analysis of BV-associated proteins has been conducted. In this study, the protein composition of the BV of Autographa californica nucleopolyhedrovirus (AcMNPV), the type species of baculoviruses, was analyzed by various mass spectrometry (MS) techniques, including liquid chromatography-triple quadrupole linear ion trap (LC-Qtrap), liquid chromatography-quadrupole time of flight (LC-Q-TOF), and matrix-assisted laser desorption ionization-time of flight (MALDI-TOF). SDS-PAGE and MALDI-TOF analyses showed that the three most abundant proteins of the AcMNPV BV were GP64, VP39, and P6.9. A total of 34 viral proteins associated with the AcMNPV BV were identified by the indicated methods. Thirteen of these proteins, PP31, AC58/59, AC66, IAP-2, AC73, AC74, AC114, AC124, chitinase, polyhedron envelope protein (PEP), AC132, ODV-E18, and ODV-E56, were identified for the first time to be BV-associated proteins. Western blot analyses showed that ODV-E18 and ODV-E25, which were previously thought to be ODV-specific proteins, were also present in the envelop fraction of BV. In addition, 11 cellular proteins were found to be associated with the AcMNPV BV by both LC-Qtrap and LC-Q-TOF analyses. Interestingly, seven of these proteins were also identified in other enveloped viruses, suggesting that many enveloped viruses may commonly utilize certain conserved cellular pathways.

FIG. 1.

SDS-PAGE profile of purified AcMNPV budded virions and samples for MS analysis. M1 to M3 indicate the bands analyzed by MALDI-TOF, and L1 to L14 indicate the bands analyzed by LC-Qtrap.

FIG. 2.

Western blot analyses of AcMNPV ODV/BV nucleocapsid (NC) and envelope (E) fractions with anti-ODV-E25 and anti-ODV-E18 antibodies. Healthy cells (H) and virus-infected cells (I) were loaded as negative and positive controls, respectively. GP64, VP39, and ODV-E66 were detected by their respective antibodies to illustrate the BV E-specific, NC-specific, and ODV E-specific proteins, respectively. M indicates molecular mass markers.