Autographa californica Multiple Nucleopolyhedrovirus 38K Is a Novel Nucleocapsid Protein That Interacts with VP1054, VP39, VP80, and Itself

Abstract

It has been shown that the Autographa californica multiple nucleopolyhedrovirus (AcMNPV) 38K (ac98) is required for nucleocapsid assembly. However, the exact role of 38K in nucleocapsid assembly remains unknown. In the present study, we investigated the relationship between 38K and the nucleocapsid. Western blotting using polyclonal antibodies raised against 38K revealed that 38K was expressed in the late phase of infection in AcMNPV-infected Spodoptera frugiperda cells and copurified with budded virus (BV) and occlusionderived virus (ODV). Biochemical fractionation of BV and ODV into the nucleocapsid and envelope components followed by Western blotting showed that 38K was associated with the nucleocapsids. Immunoelectron microscopic analysis revealed that 38K was specifically localized to the nucleocapsids in infected cells and appeared to be distributed over the cylindrical capsid sheath of nucleocapsid. Yeast two-hybrid assays were performed to examine potential interactions between 38K and nine known nucleocapsid shell-associated proteins (PP78/83, PCNA, VP1054, FP25, VLF-1, VP39, BV/ODV-C42, VP80, and P24), three non-nucleocapsid shell-associated proteins (P6.9, PP31, and BV/ODV-E26), and itself. The results revealed that 38K interacted with the nucleocapsid proteins VP1054, VP39, VP80, and 38K itself. These interactions were confirmed by coimmunoprecipitation assays in vivo. These data demonstrate that 38K is a novel nucleocapsid protein and provide a rationale for why 38K is essential for nucleocapsid assembly.

Autographa californica Multiple Nucleopolyhedrovirus ORF11 Is Essential for Budded-Virus Production and Occlusion-Derived-Virus Envelopment

“…ac93 encodes a BV-and ODV-associated nucleocapsid protein that is present in the BV envelope fraction (33). In addition, ac9 (p78/83) (34), ac54 (vp1054) (35), ac77 (vlf-1) (36), ac98 (38k) (20), ac142 (37), and ac146 (38), which are essential for BV production, have been shown to encode nucleocapsid proteins located in both BV and ODV. Interestingly, all of these genes are not essential for …”

“…The resulting supernatant was used for BV purification, and the pellet was used for ODV purification. The purification of BVs and ODVs and the fractionation of the envelope and nucleocapsid of BV and ODV were performed as previously described (7,20).…”

Bombyx mori nucleopolyhedrovirus BmP95 plays an essential role in budded virus production and nucleocapsid assembly

“…It has been proposed that P6.9 (Wang et al, 2010a), VP39 (Thiem & Miller, 1989), VP1054 (Olszewski & Miller, 1997), P78/83 , VLF-1 (Vanarsdall et al, 2006), FP25 (Braunagel et al, 1999(Braunagel et al, , 2001, BV/ODV-C42 (Braunagel et al, 2001), ODV-EC27 , vp80 (Marek et al, 2011), P24 (Wolgamot et al, 1993), AC141 (EXON0; Fang et al, 2007), AC142 (McCarthy et al, 2008) and AC98 (38K; Wu et al, 2008) are associated with BV and ODV nucleocapsids. P95, a homologue of Ac83 of Autographa californica multicapsid nucleopolyhedrovirus (AcMNPV), is a virion capsid protein of both BV and ODV and was originally characterized as VP91 in Orgyia pseudotsugata MNPV (OpMNPV; .…”

Autographa californicaNucleopolyhedrovirus Ac76: a Dimeric Type II Integral Membrane Protein That Contains an Inner Nuclear Membrane-Sorting Motif

“…Samples were infiltrated with 40, 70, and 100% LR White-ethanol series at Ϫ20°C for 1 h per step, followed by infiltration with 100% LR White at Ϫ20°C overnight. The resin was then polymerized in gelatin capsules by UV irradiation (320 nm) at Ϫ20°C for 72 h and at room temperature (RT) for 24 h. Ultrathin sections were immunolabeled and stained as previously described (22). Immunolabeling was performed with a mouse monoclonal anti-HA antibody (1:50; Abmart) or mouse monoclonal anti-GFP antibody (1:50; Abmart), followed by incubation with goat anti-mouse IgG conjugated to 10-nm gold particles as the secondary antibody (1:50; Sigma).…”