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Autographa californica Multiple Nucleopolyhedrovirus Nucleocapsid Protein BV/ODV-C42 Mediates the Nuclear Entry of P78/83

State Key Laboratory of Virology, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan 430071, People's Republic of China,¹ Graduate University of the Chinese Academy of Sciences, Beijing 100039, People's Republic of China²

Received 11 November 2007/ Accepted 11 February 2008

Autographa californica multiple nucleopolyhedrovirus (AcMNPV) BV/ODV-c42 (orf101; c42), which encodes a 41.5-kDa viral nucleocapsid protein with a putative nuclear localization signal (NLS) motif at the C terminus, is a highly conserved gene among members of the Baculoviridae family. C42 is demonstrated to be essential for AcMNPV propagation and can bind to nucleocapsid protein P78/83, a viral activator for the actin-related protein 2/3 (ARP2/3) complex to initiate nuclear actin polymerization, which is essential for viral nucleocapsid morphogenesis during AcMNPV infection. Here, we report the identification of a novel pathway through which c42 functions in nucleocapsid assembly. Cotransfection of Sf9 cells with c42 and p78/83 plasmids demonstrated that C42 was capable of recruiting P78/83 to the nuclei of uninfected cells and that the NLS motif of C42 was essential for this process. To validate this nuclear relocation mode in bacmid-transfected cells, a c42-disrupted bacmid (vAc^c42ko-gfp) and rescued bacmids with wild-type c42 (vAc^c42res-gfp) or with NLS coding sequence-mutated c42 (vAc^c42nls-gfp) were prepared. By immuno-staining, P78/83 was found to be localized in the cytoplasm of either vAc^c42ko-gfp- or vAc^c42nls-gfp-transfected cells, whereas P78/83 was relocated to the nuclei of vAc^c42res-gfp-transfected cells. Furthermore, F-actin-specific staining confirmed that there was no actin polymerization activity in the nuclei of either vAc^c42ko-gfp- or vAc^c42nls-gfp-transfected cells, which might be attributed to the absence of nuclear P78/83, an activator of the ARP2/3 complex to initiate nuclear actin polymerization. We therefore hypothesize a mode of action where C42 binds to P78/83 in the cytoplasm to form a protein complex and cotransports to the nucleus under the direction of the NLS motif in C42 during AcMNPV infection.

Published ahead of print on 20 February 2008.

^# Present address: Department of Biochemistry & Molecular Biology, Nanjing Medical University, Nanjing, China.