Systematic analysis of nuclear localization of Autographa californica multiple nucleopolyhedrovirus proteins

Abstract

Baculoviruses are large DNA viruses that replicate within the nucleus of infected host cells. Therefore, many viral proteins must gain access to the nucleus for efficient viral genome replication, gene transcription and virion assembly. To date, the global protein localization pattern of baculoviral proteins is unknown. In this study, we systematically analysed the nuclear localization of 154 ORFs encoded by the prototypic baculovirus, Autographa californica multiple nucleopolyhedrovirus (AcMNPV), either during transient expression or with super-infection of the virus. By transient expression of vectors containing egfp-fused ORFs, we found that in the absence of virus infection, 25 viral proteins were localized in the nucleus. Most of these, which we called ‘auto-nuclear localization’ proteins, are related to virus replication, transcription or virion structure, and 20 of them contain predicted classical nuclear localization signal. Upon virus infection, 11 proteins, which originally localized in the cytoplasm or both cytoplasm and nucleus in the transfection assays, were completely translocated into the nucleus, suggesting that their nuclear import is facilitated by other viral or host proteins. Further co-transfection experiments identified that four of the 11 proteins, including P143, P33, AC73 and AC114, were imported into the nucleus with the assistance of the auto-nuclear localization proteins LEF-3 (for P143), TLP (for P33) and VP80 (for both AC73 and AC114). This study presents the first global nuclear localization profile of AcMNPV proteins and provides useful information for further elucidation of the mechanisms of baculovirus nuclear entry and gene functions.

Identification of the Key Functional Domains of Bombyx mori Nucleopolyhedrovirus IE1 Protein

“…He et al showed that 25 viral proteins were localized in the nucleus by transient transfection experiments in the absence of AcMNPV infection. Most of these proteins are involved in viral DNA replication, transcription, and virion structure, and 20 of them contain a predicted classical nuclear localization signal [ 25 ]. IE1 acts as an immediate early protein and major transcriptional activator, and nuclear localization signal is required for its function.…”

“…Moreover, BmNPV IE1 539–559 as a secondary nuclear localization element includes the two arginines, which may explain why the absence of IE1 539–559 weakened the nuclear localization of BmNPV IE1; however, these amino acids are not requisites for the nuclear import of BmNPV IE1. AcIE1 is predicted to have a bipartite nuclear localization signal at residues 130–151 [ 25 ], which may account for the ability of BmNPV IE1 1–157 to partially enter the nucleus. Although we identified three nuclear localization elements of BmNPV IE1, further truncations and point mutations need to be performed to identify the key amino acids that perform nuclear localization.…”

A SLiM-dependent conformational change in baculovirus IE1 controls its focus formation ability

“…The original BDI (Basic domain I; residues 152 to 161 in AcMNPV) is a highly conserved domain required for IE1 binding to the hr enhancer element as well as hr-dependent transactivation and focus formation (6,7). BDI was recently predicted (8,9) to comprise a bipartite NLS (Fig. 1A).…”

“…BDI [Basic domain I; residues 152 to 161 in the Autographa californica multiple nucleopolyhedrovirus; (AcMNPV)] was originally identified as a highly conserved domain required for IE1 interactions with the hr enhancer element and subsequent formation of an hr -dependent focus (6, 7). A recent report using a nuclear localization signal (NLS) prediction algorithm identified a putative bipartite NLS in BDI (8, 9), which we propose to subdivide into BDIa and BDIb with the original BDI re-defined BDIb as shown in Figure 1A. Additional stretches of basic amino acids encompassing the C termini of BDII (residues 521 to 538 in AcMNPV) and HLH (Helix-loop-helix dimerization motif; residues 543 to 568 in AcMNPV) play roles in DNA binding and nuclear import (10, 11).…”

Identification and characterization of BmNPV Bm5 protein required for the formation of nuclear vesicle structures

“…The relationships between viral proteins, small NMV formation and nucleocytoplasmic transport have been established extensively, however, the molecular mechanism of large NMV formation is poorly understood. Recently, several studies indicated that Bombyx mori nucleopolyhedrovirus (BmNPV) Bm5 protein and its homologue Ac13 are shown to concentrate on both INM and ONM [21][22][23]. Deletion of Bm5 and Ac13 results in fewer BV and ODV productions as well as reducing the accumulation of lipids in the nucleus [22][23][24].…”