The baculovirus Autographa californica nucleopolyhedrovirus (AcNPV) has two viral forms, budded virus (BV) and occlusion derived virus (ODV). The envelopment of these two viral forms occurs at different locations: BV acquires envelopes at the plasma membrane while ODV acquires envelopes in the nucleus. The two viral forms carry out different functions in the viral life cycle. The purpose of this study is to investigate how viral envelope proteins sort/traffic to the nucleus. Of particular interest is BV/ODV E26 (E26). E26 is an envelope protein of both BV and ODV (Braunagel and Summers, 1994); therefore it must traffic to the plasma membrane and the nucleus during infection. Thus, E26 is a bi-directional trafficking protein, which interacts with membranes in both locations of the cell. As such it has been shown that there are several immunoreactive forms of E26 (Beniya, Braunagel, and Summers, 1998). The da26 gene produces at least 2 protein products of 26 and 28 kDa with different functions respectively, which correlate with localization, solubility, membrane association, and temporal requirements. The 28 kDa form is likely a soluble protein that interacts with transcriptional activators and DNA in the nucleus in the early stages of infection. A part of the 26 kDa population is a membrane bound form interacting with an integral membrane protein in the ER and likely functions as an INM sorting factor. The 26 kDa membrane bond form is also found in the inner nuclear membrane, intra-nuclear microvesicles, ODV envelopes, and ODV in the nucleus.

The amino-terminal 33 amino acids of the baculovirus integral membrane protein, ODV-E66, are sufficient for localization of fusion proteins to viralinduced intranuclear microvesicles (MV) and occlusion derived virus envelopes during infection, and has been termed the sorting motif (SM). When abundantly expressed, SM-fusions are also detected in the inner nuclear membrane (INM), outer nuclear membrane and endoplasmic reticulum of infected cells, suggesting proteins with the SM use the same trafficking pathway as cellular INM proteins to traffic to nuclear membranes. This study identifies the essential characteristics required for sorting of the SM to the INM of uninfected cells, and the MV and ODV envelopes of infected cells. These features are an 18 amino acid transmembrane sequence that lacks polar and charged amino acids (a.a.) with a cluster of charged a.a. spaced 5-11 residues from the end of the transmembrane sequence. A comparison of the a.a. sequence of these SM features with cellular INM proteins shows the features are conserved. The model of INM protein sorting and localization predicts the only known sorting event during INM protein trafficking is immobilization/retention in the INM. This study uses confocal microscopy and fluorescence recovery after photobleaching to compare the localization and mobility of lamin B receptor (LBR) fusions (which contain SM-like sequences) to a viral SM fusion when expressed in either mammalian or insect cells. The results show that immobilization is not necessarily required for accumulation of proteins in the INM. Furthermore, the results from infected cells show that an active sorting event, likely independent of immobilization, can distinguish the viral SM from cellular sequences similar to the SM. The results of this study show that sorting of proteins to the INM can be mediated by the viral SM or INM protein SM-like sequences that can function either independent of, or in addition to, immobilization. These data combined with recent reports suggest that in addition to diffusion:retention a signal mediated mechanism for sorting and localization to the INM can occur.