ROLE OF BACULOVIRUS IE2 AND ITS RING FINGER IN CELL-CYCLE ARREST

The ie2 gene of Autographa californica nuclear polyhedrosis virus (AcM NPV) is known to transactivate transient expression from viral promote rs in a host cell-specific manner. We report that transfection of Spod optera frugiperda (SF-21) cells with ie2 was sufficient to arrest the cell cycle, resulting in the accumulation of enlarged cells with abnor mally high DNA contents. By 72 h posttransfection, more than 50% of ie 2-transfected cells had DNA contents greater than 4N. There was no evi dence of mitotic spindle formation in these cells, and expression of i e2 appeared to block cell cycle progression in S phase. Several ie2 mu tants were analyzed to further define the region of IE2 responsible fo r arresting the cell cycle. Analysis of these mutants showed that dele tion of the RING finger motif eliminated the ability of IE2 to arrest the cell cycle but did not affect its ability to transactivate the ie1 promoter. Moreover, mutation of a single conserved cysteine (C251) of the RING finger motif abolished the ability of IE2 to block cell cycl e progression but had no apparent effect on its transregulatory activi ty. In contrast, a mutant of IE2 containing a deletion of residues 94 to 173 was able to block cell division but lacked trans-regulatory act ivity. Thus, the ability of IE2 to arrest the cell cycle depended on t he integrity of the RING finger motif and was distinct from and indepe ndent of its ability to trans-activate the ie1 promoter. IE2 also arre sted the division of cells derived from other insect species, Trichopl usia ni (TN-368 and BTI-TN-5B1-4) and Helicoverpa zea (Hz-AM1).