Assembly of herpes simplex virus replication proteins at two distinct intranuclear sites

Herpes simplex virus DNA replication proteins amplify the viral genome in large globular replication compartments within infected cell nuclei. In the absence of viral DNA synthesis, the replication proteins accumulate at punctate foci throughout the nucleus referred to as prereplicative sites. To more thoroughly understand the nature of this nuclear assembly process, we have examined the viral and cellular factors involved. First, we demonstrate that six viral replication proteins are sufficient for formation of functional replication compartments in transfected cells in the absence of viral origin-containing DNA. Second, we show that the viral replication proteins form two distinct types of prereplicative sites within infected cells. One type of punctate structure assembles in S-phase cells, colocalizes with cellular DNA synthesis, and contains components of the host-cell replication apparatus as indicated by the presence of Replication Protein A. However, the other class of prereplicative sites is independent of host-cell DNA synthesis as evidenced by their formation in cells arrested in G1 by n-butyrate. These complexes are significantly less abundant and closely correspond with cellular Nuclear Domain 10 structures to which viral DNA has recently been demonstrated to be targeted early in infection (G. G. Maul, A. M. Ishov, and R.D. Everett, 1996, Virology 217, 67-75). Hence, this second type appears to represent the subset of prereplicative sites destined to become replication compartments.