The three immediate-early proteins of HSV-1, ICPO, ICP22, and ICP27, have specific and pivotal functions in transcriptional activation and inhibition, multiple regulatory and control processes of viral genes. In this paper, the expression and localization of these three proteins were studied in neuroblastoma cells using biochemical assays, and their possible and potential interactive functions are discussed. The data show that the three proteins are localized in different structures, specifically in the PML-NB-associated structure, which is a specific nuclear structure composed of many protein molecules and bound tightly to the nuclear matrix in neuroblastoma cells. The results suggest that the activating and suppressive functions of ICPs are mostly dependent on their transcriptional and regulatory roles, including the PML-NB-associated structure.