The initiation and termination of vaccinia virus postreplicative transcription have been reported to require cellular proteins, some of which are believed to be nuclear proteins. Vaccinia virus replicates in the cytoplasmic compartment of the cell, raising questions as to whether vaccinia virus has access to nuclear proteins. This was addressed here by following the fate of several nuclear proteins after infection of cells with vaccinia virus. The nuclear transcription factors YY1, SP1, and TATA binding protein were found to colocalize with virus replication complexes in the cytoplasm of infected cells. In addition, the nuclear proteins RNA polymerase II, TAFIIp32, and histone deacetylase 8, but not the structural protein lamin B, also were found in the cytoplasm of the cell. The association of YY1 with replication complexes was dependent on DNA replication and required only the DNA binding domain of the protein, indicating that DNA binding alone may be responsible for the association of nuclear transcription factors with viral replication complexes in the cytoplasm. The cytoplasmic localization of YY1 was resistant to the nuclear export inhibitor leptomycin B. Evidence is presented indicating that nuclear import and export pathways were not adversely affected by vaccinia virus infection. These observations indicate that vaccinia virus replication complexes have ready access to nuclear proteins by allowing leakage from the nucleus.