During
infection by herpes simplex virus type 1 (HSV-1), the virion
protein VP16 activates the transcription of viral immediate-early (IE) genes. Genetic and biochemical assays have shown that the potent transcriptional activation domain of
VP16 can associate with
general transcription factors and with
chromatin-modifying coactivator
proteins of several types. The latter interactions are particularly intriguing because previous reports indicate that HSV-1
DNA does not become nucleosomal during lytic
infection. In the present work, chemical cross-linking and immunoprecipitation assays were used to probe the presence of activators,
general transcription factors, and
chromatin-modifying coactivators at IE gene promoters during
infection of HeLa cells by wild-type HSV-1 and by RP5, a viral strain lacking the
VP16 transcriptional activation domain. The presence of
VP16 and Oct-1 at IE promoters did not depend on the activation domain. In contrast, association of
RNA polymerase II, TATA-binding protein,
histone acetyltransferases (p300 and CBP), and
ATP-dependent remodeling
proteins (BRG1 and hBRM) with IE gene promoters was observed in wild-type
infections but was absent or reduced in cells infected by RP5. In contrast to the previous evidence for nonnucleosomal HSV-1
DNA,
histone H3 was found associated with
viral DNA at early times of
infection. Interestingly,
histone H3 was underrepresented on IE promoters in a manner dependent on the
VP16 activation domain. Thus, the
VP16 activation domain is responsible for recruiting
general transcription factors and coactivators to IE promoters and also for dramatically reducing the association of
histones with those promoters.