Molecular proof that bacteriophage T4 alc and unf genes are the same gene

Abstract

The DNA of bacteriophage T4 normally has a substituted base, hydroxymethylcytosine, instead of the usual cytosine. The bacteriophage shuts off host transcription after infection presumably by specifically blocking transcription of cytosine DNA. If T4 incorporates cytosine into its own DNA, this shutoff mechanism is directed back at itself and blocks its own transcription. Mutations which overcome this transcriptional block are in the T4 alc gene, and alc mutations allow the propagation of T4 with cytosine in their DNA (L. Snyder, L. Gold, and E. Kutter, Proc. Natl. Acad. Sci. USA 73:3098-3102, 1976). By genetic criteria, alc is the same as another gene, unf, whose product is required for the unfolding of the bacterial nucleoid after infection (K. Sirotkin, J. Wei, and L. Snyder, Nature [London] 265:28-32, 1977; D. P. Snustad, M. A. Tigges, K. A. Parson, C. J. H. Bursch, F. M. Caron, J. F. Koerner, and D. J. Tutas, J. Virol. 17:622-641, 1976; M. Tigges, C. J. H. Bursch, and D. P. Snustad, J. Virol. 24:775-785, 1977). The product of the alc gene has been identified as a 19-kilodalton protein (R. E. Herman, N. Haas, and D. P. Snustad, Genetics 108:305-317, 1984; E. Kutter, R. Drivdahl, and K. Rand, Genetics 108:291-304, 1984), and an open reading frame has been proposed to be the alc gene based on its size and map position (E. Kutter, R. Drivdahl, and K. Rand, Genetics 108:291-304, 1984). We used marker rescue techniques and DNA sequencing to confirm that this open reading frame is the alc gene. We also present a molecular proof that alc and unf are the same gene. While these results do not rigorously exclude the possibility that Unf and Alc are different activities of the same protein, they strongly support the conclusion that the unfolding of the bacterial nucleoid the blockage of transcription are but different manifestations of the same activity.