Role of forward translocation in nucleoside triphosphate phosphohydrolase I (NPH I)-mediated transcription termination of vaccinia virus early genes.

Termination of transcription of vaccinia virus early genes requires the virion form of the viral RNA polymerase (RNAP), a termination signal (UUUUUNU) in the nascent RNA, vaccinia termination factor, nucleoside triphosphate phosphohydrolase I (NPH I), and ATP. NPH I uses ATP hydrolysis to mediate transcript release, and in vitro, ATPase activity requires single-stranded DNA. NPH I shows sequence similarity with the DEXH-box family of proteins, which includes an Escherichia coli ATP-dependent motor protein, Mfd. Mfd releases transcripts and rescues arrested transcription complexes by moving the transcription elongation complex downstream on the DNA template in the absence of transcription elongation. This mechanism is known as forward translocation. In this study, we demonstrate that NPH I also uses forward translocation to catalyze transcript release from viral RNAP. Moreover, we show that NPH I-mediated release can occur at a stalled RNAP in the absence of vaccinia termination factor and U(5)NU when transcription elongation is prevented.