FIG. 1.

pRb, but not p53, inhibits the replication of Py T Ag in vitro. (A) Increasing concentrations of pRb were added to replication mixtures containing FM3A extract (300 μg), Py ori-DNA (0.2 μg), and Py T Ag (0.6 μg). After a 3-h incubation at 33°C, 5-μl aliquots were acid precipitated and counted by scintillation. A background of 0.5 pmol has been subtracted. (B) DNA replication products were purified from the replication mixtures, linearized, digested with DpnI, and analyzed on 1% agarose gels followed by autoradiography. Linear DNA (form I) is indicated on the left. (C) Replication mixtures containing FM3A extract (300 μg), Py ori-DNA (0.2 μg), and Py T Ag (0.6 μg) received increasing concentrations of pRb or p53 purified from insect cells or from bacteria (GST-Rb, GST-p53, or GST). pRb¹ and pRb³ are different preparations of pRb. 1 (0.5:1), 2 (1:1), and 3 (2:1) denote the ratios of pRb or p53 to Py T Ag. After a 3-h incubation at 33°C, the DNA products were analyzed as described for panel B. Form I and form II DNAs are indicated on the left.