Fig 4.

Rtt109C is required by Asf1 to enhance H3K56ac. (A) Rtt109(1–424) is not efficient in acetylating H3K56 in the presence of Asf1 in vitro. HAT assays were performed as described in the legend of Fig. 3C except that the same series of dilutions of full-length 6×HIS-Rtt109 and 6×HIS-Rtt109(1–424) were incubated with a constant concentration of 6×HIS-Asf1. Reaction products were separated by SDS-PAGE, transferred onto nitrocellulose, and immunoblotted with anti-HIS and anti-H3K56ac. The two boxed images were grouped together from the same SDS-PAGE gel. (B) In vitro, Asf1 and Asf1N but not Vps75 require Rtt109C to enhance the in vitro activity of Rtt109. Serial dilutions of equal amounts of full-length 6×HIS-Rtt109 and 6×HIS-Rtt109(1–424) were incubated with either of 6×HIS-Asf1, 6×HIS-Asf1N, or 6×HIS-Vps75. HAT assays were performed and analyzed as described in the legend of Fig. 3C. The asterisk indicates a probable dimeric form of 6×HIS-Rtt109 and 6×HIS-Rtt109(1–424); the dagger (†)indicates a breakdown product of 6×HIS-Asf1.